CN1715413A

CN1715413A - Corynebacterium glutamicum genes encoding metabolic pathway proteins

Info

Publication number: CN1715413A
Application number: CNA2005100759145A
Authority: CN
Inventors: M·波姆佩朱斯; B·克雷格尔; H·施雷德; O·策尔德; G·哈贝豪尔
Original assignee: BASF SE
Current assignee: BASF SE
Priority date: 1999-06-25
Filing date: 2000-06-23
Publication date: 2006-01-04
Also published as: CN1800397A; ZA200200645B; CN1766111A; CN101078016A; ZA200200646B; CN1807634A; CN101078017A; CN101082049A; ZA200200585B; CN101074441A; CN1962870A; CN1920040A; ZA200200584B; CN1990868A; ZA200200647B; CN101082050A

Abstract

The invention describes the MP protein nuclei acid molecular of new strain corynebacterium of separated code. The molecular is called MP nucleic acid molecular. The invention also provides an anti sense nucleic acid molecular, a recombinant expression vector containing the MP nucleic acid molecular and a host cell that has been introduced into the vector. The invention further provides the separated MP protein, MP mutant protein, fusion protein and antigen peptide and a method for manufacturing a required compound, which is processed by an organism base on the improvement of MP gene genetic engineering of the nuclei acid molecular.

Description

Encoding metabolic pathway gene protein of Corynebacterium glutamicum

This application is filed on June 23, 2000 the Chinese Patent Application 00811981.3 "Program Code metabolic pathways of Corynebacterium glutamicum protein gene "divisional application.

RELATED APPLICATION

This application claims the priority of the following apply: prior filed U.S. Provisional Application Serial No. 60/141 031, filing date June 25, 1999, U.S. Provisional Application Serial No. 60/142101, the applicant Please date July 2, 1999, U.S. Provisional Application Serial No. 60/148613, the filing date of August 1999 On 12 May, and U.S. Provisional Application Serial No. 60/187970, filing date March 9, 2000. This application requires the following priority application: filed earlier German Patent Application No. 19930476.9, filing date July 1,1999, German Patent Application No. 19931415.2, filing date July 8, 1999, the German Patent Application No. 19931418.7, filing date July 8, 1999, German Patent Application No. 19931419.5, filing date July 8, 1999, the German Patent Application No. 19931420.9, filing date July 8, 1999, the German Patent Application No. 19931424.1, apply Day 8 July 1999, the German Patent Application No. 19931428.4, filing date July 8, 1999 , The German Patent Application No. 19931434.9, filing date July 8, 1999, the German Patent Application Please No. 19931435.7, filing date July 8, 1999, the German Patent Application No. 19931443.8, Filing date July 8, 1999, the German Patent Application No. 19931453.5, filing date July 1999 On 8 May, the German Patent Application No. 19931457.8, filing date July 8, 1999, the German expert Patent Application No. 19931465.9, filing date July 8, 1999, the German Patent Application No. 19931478.0, filing date July 8, 1999, the German Patent Application No. 19931510.8, apply Day 8 July 1999, the German Patent Application No. 19931541.8, filing date July 8, 1999 , The German Patent Application No. 19931573.6, filing date July 8, 1999, the German Patent Application Please No. 19931592.2, filing date July 8, 1999, the German Patent Application No. 19931632.5, Filing date July 8, 1999, the German Patent Application No. 19931634.1, filing date July 1999 On 8 May, the German Patent Application No. 19931636.8, filing date July 8, 1999, the German expert Patent Application No. 19932125.6, filing date July 9, 1999, the German Patent Application No. 19932126.4, filing date July 9, 1999, the German Patent Application No. 19932130.2, apply Day 9 July 1999, the German Patent Application No. 19932186.8, filing date July 9, 1999 , The German Patent Application No. 19932206.6, filing date July 9, 1999, the German Patent Application Please No. 19932227.9, filing date July 9, 1999, the German Patent Application No. 19932228.7, Filing date July 9, 1999, the German Patent Application No. 19932229.5, filing date July 1999 On 9 May, German Patent Application No. 19932230.9, filing date July 9, 1999, the German expert Patent Application No. 19932922.2, filing date July 14, 1999 German Patent Application No. 19932926.5, filing date July 14, 1999, German Patent Application No. 19932928.1, Shen Please date July 14,1999, German Patent Application No. 19933004.2, filing date July 14, 1999 , The German Patent Application No. 19933005.0, filing date July 14, 1999, the German Patent Application Please No. 19933006.9, filing date July 14, 1999, German Patent Application No. 19940764.9, Filing Date August 27, 1999, German Patent Application No. 19940765.7, filing date August 1999 On 27 May, the German Patent Application No. 19940766.5, filing date August 27, 1999, Germany Patent Application No. 19940832.7, filing date August 27, 1999, German Patent Application No. 19941378.9, filing date August 31, 1999, German Patent Application No. 19941379.7, Shen Please date August 31, 1999, German Patent Application No. 19941380.0, filing date August 1999 On 31 May, the German Patent Application No. 19941394.0, filing date August 31, 1999, Germany Patent Application No. 19941396.7, filing date August 31, 1999, German Patent Application No. 19942076.9, filing date September 3, 1999, the German Patent Application No. 19942077.7, apply Day 3 September 1999, the German Patent Application No. 19942079.3, filing date September 03, 1999 , The German Patent Application No. 19942086.6, filing date September 3, 1999, the German Patent Application Please No. 19942087.4, filing date September 3, 1999, the German Patent Application No. 19942088.2, Filing date September 3, 1999, the German Patent Application No. 19942095.5, filing date September 1999 On 3 May, German Patent Application No. 19942124.2, filing date September 3, 1999, and Germany Patent Application No. 19942129.3, filing date September 3, 1999. Entire contents of the above application Incorporated herein by reference. ...

The present invention relates to an isolated Corynebacterium glutamicum coding for a novel protein nucleic acid molecule MP, the MP nucleic acid molecule molecule is known. The present invention also relates to an antisense nucleic acid molecule comprising a nucleic MP The recombinant expression vector molecule, and has been introduced into an expression vector host cells. The present invention also further MP step relates to an isolated protein, MP muteins, fusion proteins, antigenic peptides, and on the Corynebacterium glutamicum MP genetic engineering to improve the organism by the production of the desired compound Approach.

BACKGROUND

Naturally present in cell metabolism of the specific products and by-products in many industries With uses including food, feed, cosmetics and pharmaceutical industries. These molecules are referred to as "fine Fine chemical ", include an organic acid, a source of protein and non-protein sources of amino acids, nucleotides and Nucleosides, lipids and fatty acids, diols, carbohydrates, aromatic compounds, vitamins and cofactors in And enzymes. Can produce and secrete a large number of large-scale cultivation required specific molecules of bacteria, most parties The preparation of these products will be. For this purpose is a particularly useful organisms glutamate Corynebacterium (Corynebacterium glutamicum), a Gram-positive non-pathogenic bacteria. Through Strains over, a number of generating a large number of mutants of the desired compound. However, to improve the Particular molecule carried Strains production, is a time-consuming and difficult process. ...

The present invention provides a new bacterial nucleic acid molecules that have a variety of uses. These use Uses include identification can produce fine chemicals microorganisms, regulation or pro Corynebacterium glutamicum Edge in bacteria fine chemical substances, Corynebacterium glutamicum or genetic typing of bacteria and Identified as Corynebacterium glutamicum drawing reference point genome. These new nucleic acid molecules Encoding a protein, referred to herein as metabolic pathway (MP) proteins.

The present invention provides a new bacterial nucleic acid molecules that have a variety of uses. These use Uses include identification can produce fine chemicals microorganisms, regulation or pro Corynebacterium glutamicum Edge in bacteria fine chemical substances, Corynebacterium glutamicum or genetic typing of bacteria and Identified as Corynebacterium glutamicum drawing reference point genome. These new nucleic acid molecules Encoding a protein, referred to herein as metabolic pathway (MP) proteins....

MP nucleic acid molecule of the present invention is a microorganism that can be used to identify whether or Corynebacterium glutamicum By its genetic strains, or identification of mixed microbial populations glutamicum bacteria or their relatives Strain exists. The present invention provides a number of the gene of Corynebacterium glutamicum nucleic acid sequence; the stringency Conditions, the probe with a probe from a single group of microorganisms, or mixed microbial culture of the base was extracted Genomic DNA, this probe covers a Corynebacterium glutamicum Ji Yinte some regions can be determined Whether the organism exists. Although Corynebacterium glutamicum itself is non-pathogenic, but it Pathogens in humans related species, such as Corynebacterium diphtheria (Corynebacterium diphtheriae) (Diphtheria pathogen); detect this microorganism has significant clinical utility. ...

MP nucleic acid molecule of the present invention can be used as draw Corynebacterium glutamicum genome parameters According to point, or draw its genetic strain genome reference point. Similarly, these molecules, Or a variant or portion thereof, can be used as genetic engineering of Corynebacterium or Brevibacterium genetic markers Remember. For example, the present invention is a novel nucleic acid molecule encoding the protein can be certain MP Fine Chemicals Qualitative enzymatic metabolism step, said fine chemicals including amino acids, vitamins, cofactors, Nutritional factors (nutraceutical), nucleotides, nucleosides, and trehalose. Taking into account that in C. glutamicum Coli cloning vector used in practicality, for example, in Sinskey et al., U.S. Patent No. 4,649,119 in the open, and taking into account genetic Corynebacterium glutamicum and Brevibacterium species (eg Brevibacterium lactofermentum) genetic manipulation techniques (Yashihama et al, J.Bacteriol.162: 591 - 597 (1985); Katsumata et al., J.Bacteriol.159 :306-311 (1984); and Santamaria et al., J.Gen.Microbiol.130 :2237-2246 (1984)), the present invention is a nuclear Acid molecules can be used for the genetic engineering of living organisms, to make it one or more fine chemicals Better quality, or more efficient producers. ...

Fine chemicals can be produced to improve or effective operation of the present invention, the direct effect of gene Operation or indirect effect of this gene. Specifically, Corynebacterium glutamicum amino acids, vitamins, Cofactors, nucleotide changes in metabolic pathways and trehalose biological production of such one or more of these These compounds have a direct effect desired. For example, optimizing the lysine biosynthetic pathway protein Activity Or lower lysine breakdown pathway protein activity can lead to the production of this bio-engineered Lysine yield or efficiency. These changes in metabolic pathway proteins will also need refinement Chemicals production or efficiency have an indirect effect. For example, to produce the desired intermediate molecular competition The reaction may be eliminated contention, or to produce the desired intermediate compound of specific ways to Be optimized. In addition, amino acids, vitamins, or nucleotide biosynthesis or degradation can be increased regulation Increase the production and the ability to divide, thereby increasing the number of microorganisms in the culture and / or production capacity, And increase the possibility of the desired fine chemical production. ...

The nucleic acid and protein of the present invention can be used to directly improve the molecule Corynebacterium glutamicum of one or More desired fine chemicals production or productivity. Using recombinant techniques known in the art Surgery, one or more of the present invention, amino acids, vitamins, cofactors, nutritional factors, nucleotides, Nucleosides or trehalose biosynthetic enzyme may be changed or degradation, thereby regulating their functions. For example, Biosynthetic enzymes can increase the efficiency or destruction of its allosteric control region so as to prevent production of the compound Feedback inhibition. Similarly, a degradative enzyme can be substitution, deletion or increases are deleted or modified, The desired compound thus the degradation of its activity decreased, without affecting cell viability. In a variety of intelligence State, the desired fine chemical production, or production rate is increased. ...

Protein of the invention and changes in the nucleotide molecule may also increase the ammonia by indirect mechanisms Amino acids, vitamins, cofactors, nutritional factors, nucleotides, nucleosides, or other than trehalose Fine chemicals production. Metabolism of any one compound bound to the intracellular synthesis of other biological And associated degradation pathways, in a way necessary cofactors, intermediates, or the substrate may be provided by other Supply or subject to restrictions similar way. Therefore, by adjusting one or more of the activities of the protein of the invention Resistance, another fine chemical biosynthetic pathway or degradation activity or efficiency of production may be Affected. For example, amino acids may be used as structural units of all proteins, but in the cell The level of the presence of protein synthesis may be restricted; Thus, by increasing the intracellular one or more Productivity of amino acids or rate of production, such as protein biosynthesis or degradation proteins easier Easy synthesis. Similarly, the metabolic pathway enzymes specific adverse change makes more favorable or unfavorable when Result of one or more substances for the production of the desired fine chemical intermediates or compounds over a substrate Volume production or insufficient production. ...

The present invention provides a new nucleic acid molecule encoding the protein, a protein referred to herein as the Metabolic pathway (MP) protein, they are able to complete the normal function of cells important molecules such as Amino acids, vitamins, cofactors, nucleotides and nucleotide metabolism or the enzymatic trehalose Steps. A nucleic acid molecule encoding proteins MP MP referred to herein as a nucleic acid molecule. In a preferred embodiment Case, MP proteins perform one or more of the following substances enzymatic metabolism steps: amino Acids, vitamins, cofactors, nutritional factors, nucleotides, nucleosides, and trehalose. These proteins Examples, including those listed in Table 1 of the protein encoded by the gene. ...

Accordingly, an aspect of the present invention, comprising a separate encoding a protein or MP Part of its biological activity by the nucleic acid sequence a nucleic acid molecule (e.g., cDNA, DNA, or RNA), and the separation is suitable as a detection or amplification of a nucleic acid encoding MP (such as DNA or RNA) primers or hybridization probes for nucleic acid fragment. In a particularly preferred embodiment, the sub- The nucleic acid molecule from a column containing the sequence in the sequence listing the nucleic acid sequence is an odd number (for example, Eg, SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7 ...) or a The nucleotide sequence of the coding region or the complementary sequence thereof. In other particularly preferred embodiment Solution, the isolated nucleic acid molecule of the present invention contains the sequence number of the sequence table is odd nucleoside Acid sequence (e.g., SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7 ...) or part thereof has at least about 50% homology, preferably at least about 60% of the Homology, more preferably at least about 70%, 80%, or 90% homology, even more preferably To have at least about 95%, 96%, 97%, 98%, 99% or higher homology. In other preferred Embodiments, the isolated nucleic acid molecule encoding the sequence shown in the sequence table, the even number group Acid sequence (e.g., SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8 ...). Preferred proteins of the present invention is also preferably MP described herein having at least one MP Activity. ...

In another embodiment, the isolated nucleic acid molecule encoding a protein or a portion Points, in which the protein or part thereof containing an amino acid sequence that is the present invention The amino acid sequence (for example, even in the sequence listing SEQ ID sequence) sufficient homology For example, the amino acid sequence of the present invention, there is sufficient homology such that the protein or the portion Points with MP activity. Preferably, the nucleic acid molecule encoding a protein or part thereof, maintaining for Amino acids, vitamins, cofactors, nucleotides and nucleotide metabolic pathway or the enzymatic trehalose Response capabilities. In one embodiment, the nucleic acid molecule encoding a protein of the present invention ammonia Amino acid sequence (for example, from the even sequence table serial number sequence selected from the complete amino acid sequence Column) at least about 50% homology, preferably at least about 60% homology, more preferably There are at least about 70%, 80%, 90% homology, most preferably at least about 95%, 96%, 97%, 98%, 99% or higher homology. In another preferred embodiment, the protein Corynebacterium glutamicum is the full length protein, the protein of the present invention the full length amino acid sequence (from the The corresponding sequence shown in the odd sequence number in nucleic acid sequence (e.g., SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7 ...) encoded by open reading frame) with sufficient Sources. ...

In another preferred embodiment, the isolated nucleic acid molecule from Corynebacterium glutamicum, And encodes a protein (for example, a MP fusion protein), the protein contains a biological The active region, which the present invention is an amino acid sequence (e.g., the even sequence Sequence A sequence of the sequence number) of at least about 50% or more homology, and the protein Substance capable of catalyzing the amino acids, vitamins, cofactors, nucleotides and nucleotide metabolic pathways or trehalose Diameter of the reaction, or have one or more listed in Table 1 of the activity, and the protein Further comprising a heterologous polypeptide coding regulatory region or heterologous nucleic acid sequence. ...

Another aspect of the present invention, the carrier, such as those containing a recombinant nucleic acid molecule of the present invention. Expression vector, and this vector is introduced into host cells. In one embodiment, by Carried out in a suitable culture medium, such host cell proteins are used to produce MP. Then From the medium or the host cell can be isolating the MP protein.

Further, another aspect of the present invention relates to a genetically altered microorganism, MP Wherein the gene has been introduced or has been changed. In one embodiment, by introducing for the The transgene coding for the wild-type or mutant sequences MP nucleic acid molecule of the invention, changing the The genome of the microorganism. In another embodiment, the change in the genome of the microorganism MP endogenous gene, for example, by using the MP gene has been altered by homologous recombination for Functional damage. In another embodiment, the microorganism of endogenous or introduced MP Gene through one or more point mutations, deletions or inversions is changed, but still can be compiled Code function MP proteins. In another embodiment, the change in a gene or a microorganism MP By a plurality of adjustment areas (e.g., promoter, repressor, or inducer), thus regulating MP Gene expression. In a preferred embodiment, the microorganism belongs to the species Corynebacterium or Brevibacterium species, Particularly preferably Corynebacterium glutamicum. In a preferred embodiment, it is also produced by using a microorganism To be compounds such as amino acids, particularly preferably lysine. ...

Further, another aspect of the present invention, the MP protein has been isolated or part, for example, Biological activity as its part. In a preferred embodiment, the isolated protein or the MP Part can be catalytic amino acids, vitamins, cofactors, nutritional factors, nucleotides, nucleosides, or sea Fucose metabolism pathway in one or more of the enzymatic reaction. In another preferred embodiment, In the separated protein or part MP of the invention an amino acid sequence (e.g., Sequence SEQ ID sequence, even a sequence) with sufficiently high homology, so that the protein Or a part of the catalytic mass to maintain amino acids, vitamins, cofactors, nutritional factors, nucleotides, Nucleoside or trehalose metabolism pathway in one or more of the ability of the enzymatic reaction. ...

MP present invention also provides isolated protein products. In a preferred embodiment, MP egg White comprise the amino acid sequence of the invention (e.g., SEQ ID sequence even in an ordered sequence Column). In another preferred embodiment, the present invention relates to the isolated full-length protein, The protein and the complete amino acid sequence of the present invention (having the sequence SEQ ID sequence even a Sequence) (the sequence shown in the sequence corresponding to the odd number of open reading frame encoding) Have a very high homology. Further, in another embodiment, the protein of the present invention the complete Full amino acid sequence (e.g., the sequence of SEQ ID sequence even) at least about 50% homology , It is preferable to have at least about 60% homologous, more preferably at least about 70%, 80%, or 90% homology, most preferably at least about 95%, 96%, 97%, 98%, or 99%, or more High homology. In another embodiment, the isolated protein comprises the amino acid sequence of MP And the amino acid sequence of the present invention is one (e.g., the sequence of the even sequence number of a sequence) At least about 50% or more homology to the catalytic amino acids and vitamins, auxiliary Factors, nutritional factors, nucleotides, nucleosides, or trehalose in the enzymatic reaction pathway, or Listed in Table 1 in one or more of the active. ...

MP polypeptide or a biologically active portion, can be effectively connected to the polypeptide and form of MP A fusion protein. In a preferred embodiment, the fusion protein having a different MP alone Protein activity itself. In another preferred embodiment, the fusion protein is introduced glutamyl Corynebacterium amino acids, vitamins, cofactors, nutritional factors metabolic pathway, causing acid Corynebacterium fine chemicals desired yield, production and / or production efficiency increases. In particularly preferred Selected embodiments, the fusion protein of the amino acid integrated into the host cell, microorganism, auxiliary Factors, nutritional factors, nucleotides, nucleosides, or trehalose metabolic pathways in the cell can be adjusted Production of desired compound. ...

Another aspect of the invention relates to a method for the production of fine chemicals. The method involves training Cultured cells comprising a vector, the vector instructions MP nucleic acid molecule of the present invention the expression, thereby Produce fine chemicals. In a preferred embodiment, the method further comprises obtained contains The step of the support cell, in this step, the use of a nucleic acid molecule can direct the expression of MP load Transfected cells. In another preferred embodiment, the method further comprises back from the medium The step of closing fine chemicals. In a particularly preferred embodiment, the cells are Corynebacterium Brevibacterium species or species listed in Table 3 or is selected from those strains. ...

Another aspect of the present invention is a molecule produced microorganism adjustment method. In this way France including the use of adjustable MP or MP nucleic acid protein active pharmaceutical contacting cells expressing such Cells in the absence of such activity with respect to the active agent changed. In a preferred Embodiment, the adjustment of Corynebacterium glutamicum cells, one or more amino acids, vitamins, Cofactors, nutritional factors, nucleotides, nucleosides, or trehalose metabolic pathways, such that this microorganism Yield of the desired fine chemical substances or generation efficiency is improved. Regulating protein activity MP Agents, which can be stimulated MP or MP nucleic acid protein activity expressed agents. Stimulation MP protein MP nucleic acid expression or activity of the drug of examples, including small molecules, proteins MP activity in And encodes the protein into cells MP nucleic acid. MP protein activity or inhibiting the expression of drug Agents include small molecules and antisense nucleic acid molecule MP. ...

Another aspect of the invention, the desired compound involved in the regulation of cell production methods, including Comprising the wild type or mutant MP gene into the cells, the gene or kept in a separate mass Tablets, or integrated into the host cell genome. If integrated into the host cell genome, This integration may be arbitrary, or by homologous recombination occurs, so that the introduction of Copy replace the natural gene, resulting in the generation of the desired compound in the cell is adjusted. In an excellent Selected embodiments, the production has been increased. In another preferred embodiment, said Fine chemicals are amino acids. In a particularly preferred embodiment, said amino Acid is L-lysine. ...

MP The present invention provides nucleic acids and protein molecules, their participation in a Corynebacterium glutamicum These fine chemicals including amino acids, vitamins, cofactors, nutritional factors, nucleotides, nuclear Glycosides and trehalose metabolism. Molecules of the invention can be used to adjust the microorganisms such as Corynebacterium glutamicum Production of fine chemicals, such an adjustment may be direct (e.g., lysine biosynthesis The adjustment of the protein into the lysine-producing organism or production efficiency has a direct impact), Or have indirect effects, but will result in production of the desired compound, or an increase in productivity (for example, When the nucleotide biosynthesis regulation of protein activity against bacterial production of organic acids or fatty acid shadow Ring, may be improved because of the growth or necessary cofactors, energy compound or precursor molecule for Should be increased). The various aspects of the present invention is described in further detail below. ...

"Fine chemical" this word is known in the art, including in various organisms Molecular species used in industry, such as, but not limited to, pharmaceuticals, agriculture, and cosmetics industries. Such compounds include organic acids, such as tartaric acid, itaconic acid, and diaminopimelic acid, protein Source and non-protein source of amino acids, purine and pyrimidine bases, nucleosides and nucleotides (case Like the device described in Kuninaka, A. (1996) Nucleotides and related compounds, p. 561-612, in Biotechnology vol.6, Rehm et al., Eds.VCH: Weinheim and its Included in the references), lipids, saturated and unsaturated fatty acids (e.g., arachidonic acid), Diols (e.g., propane diol and butane diol), aromatic compounds (e.g., aromatic amines, Vanillin and indigo), vitamins and cofactors (see Ullmann's Encyclopedia of Industrial Chemistry, vol.A27, "Vitamins", p.443-613 (1996) VCH: Weinheim and references therein; and Ong, AS, Niki, E. & Packer, L. (1995) "Nutrition, Lipids, Health, and Disease "Proceedings of the UNESCO / Confederation of Scientific and Technological Associations in Malaysia, and the Society for Free Radical Research-Asia, held Sept. 1-3,1994 at Penang, Malaysia, AOCS Press, (1995)), enzymes, polyketides (Ployketides) (Cane et al., (1998) Science 282:63-68), and all the Gutcho (1983) Chemicals by Fermentation, Noyes Data Corporation, ISBN: 0818805086 and The reference substances described in the literature. Some of these fine chemicals into the metabolism and uses Described in detail further below. ...

Amino acids include basic structural units of all proteins, are also all organisms normally Cell biological features necessary. "Amino acid" is a term known in the art. Protein Source 20 kinds of amino acids, protein structural units, separated by a peptide bond is connected to, not the Source of protein amino acids (there are hundreds of known) would not normally occur in proteins (reference See Ulmann's Encyclopedia of Industrial Chemistry, vol.A2, p.57-97 VCH: Weinheim (1985)). While the natural L-amino acids are usually the only type of protein, However, the D-amino acid may or L-optical configuration. 20 kinds of amino acids in the protein source in each of Biosynthesis or degradation pathways are in prokaryotic cells or eukaryotic cells have their own characteristics (for example, Eg see Stryer, L.Biochemistry, 3 ...^rdedition, pages 578-590 (1988)). "Necessary" Amino acids (histidine, isoleucine, leucine, lysine, methionine, phenylalanine, Threonine, tryptophan and valine) is so named because these amino acid biosynthesis Into complex is usually necessary nutritional conditions, they can be simply converted biosynthetic pathway 11 kinds of the remaining "non-essential" amino acids (alanine, arginine, asparagine, aspartic Acid, cysteine, glutamic acid, glutamine, glycine, proline, serine, tyrosine). Although higher organisms does have the ability to synthesize some of these amino acids, but for normal egg White matter from the diet must be supplemented synthetic essential amino acids. ...

In addition to their function in protein synthesis, these amino acids is interesting in its own right Chemicals, and many of them in the food, feed, chemicals, cosmetics, agriculture and The pharmaceutical industry has a variety of applications. Lysine in nutrition not only for human beings is an important Amino acids, such as poultry and pigs but also for monogastric animals is so important. Glutamate is The most common flavor additive (monosodium glutamate, MSG), and is widely used throughout the food Industry, as aspartic acid, glycine, cysteine same. Glycine, L-methionine and All tryptophan used in the pharmaceutical industry. Glutamine, valine, leucine, isoleucine, group Acid, arginine, proline, serine and alanine are used in the pharmaceutical and cosmetic industry production Sector. Threonine, tryptophan and methionine are commonly used D/L- feed additive (Leuchtenberger, W. (1996) Amino aids-technical production and use, p. 466-502 in Rehm et al. (Eds.) Biocemistry vol.6, chapter 14a, VCH: Weinheim). In addition, these amino acid and protein synthesis as a synthetic precursor is Is useful, for example, N-acetylcysteine, S-carboxymethyl-L-cysteine, (S) -5 - serotonin Acid, and the other in Ulmann's Encyclopedia of Industrial Chemistry, vol.A2, p. 57-97 VCH: Weinheim, 1985 molecules described. ...

Natural amino acids in organisms capable of producing, for example bacteria, these natural amino acids Biosynthesis have learned very full (bacterial amino acid biosynthesis and its regulation, see Umbarger, HE (1978) Ann.Rev.Biochem.47 :533-606). Aspartic acid from the α-keto Reduction amination of glutaric acid, which is the citric acid cycle intermediates. Glutamine, proline Glutamic acid and arginine are made sequentially generated. The biosynthesis of serine is a three-step Process starts at 3 - phosphoglycerate (glycolytic intermediates), after oxidation, aminotransferase , Hydrolysis of the step, terminates at the amino acids. Cysteine and glycine by Serine produced; former by the condensation of homocysteine with serine, which is the β-carbon of the side chain Atom transfer to tetrahydrofolate obtained, the reaction is catalyzed by serine hydroxymethyl transferase enzyme catalyzed. Phenylalanine and tyrosine, from 4 - erythritol phosphate and phosphoenolpyruvate in a 9-step of raw Synthesized in biosynthetic pathway, which are glycolytic pathway and pentose phosphate pathway precursors, these two The synthetic route is different after prephenate. Tryptophan can also be produced by these two initial molecular Students, but their synthesis is a 11-step approach. Tyrosine can also be synthesized by a phenylalanine, the The reaction is catalyzed by the phenylalanine hydroxylase. Alanine, valine and leucine are glycolysis The end product of the biosynthesis of pyruvate. Aspartic acid synthesis by the oxaloacetate, which is lemon Acid cycle intermediates. Asparagine, methionine, threonine and lysine by aspartic acid Transformed. Isoleucine by a threonine formed. Through a complex 9-step pathway, can From a reactive sugar, 5 - phosphoribosyl -1 - pyrophosphate, produce histidine. ...

Natural amino acids in organisms capable of producing, for example bacteria, these natural amino acids Biosynthesis have learned very full (bacterial amino acid biosynthesis and its regulation, see Umbarger, HE (1978) Ann.Rev.Biochem.47 :533-606). Aspartic acid from the α-keto Reduction amination of glutaric acid, which is the citric acid cycle intermediates. Glutamine, proline Glutamic acid and arginine are made sequentially generated. The biosynthesis of serine is a three-step Process starts at 3 - phosphoglycerate (glycolytic intermediates), after oxidation, aminotransferase , Hydrolysis of the step, terminates at the amino acids. Cysteine and glycine by Serine produced; former by the condensation of homocysteine with serine, which is the β-carbon of the side chain Atom transfer to tetrahydrofolate obtained, the reaction is catalyzed by serine hydroxymethyl transferase enzyme catalyzed. Phenylalanine and tyrosine, from 4 - erythritol phosphate and phosphoenolpyruvate in a 9-step of raw Synthesized in biosynthetic pathway, which are glycolytic pathway and pentose phosphate pathway precursors, these two The synthetic route is different after prephenate. Tryptophan can also be produced by these two initial molecular Students, but their synthesis is a 11-step approach. Tyrosine can also be synthesized by a phenylalanine, the The reaction is catalyzed by the phenylalanine hydroxylase. Alanine, valine and leucine are glycolysis The end product of the biosynthesis of pyruvate. Aspartic acid synthesis by the oxaloacetate, which is lemon Acid cycle intermediates. Asparagine, methionine, threonine and lysine by aspartic acid Transformed. Isoleucine by a threonine formed. Through a complex 9-step pathway, can From a reactive sugar, 5 - phosphoribosyl -1 - pyrophosphate, produce histidine. ...^rded.Ch. 21 "Amino Acid Degradation and the Urea Cycle" p.495-516 (1988)). Exhausted Duct cells can be transformed into useful extra amino acid metabolic intermediates, but the resulting amino acids to eliminate Consume a lot of energy, precursor molecules and the enzymes required for synthesis. Hence the feedback inhibition to regulate amino Acid biosynthesis is not surprising, particularly the presence of amino acids can slow down or completely stop For the generation of its own (for amino acid biosynthesis pathway feedback mechanisms reviews, see Stryer, L.Biochemistry 3^rded.Ch.24 "Biosynthesis of Amino Acid and Heme" p. 575-600 (1988)). Thus, the output of any particular amino acid are in the group of memory cells Acid number is limited.

B. vitamins, cofactors, and metabolism and the use of nutritional factors

Vitamins, cofactors, and another group of molecules, including nutritional factors, although other organisms, such as Bacteria, these molecules can be synthesized, but the higher animals have lost the ability of their synthesis can Intake. These molecules or biologically active substance which is itself, or the biologically active substance before Body, the biological active substance may be an electron carrier or a variety of metabolic pathways intermediates. In addition to Its nutritional value, these compounds as pigments, antioxidants and catalysts or other processing aids Agent also has a significant industrial value. (For these compounds the structure, activity, and industrial applications ratings Above, see for example, Ullmann's Encyclopedia of Industrial Chemistry, "Vitamins" vol.A27, p.443-613 VCH: Weinheim 1996.) "vitamin" the word is in the field Is well known, including the normal functions of the organism but can not be synthesized nutrients. Dimension Antibiotics can include compounds of nutritional factors and cofactors. The term "cofactor" includes conduct regular Activity often desired non-protein compounds. These compounds may be inorganic or organic A; cofactor molecules of the invention are preferably organic. "Trophic factor" is the word contains a Plants and animals, especially beneficial dietary supplements. Examples of such molecules are vitamins, Antioxidants and certain lipids (such as multi-unsaturated fatty acids). ...

Thiamine (vitamin B1) is a pyrimidine and thiazole produced by chemical connection. Riboflavin (Victoria Vitamin B2) by the 5'-triphosphate and guanosine 5'-ribose synthesis. Riboflavin sequential use in Synthesis of flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD). Collectively referred to as "dimension Vitamin B6 "a group of compounds (e.g., pyridoxine, pyridoxamine, pyridoxal 5'-phosphate, and Commercial pyridoxal hydrochloride) are common structural unit 5 - hydroxy-6 - methyl-pyridine derivatives. Pantothenate (pantothenic acid, (R) - (+)-N-(2,4 - dihydroxy-3 - dimethyl-1 - oxobutyl)-β-alanine) By chemical synthesis or fermentation. Pantothenate biosynthesis drive the final step, the ATP The β-alanine and pantoic acid condensation. Charge into pantoic acid and β-alanine enzyme, and reduced Pantothenate synthesis enzymes are known. Metabolic active form of pantothenate is coenzyme A, the biological Enzymatic synthesis process of step 5. Pantothenate, pyridoxal 5'-phosphate, cysteine and ATP are Coenzyme A precursor. These enzymes not catalyzed the formation of pantothenate, but also the catalytic (R) - pantoic acid, (R) - pantolacton, (R) - panthenol (provitamin B5) cysteamine panthenol (and derivatives thereof) of Production. ...

In microorganisms from the precursor molecule pimeloyl coenzyme A to biotin biosynthesis very Specifically, several genes involved have been identified. Many proteins have been found in the corresponding parameter And clusters of iron (Fe-cluster) synthesis, and is nifS protein family member. Lipoic acid to Since acid, used as a coenzyme in energy metabolism, can be a pyruvate dehydrogenase complex and the α-keto Glutarate dehydrogenase complex part. Folate is a folic acid derivative, followed by L-glutamic acid, p-aminobenzoic acid and 6 - methyl neopterin. Starting in metabolic intermediates guanosine 5'-triphosphate Purine (GTP), L-glutamic acid and amino acid biosynthesis of folic acid and its derivatives, In certain microorganisms in detail. ...

Although some of these compounds can also be produced by microbial culture scale, for example riboflavin , Vitamin B6, pantothenic acid and biotin, but large scale production of these compounds also largely Dependent on cell-chemical system. Only vitamin B12, as the complexity of its synthesis, only By fermentation. Vitro methods require considerable material and time investment, often costly.

C. purine, pyrimidine, nucleoside and nucleotide metabolism and uses

Purine and pyrimidine metabolism of genes and their corresponding proteins, tumor diseases and viral influenza Important therapeutic target dye. The term "purine" and "pyrimidine", contains a nucleic acid, secondary Enzymes and nucleotide composition of the nitrogenous base. The term "nucleotide" the basic structure of a nucleic acid molecule that contains a single Element, the nucleic acid molecule consists of nitrogenous bases, pentose sugars (for RNA, the pentose sugar is ribose; For DNA, The pentose sugar is deoxyribose) and phosphate components. The term "nucleoside" includes a nucleotide precursors Molecule, but the lack of a phosphate portion of nucleotides. These molecules by inhibiting biological Synthesis, or inhibition of nucleic acid molecules for the synthesis of the movement carried out, may be inhibition of RNA and DNA Synthesis; through targeted tumor cells to inhibit the active mode, the tumor cell division and replication The energy may be suppressed. In addition, some nucleotides not used to form a nucleic acid, but as Energy storage (eg AMP) or coenzyme (such as FAD and NAD). ...

Some publications describe by influencing purine and / or pyrimidine metabolism, these chemicals as The use of these medical indications (eg, Christopherson, RIand Lyons, SD (1990) "Potent inhibitors of de novo pyrimidine and purine biosynthesis as chemotherapeutic agents. "Med.Res.Reviews 10:505-548). involving purine and pyrimidine Pyridine metabolic enzymes studies focused on the development of new drugs that can be used above, for example, as an immune Inhibitor or antiproliferative agents (Smith, JL, (1995) "Enzyme in nucleotide synthesis. "Curr.Opin.Struct.Biol.5 :752-757; (1995) Biochem Soc. Transact.23 :877-902). However, purine and pyrimidine bases, nucleosides and nucleotides also have other Outside effects: as many fine chemicals biosynthetic intermediates (e.g., thiamine, S-adenosyl methionine, folic acid, or riboflavin), as a cell energy carrier (such as ATP Or GTP), and as a chemical substance itself, usually used as flavor enhancers (for example IMP or By GMP) or several medical applications (see, for example, Kuninaka, A. (1996) Nucleotides and Related Compounds in Biotechnology vol.6, Rehm et al., eds.VCH: Weinheim,, p.561-612). Similarly, involving purine, pyrimidine, nucleoside or nucleotide metabolism of Enzymes, becoming developed chemicals for crop protection role of objectives, which Chemicals including fungicides, herbicides and insecticides. ...

The metabolism of these compounds in bacteria has a characteristic (a review see, for example, Zalkin, H.and Dixon, JE (1992) "de novo purine nucleotide biosynthesis", in: Progress in Nucleic Acid Research and Molecular Biology, vol.42, Academic Press:, p.259-287; and Michal, G. (1999) "Nucleotides and Nucleosides", Chapter 8 in: Biochemical Pathways: An Atlas of Biochemistry and Molecular Biology, Wiley: New York). Purine metabolism has been the focus of research, and it is necessary for normal cell function. Higher Purine metabolism in animals can cause damage serious diseases, such as gout. Purine nucleotides by the Synthesis of 5'-phosphoribosyl, through a series of steps through the intermediate inosine 5'-monophosphate (IMP), ultimately produce guanosine 5'-monophosphate (GMP) and adenosine 5'-monophosphate (AMP), They are formed by the nucleotide triphosphate form used. These compounds are also used as an energy store, Degradation of the cell to provide a variety of biochemical processes of energy. Pyrimidine biosynthesis, is through Had formed by the 5'-ribose uridine 5'-monophosphate (UMP). UMP into the next Cytosine 5'-triphosphate (CTP). All of these nucleotides deoxy forms are to undergo a step further Reduction reaction generated by the nucleotide diphosphate ribose form of the nucleotide diphosphate deoxyribose Form. Upon phosphorylation, these molecules can participate in DNA synthesis of. ...

Trehalose consists of two glucose molecules by α, α-1, 1 connection. Usually in the food industry Used as sweetening agents, dry foods or frozen food additives and beverages which. Furthermore, It is also used in the pharmaceutical, cosmetics and biotechnology industries (see, for example Nishimoto et al., (1998) USPatent No.5, 759,610; Singer, MAand Lindquist, S. (1998) Trends Biotech.16 :460-467; Paiva, CLAand Panek, AD (1996) Biotech. Ann.Rev.2 :293-314; and Shiosaka, M. (1997) J.Japan 172:97-102). Many The enzyme can produce a microorganism trehalose, and released naturally into the surrounding medium can Which technically well known methods using for collection.

II. Components and methods of the present invention,

The present invention is based at least in part on the discovery of new molecules on the basis, where it is called MP Nucleic acid and protein molecules of one or more cells are metabolic pathways play a role or function Energy. In one embodiment, MP Molecular Catalysis of one or more amino acids, vitamins, cofactors Son, nutritional factors, nucleotides, nucleosides, or trehalose in the enzymatic reaction pathway. In the preferred Embodiment, the present invention is one or more of Corynebacterium glutamicum amino acids, vitamins, cofactors Son, nutritional factors, nucleotides, nucleosides, or trehalose in the metabolic pathway of a molecule active MP The microbial production of fine chemicals needed impact. In a particularly preferred embodiment, , The present invention is adjusted MP activity of molecules, so that this invention is proteins involved in the metabolism of an MP Efficiency or production pathway is regulated, which will directly or indirectly affect one or Corynebacterium glutamicum Variety of fine chemicals production and production efficiency. ...

The term "MP protein" or "MP polypeptide" included in the one or more amino acids, Victoria Vitamin, cofactor, nutritional factors, nucleotides, nucleosides, or play a role in metabolic pathways, such as trehalose Wherein the enzymatic reaction catalyzed protein. MP proteins include those listed in the sequence listing by the MP sequence number is odd genes encoding proteins. The term "MP gene" or "MP Nuclear Acid sequence "includes a nucleic acid sequence encoding MP protein, which contains the coding region and the corresponding Non-translated 5 'and 3' sequence regions. MP genes include those listed in Table 1 of the base Result. The term "production" or "productivity" is known in the art, including at a given time And a given volume of fermentation, fermentation products (eg, the desired fine chemical) concentration (for example, For example, kg product per hour per liter). The term "productivity" contains the, to achieve a particular The time required for production levels (for example, how long it takes the cell reaches a certain fine Chemical substances). The term "income", "product / carbon yield" is known in the art, including the Carbon conversion products (for example, fine chemicals) efficiency. For example, often writing kg capacity Substance per kg carbon. By increasing revenue or production compounds, increase the number of molecules recovered , Or increase in a given period of time a given quantity of the compound in the culture of molecules useful in recovery Number. The term "biosynthesis" or a "biosynthetic pathway" are known in the art, including Contained in the cells may be from the intermediate compound through a multi-step process and is highly regulated, Synthetic compounds, in particular compounds. The term "degradable" or a "degradation pathway" Is known in the art, including in cells, probably through multi-step and is highly regulated Process, the compound is preferably an organic compound, the decomposition of degradation products (in general, the Smaller or less complex molecules). The term "metabolic" is known in the art, including the Organisms biochemical reactions that occur in all. Thus, the metabolism of specific compounds (e.g., Metabolism of amino acids such as glycine this) cells with the compound, including all related biological Synthesis, modification and degradation pathways. ...

In another embodiment, the present invention is to regulate the microorganism MP molecules, such as Corynebacterium glutamicum, the desired compound such as the production of fine chemicals. Using recombinant genetic Technique of the present invention can operate one or more amino acids, vitamins, cofactors, nutrition because Son, nucleotides, nucleosides, or trehalose biosynthesis or degradation enzymes, thereby regulating their activity. For example, Biosynthetic enzymes can increase the efficiency or destruction of its allosteric control region so as to prevent production of the compound Feedback inhibition. Similarly, a degradative enzyme can be substitution, deletion or increases are deleted or modified, The desired compound thus the degradation of its activity decreased, without affecting cell viability. In a variety of intelligence State, the desired fine chemical production, or production rate is increased. ...

Proteins and nucleotides of the invention may also improve the molecule in addition to changes in the amino acid, vitamin Factors, cofactors, nutritional factors, nucleotides, nucleosides, or trehalose fine chemicals other than Qualitative production. Metabolism of any one compound bound to the intracellular synthesis and degradation pathways of other organisms Association, in a way necessary cofactors, intermediates, or the substrate may be provided by other similar means for Give or be limited. Therefore, by adjusting one or more of the active protein of the invention, another Fine chemicals or degradation pathway biosynthetic activity or efficiency of production may be affected. For example, amino acids may be used as structural units of all proteins, but its presence in the cell Level may restrict protein synthesis; Thus, by increasing the intracellular one or more amino acids of the Production efficiency or productivity, such as the protein biosynthesis or degradation can be more easily synthesized protein. With Kind, metabolic pathway enzymes specific adverse change makes more favorable or unfavorable, it will lead to one or Required for the production of a variety of fine chemical intermediates or compounds substrate excess production or raw Production is insufficient. ...

The present invention also related to such a protein, the amino acid sequence of the protein of the present invention Sufficient amino acid sequence homology (for example, the sequence of the even sequence of the sequence). As As used herein, has the amino acid sequence selected have sufficient homology to the amino acid sequence Column of protein, and the amino acid sequence selected, for example, the amino acid sequences of selected with At least about 50% homology. Having the amino acid sequence selected a great homology group Acid sequence of the protein, but also with an amino acid sequence selected has at least about 50-60%, preferably Selected to have at least about 60% homologous, more preferably at least about 70%, 80%, 90% of the Homology, most preferably at least about 95%, 96%, 97%, 98%, 99% or higher homology Sex. ...

The following sections describe in more detail the various aspects of the present invention:

Isolated nucleic acid molecule A.

One aspect of the invention relates to an isolated polypeptide encoding MP or parts of its biological activity Nucleic acid molecules, and sufficient for the hybridization probe or primer is a nucleic acid molecule fragments of MP for the identification or coding nucleic acid amplification (eg MP DNA). As used herein, that Like, the term "nucleic acid molecule" means comprising a DNA molecule (e.g., cDNA or gene Groups DNA) and RNA molecules (e.g. mRNA), and the resulting DNA nucleotide analogs Or RNA analogs. The term also includes gene coding region in the 3 'and 5' untranslated end of the Sequence: coding region of the 5 'end of the sequence upstream of at least 100 nucleotides, and gene coding region 3 'end of the sequence downstream of at least 20 nucleotides. The nucleic acid molecule may be single-stranded or double-stranded , But preferably is double-stranded DNA. An "isolated" nucleic acid molecule refers to those present in the nucleus Other natural sources acid a nucleic acid molecule to an isolated nucleic acid molecule. Preferably, the "isolated" Nucleic acids do not contain genomic DNA of the organism in a natural nucleic acid sequences on both sides (for example, in DNA 5 'and 3' ends of the sequence), the nucleic acid is obtained from the organism. For example, in each Colors embodiment, the isolated nucleic acid molecule can contain less than MP about 5kb, 4kb, 3kb, 2kb, 1kb, 0.5kb or 0.1kb nucleotide sequence which naturally located in the cell genome DNA On both sides of the nucleic acid molecule, the nucleic acid is from these cells (e.g., Corynebacterium glutamicum cells) Obtained. In addition, "isolated" nucleic acid molecule, such as DNA molecules, recombinant techniques when raw Production can substantially contain other cellular material or culture medium, when the chemical synthesis can not Combined chemical precursors or other chemicals. ...

One aspect of the invention relates to an isolated polypeptide encoding MP or parts of its biological activity Nucleic acid molecules, and sufficient for the hybridization probe or primer is a nucleic acid molecule fragments of MP for the identification or coding nucleic acid amplification (eg MP DNA). As used herein, that Like, the term "nucleic acid molecule" means comprising a DNA molecule (e.g., cDNA or gene Groups DNA) and RNA molecules (e.g. mRNA), and the resulting DNA nucleotide analogs Or RNA analogs. The term also includes gene coding region in the 3 'and 5' untranslated end of the Sequence: coding region of the 5 'end of the sequence upstream of at least 100 nucleotides, and gene coding region 3 'end of the sequence downstream of at least 20 nucleotides. The nucleic acid molecule may be single-stranded or double-stranded , But preferably is double-stranded DNA. An "isolated" nucleic acid molecule refers to those present in the nucleus Other natural sources acid a nucleic acid molecule to an isolated nucleic acid molecule. Preferably, the "isolated" Nucleic acids do not contain genomic DNA of the organism in a natural nucleic acid sequences on both sides (for example, in DNA 5 'and 3' ends of the sequence), the nucleic acid is obtained from the organism. For example, in each Colors embodiment, the isolated nucleic acid molecule can contain less than MP about 5kb, 4kb, 3kb, 2kb, 1kb, 0.5kb or 0.1kb nucleotide sequence which naturally located in the cell genome DNA On both sides of the nucleic acid molecule, the nucleic acid is from these cells (e.g., Corynebacterium glutamicum cells) Obtained. In addition, "isolated" nucleic acid molecule, such as DNA molecules, recombinant techniques when raw Production can substantially contain other cellular material or culture medium, when the chemical synthesis can not Combined chemical precursors or other chemicals. ...^nd, Ed.Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, In 1989) were isolated. Further, the present invention includes a nucleic acid sequence (e.g., sequence The nucleotide sequence of SEQ ID number) all or part of the nucleic acid molecule can be poly Synthase chain reaction, based on the sequences using oligonucleotide primers isolated (Example For example, the present invention includes a nucleic acid sequence (e.g., the sequence of the nucleotide sequence of the odd sequence number Column) all or part of a nucleic acid molecule that can by polymerase chain reaction based The sequences of the same oligonucleotide primers, isolated). For example, mRNA can Isolated normal endothelial cells (for example, using Chirgwin et al. (1979) Biochemistry 18: 5294-5299 guanidine thiocyanate extraction method in), DNA by reverse transcriptase (for example, Gibco / BRL, Bethesda, MD provided Moloney MLV reverse transcriptase; or Seikagaku America, Inc., St.Peterburg, FL supplied AMV reverse transcriptase) thereof. As polymerase Synthesized oligonucleotide chain reaction primers may be based on the sequence of a polynucleotide listed in Table Sequence Design. The nucleic acid of the invention may be used as an alternative or cDNA gene Genomic DNA as a template and appropriate oligonucleotide primers according to standard PCR amplification techniques to extend Increase. Such amplified nucleic acid can be cloned into a suitable vector and the DNA sequence analysis Identify its characteristics. Further, the nucleotide sequence corresponding to the MP oligonucleotides can be used standard Quasi-synthesis technology to prepare, for example using an automatic DNA synthesizer. ...

, Ed.Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, In 1989) were isolated. Further, the present invention includes a nucleic acid sequence (e.g., sequence The nucleotide sequence of SEQ ID number) all or part of the nucleic acid molecule can be poly Synthase chain reaction, based on the sequences using oligonucleotide primers isolated (Example For example, the present invention includes a nucleic acid sequence (e.g., the sequence of the nucleotide sequence of the odd sequence number Column) all or part of a nucleic acid molecule that can by polymerase chain reaction based The sequences of the same oligonucleotide primers, isolated). For example, mRNA can Isolated normal endothelial cells (for example, using Chirgwin et al. (1979) Biochemistry 18: 5294-5299 guanidine thiocyanate extraction method in), DNA by reverse transcriptase (for example, Gibco / BRL, Bethesda, MD provided Moloney MLV reverse transcriptase; or Seikagaku America, Inc., St.Peterburg, FL supplied AMV reverse transcriptase) thereof. As polymerase Synthesized oligonucleotide chain reaction primers may be based on the sequence of a polynucleotide listed in Table Sequence Design. The nucleic acid of the invention may be used as an alternative or cDNA gene Genomic DNA as a template and appropriate oligonucleotide primers according to standard PCR amplification techniques to extend Increase. Such amplified nucleic acid can be cloned into a suitable vector and the DNA sequence analysis Identify its characteristics. Further, the nucleotide sequence corresponding to the MP oligonucleotides can be used standard Quasi-synthesis technology to prepare, for example using an automatic DNA synthesizer. ...

For the purpose of the application, the sequence listed in the table can understand each of nucleic acids and amino acid sequence Column, there is a for identification RXA, RXN, RXS or RXC number, "RXA", "RXN", "RXS", or "RXC" followed by a five digits (i.e., RXA00007, RXN00023, RXS00116, or RXC00128). Every nucleic acid sequence contains up to three Part: 5 'upstream region, the coding region, downstream region. Each part of the three regions, all with Same RXA, RXN, RXS, or RXC name OK to eliminate confusion. So narrative "A number in the sequence table sequence encoding" refers to any sequence in a nucleic acid sequence, These sequences can also use their different RXA, RXN, RXS, or RXC name mutual Distinction. Each coding region of this sequence is translated into the corresponding amino acid sequence, the sequence Columns are also listed in the sequence listing, a nucleic acid sequence corresponding to immediately even after the serial number. For example, RXA02229 coding region shown in SEQ ID NO: 1, and its encoded amino acid sequences are listed in SEQ ID NO: 2. The nucleic acid molecule sequence of the present invention, with its encoded amino acid molecules by phase Same RXA, RXN, RXS, or RXC name that makes them easy to contact each other. For example, known RXA02229, RX00351, RXS02970 and the amino acid sequence RXC02390 Columns are RXA02229, RX00351, RXS02970 and RXC02390 nucleic acid molecules nuclear Nucleotide sequence of the coding region of the translation. The present invention RXA, RXN, RXS and RXC nucleotides and Correspondence between the amino acid sequences, and their serial numbers are specified in Table 1. ...

In one embodiment, the present invention, the nucleic acid molecule does not contain the compiled in Table 2, those Corynebacterium glutamicum molecules. The dapD gene, the gene sequence published in Wehrmann, A., et al. (1998) J.Bacteriol.180 (12) :3159-3165. However, the present inventors have obtained The much longer than the published version. It is said that using the wrong version published by the fact codon And therefore only the performance of the real part of the coding region.

In one embodiment, the present invention, the nucleic acid molecule does not contain the compiled in Table 2, those Corynebacterium glutamicum molecules. The dapD gene, the gene sequence published in Wehrmann, A., et al. (1998) J.Bacteriol.180 (12) :3159-3165. However, the present inventors have obtained The much longer than the published version. It is said that using the wrong version published by the fact codon And therefore only the performance of the real part of the coding region....

Also in another preferred embodiment, the isolated nucleic acid molecule of the invention, including Such a nucleotide sequence which the nucleotide sequence of the present invention (e.g., the sequence of odd Serial number The serial number) or a part, at least about 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59% or 60% homology, preferably at least about 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69% or 70% homology, more preferably have at Less some 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79% or 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 88%, 89% or 90%, or 91%, 92%, 93%, 94%, and even more preferably at least about 95%, 96%, 97%, 98%, 99% Or higher homology. Cited above range (for example 70 to 90%, or 80-95% to the consistency Consistency) in the middle of the range and consistency values, are included in the present invention. For example, this includes Consistency range, these ranges are referenced above upper and / or lower limit value combination. In another A preferred embodiment, the present invention is an isolated nucleic acid molecule comprises a nucleotide sequence that Column, the sequence can be a nucleotide sequence of the present invention or parts thereof for hybridization, for example, Hybridizes under stringent conditions. ...

Further, the nucleic acid molecule of the present invention may contain only the sequence of the coding region of the sequence SEQ ID number Part of the domain, for example, can be used as probes or primers fragment or protein encoded MP Biologically active portions of clips. The MP gene of Corynebacterium glutamicum nucleotide sequence, Probes and primers allow the generation, these probes and primers designed for the identification and / or cloning other Cell type, or other organisms in the MP homologues, other genetic material, or Corynebacterium The MP species homologues. Probe / primer typically contains relatively purified oligonucleotides. Oligo Nucleotides typically including such a nucleotide sequence region that stringent hybridization conditions Next, the nucleotide sequence of the present invention (e.g., the sequence of SEQ ID sequence number) of a sense strand, An antisense sequence of these sequences, or of naturally occurring mutants of at least about 12, preferably About 25, more preferably about 40, 50 or 75 consecutive nucleotides hybridized. Based on The nucleotide sequence of the present invention, a primer can be used to clone homologues MP PCR reaction. Based on MP nucleotide sequence probe can be used to detect the same or homologous proteins or transcription-based Genome sequence. In a preferred embodiment, the probe is attached comprises an additional marker gene Group, such as a tag group can be a radioisotope, a fluorescent compound, an enzyme or enzyme cofactors Child. Such probes can be used as part of diagnostic test kits, the kit for the identification of fault MP protein expression cells mistake, such as a sample cell by measuring the level of nucleic acid encoding the MP, For example, the detection level of MP mRNA, or genomic MP determined whether there is a sudden genes Changed or missing. ...

In one embodiment, the present invention is a nucleic acid molecule encoding a protein or parts thereof, The protein or part of the amino acid sequence of the amino acid sequence of the present invention (e.g., SEQ Sequence SEQ ID table even) have sufficient homology such that the protein or part thereof Catalyzes amino acids, vitamins, cofactors, nutritional factors, nucleotides, nucleosides, or trehalose Enzymatic reaction in the metabolic pathway. As used herein, the term "sufficient homology" is the Refers to the protein or part of the amino acid sequence, containing the minimum number of amino acid sequence of the present invention. Column consistent or equivalent (eg, having a sequence table even the amino acid sequence of SEQ ID Similar residues amino acid residue side chain) amino acid residues, so that the protein or the portion Points, capable of catalyzing the Corynebacterium glutamicum amino acids, vitamins, cofactors, nutritional factors, nuclear Nucleotide, nucleoside, or trehalose in the enzymatic reaction pathway. This metabolic pathway protein members As described herein, as its function is to catalyze one or more amino acids, vitamins, cofactors, Nutritional factors, nucleotides, nucleosides, or trehalose biosynthesis or degradation. This is described here Examples of the active species. Thus, "MP protein function" for one or more of these metabolic pathways Have a role, and / or directly or indirectly affect the one or more production of fine chemicals, raw Production and / or production efficiency. Examples MP protein activity are listed in Table 1. ...

MP nucleic acid molecule of the present invention encoding the protein moiety, preferably the biological activity of MP protein Fraction. As used herein, the term "MP protein portion of the biological activity" mean MP protein containing such a portion, for example, domains / base element, the part capable of catalyzing glutamicum Of one or more amino acid bacteria, vitamins, cofactors, nutritional factors, nucleotides, nucleosides, Or trehalose, the enzymatic reaction in the metabolic pathway or with activity in Table 1. May perform one Activity analysis to determine the protein or its biological activity MP some amino acids can catalysis, Vitamins, cofactors, nutritional factors, nucleotides, nucleosides, or in the metabolic pathway enzyme trehalose Reaction. This method of analysis the skilled person is well known, in the sample of Example 8 Described in detail. ...

Because of the degeneracy of the genetic code, as well as thus obtained, and the present invention can encode the nucleotide MP sequence encoding a protein the same protein, the present invention differs from the present invention further includes The nucleotide sequence (e.g., the sequence of SEQ ID sequence number) (and portions thereof) of the nucleic acid Child. In another embodiment, the isolated nucleic acid molecule of the present invention having such a nucleotide Sequence, the sequence having the sequence encoding the amino acid sequence listed (e.g., an even sequence number) Proteins. Also in another embodiment, the present invention is a nucleic acid molecule encoding the full length glutamic Corynebacterium acid protein, the protein of the present invention the amino acid sequence (sequence number from the sequence table Open reading frame encoding the column number) have sufficient homology. ...

In one embodiment, the present invention is not meant to include a sequence of previously known in the art Sequences, such as those listed in Table 2 or Table 4, in the present invention, before the effective Genbank Sequence, which is skilled in the art can be understood. In one embodiment, the The present invention includes such a nucleotide sequence and amino acid sequence, the nucleotide sequence of the present invention Sequence and amino acid sequence of a certain percentage of the consistency known in the percentage is greater than Sequence (e.g., Table 2 or Table 4 lists the Genbank sequence (or the sequence encoding Protein)) with the nucleotide sequence of the present invention, and the percentage of amino acid sequence identity. For example, The present invention includes and marked as RXA00115 (SEQ ID NO: 185) The nucleotide sequence of a large In and / or at least 40% identity to the nucleotide sequence, and indicated as RXA00131 (SEQ ID NO: 991) The nucleotide sequence of greater than and / or at least% identity to the nucleotide sequence, and And marked as RXA00219 (SEQ ID NO: 345) The nucleotide sequence of greater than and / or at least Consistency 39% of the nucleotide sequence. Skill in the art, listed in Table 4 by checking on At each particular sequence given three highest GPA-calculation of the percentage of matching consistency, as well as by Collected from one hundred percent in the highest GPA-calculated by subtracting the percentage of consistency, you can calculate any of the Percentage of specific sequences present invention low consistency threshold. The skilled person can sense That the percentage is greater than that calculated consistent low field values (e.g., at least 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59% or 60%, preferably at least about 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69% or 70%, more preferably to Less some 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79% or 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 88%, 89% or 90%, or 91%, 92%, 93%, 94%, and even more preferably at least about 95%, 96%, 97%, 98%, 99% Or greater consistency) the nucleic acid and amino acid sequences are also included in the present invention. ...

Skilled in the art will recognize that in addition to the sequence number in the table are listed the serial number MP nucleotide sequence of Corynebacterium glutamicum, the resulting amino acid sequence changes in MP DNA Polymorphisms in certain groups (such as Corynebacterium glutamicum groups) exists. The MP-based Due to genetic polymorphism, the conditions may be due to natural variations in the different individuals in a group Exists. As used herein, the term "gene" and "recombinant gene" refers to coding comprising MP protein open reading frame nucleic acid molecule, preferably Corynebacterium glutamicum MP protein MP proteins. This natural conditions can cause variations typically 1-5% MP nucleotide sequence Changes. Any and all the variations caused due to natural conditions, and does not change the MP protein White functional activity, such nucleotide changes, as well as amino acid polymorphism caused MP, Are within the scope of this invention. ...

Natural variants of the corresponding nucleic acid molecule, and the present invention is Corynebacterium glutamicum MP DNA non-Gu Corynebacterium acid homologues, are disclosed herein can be based with a nucleic acid Corynebacterium glutamicum MP Molecular homology with Corynebacterium glutamicum or a part of a DNA hybridization probes, in strict Cell hybridization according to standard hybridization techniques under conditions isolated. Thus, in another embodiment, Isolated nucleic acid molecule of the present invention, the length of at least 15 nucleotides, under stringent conditions, and containing The nucleotide sequence SEQ ID sequence number nucleic acid hybridization. In other embodiments, the nuclear The length of at least one acid molecule 30,50,100,250 or more nucleotides. As used herein, Herein, the term "hybridizes under stringent conditions" means hybridization and wash describe such a Conditions between the conditions at least 60% homology between the nucleotide sequence of protection Held typical hybrid. Preferably, this condition between the sequence of at least about 65%, more preferably At least about 70% and even more preferably at least about 75 or higher homology, Maintained between typical hybridization Such stringent conditions to be known to the skilled person , He may, Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, N.Y. (1989) ,6.3.1-6 .3.6 found. A preferred, but not limiting Stringent hybridization conditions in 6X sodium chloride / sodium citrate (SSC) were hybridized for about 45 ℃, Then 0.2X SSC, 0.1% SDS at 50-65 ℃ cleaned once or more times. Preferably, the separation The nucleic acid molecule of the invention, the stringent hybridization conditions with the nucleotide sequence of the present invention, hybridization, Equivalent to the naturally occurring nucleic acid molecule obtained. Such as used herein, "naturally occurring" The nucleic acid molecule refers to a nucleotide sequence present in the natural (e.g., encodes a natural protein) An RNA or DNA molecule. In one embodiment, the nucleic acid encoding the native pole glutamicum MP bacterial proteins. ...

Skilled in the art can be further realized that, in addition to the presence of naturally occurring populations of MP sequence variants other than the changes can be introduced by mutation in the nucleotide sequence of the present invention, thus MP result in amino acid sequence encoded changed without changing the MP protein. For example, the nucleotide sequence of the present invention, can be lead to "non-essential" amino acid residues Substituted amino group substituted nucleotides. "Non-essential" amino acid residue refers to a residue, The residue of the MP protein may be wild-type sequence (e.g., the sequence of even-numbered SEQ ID sequence) The change, without changing the MP protein activity, and the "essential" amino acid residue is MP Necessary for protein activity. However, other amino acid residues (e.g., those in the MP activity Results Non-conserved domain structure or just half conserved amino acid residues) may be not necessary for the activity , So you can not change the case of MP activity is changed. ...

Accordingly, another aspect of the present invention, such encoding nucleic acid molecule MP protein, The activity of the MP MP protein contains a non-essential amino acid residue changes. These proteins The amino acid sequence differs from the sequence of the even sequence number sequence, but still maintained at least one here MP activity is described. In one embodiment, the isolated nucleic acid molecule encoding a protein containing The nucleotide sequence of the white matter, wherein the amino acid sequence of the protein with the amino acid sequence of the present invention. At least about 50% homology, and capable of catalyzing the amino acids, vitamins, cofactors, business Raising factors, nucleotides, nucleosides, or trehalose, the enzymatic reaction in the metabolic pathway, or in Table 1 In one or more of the active list. Preferably, the nucleic acid molecule encoding a protein with the sequence table The amino acid sequence of SEQ ID number, there is at least about 50-60% homologous, more preferably with such There are at least about 60-70% sequence homology, even more preferably at least about with such sequences 70-80% ,80-90% ,90-95% homology, most preferably the amino acid sequence of the present invention, there is At least about 96%, 97%, 98%, or 99% homology. ...

In order to confirm the amino acid sequence (e.g., an amino acid sequence of the present invention, with projections Variant form) or both the percentage of nucleic acid sequence homology, most suitable for the purpose of comparison, Sequence comparison of the sequences (for example, to other proteins or nucleic acids with the most appropriate Sequence alignment, can be a protein or nucleic acid sequence into a gap). Then compare Amino acid positions corresponding to amino acid residue, or a nucleic acid nucleotide position. When a sequence (case For example, the amino acid sequence of the present invention a) is a position in the other sequence (e.g., ammonia Acid sequence mutant form) corresponding to the same position as the amino acid residue or nucleotide occupied , The molecules are homologous at that position (i.e., as used herein amino acid or nucleic acid "with Derived "with an amino acid or nucleic acid" identity "is the same.) The percentage between two sequences Points than homology is a sequence of the same number of positions to be sharing function (ie,% identity = Phase With position # / full position # x 100). ...

Isolated protein sequences of the present invention (e.g., the sequence of SEQ ID sequence even) with MP protein source encoding nucleic acid molecule can be through the introduction of the nucleotide sequence of the present invention, One or more nucleotide substitution, insertion, deletion and production, thus the argument in the encoded protein Into one or more amino acid substitutions, insertions, deletions. You can use standard techniques, such as fixed Mutagenesis and PCR-mediated mutagenesis, the nucleotide sequence of the present invention, the introduction of mutation. Preferably, Paul The amino acid substitution is to keep one or more expected in a non-essential amino acid residues carried. "Paul Gatekeeper amino acid substitution "refers to the amino acid residue having a similar side chain of amino acid residues taken Generations. Have similar amino acid residue side chain family is technically specified. These families include, Amino acids having basic side chains (eg, lysine, arginine, histidine), acidic Side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains with ammonia Amino acids (e.g., glycine, aspartic acid, glutamine, serine, threonine, tyrosine, Cysteine), nonpolar side chains (e.g., alanine, valine, leucine Acid, isoleucine, proline, phenylalanine, methionine, tryptophan), a β-branched Side chains (eg, threonine, valine, isoleucine) and aromatic groups Side chains (eg, tyrosine, phenylalanine, tryptophan, histidine). Accordingly, MP protein expected non-essential amino acid residues, preferably by the same side chain family which He amino acid substitution. Further, in another embodiment, the length of the coding sequence, at the MP Or in part, the introduction of random mutations, e.g., by saturation mutagenesis, according to the identification described herein MP activity in a mutant MP activity, screening the obtained mutants. In one sequence table The nucleotide sequence of odd sequence number mutagenesis, the encoded protein can be expressed recombinant protein Active substance can be, for example, using the analysis described herein (see the example in Example 8), to obtain correct Fixed. ...

In addition to the above-described nucleic acid molecule encoding a protein other than the MP, the other of the present invention, Surface also with the separation of antisense nucleic acid molecules related. "Antisense" nucleic acid encoding the protein includes "There Yi "nucleic acid complementary to the nucleotide sequences, such as double-stranded DNA molecule complementary to the coding strand, or with complementary mRNA sequences. Accordingly, an antisense nucleic acid can hydrogen bond with a sense nucleic acid connection. Antisense Nucleic acid can be complementary to the coding strand MP with all, or only some of its complementary. In one embodiment, Solution, antisense nucleic acid molecules encoding the MP protein coding strand of a nucleotide sequence of the "coding Zone "antisense term" coding region "refers translated into amino acid residues containing codon Nucleotide sequence regions (e.g., SEQ ID NO.1 (RAX 02229) including the entire coding region 1-825 nucleotides). In another embodiment, the antisense nucleic acid molecule encoding the MP A nucleotide sequence encoding an antisense chain. The term "noncoding region" refers to the coding region on both sides not turn Amino acids into the 5 'and 3' sequences (i.e., 5 'and 3' untranslated regions). ...

Considering published here MP coding strand of the coding sequence (e.g., the sequence listed in the table Odd serial number sequence), an antisense nucleic acid of the invention according to the Watson and Crick base pairing The design rules. Antisense nucleic acid molecules can be used with the entire coding region of MP mRNA interaction Fill, but more preferably is such that oligonucleotide, the oligonucleotide is only with MP mRNA Coding region or part of the non-coding region antisense. For example, the antisense oligonucleotide can MP mRNA translation initiation and complementary to a region near the position. For example, antisense oligonucleotides Length can be 5,10,15,20,25,30,35,40,45 or 50 nucleotides. This Invention, antisense nucleic acid molecules can be used procedures known in the art, or by chemical synthesis Construction of enzymatic ligation reactions. Using naturally occurring nucleotides or various of the modified core Nucleotide, chemically synthesized antisense nucleic acid (eg antisense oligonucleotides), which has been modified nuclear Nucleotide, is to increase the biological stability of the molecule, or in order to increase the sense and antisense nuclear Acid double helix formed between the physical stability and design, for example, phosphorothioate derivatives may be used Matter and acridine substituted nucleotides. Can be used to generate an antisense nucleic acid modified nucleotides examples Including 5 - fluorouracil and 5 - bromo-uracil and 5 - chloro uracil and 5 - iodo uracil, hypoxanthine, Xanthine 4 - acetyl cytosine, 5 - (carboxy-hydroxy-methyl) uracil, 5 - carboxymethyl-aminomethyl -2 - Thiouracil, 5 - carboxymethyl-aminomethyl-uracil, dihydrouracil, beta-D-galacto- Inosine, N6-isopentyl-adenine, 1 - methylguanine, 1 - methyl-inosine, 2,2 - dimethyl guanine Methotrexate 2 - methyl adenine 2 - methylguanine 3 - methylcytosine, 5 - methylcytosine, N6- Adenine, 7 - methylguanine, 5 - methyl-amino-methyl uracil, 5 - methoxy-aminomethyl-uracil -2 - thiouracil, beta-D-mannosyl queosine, 5'-methoxy-carboxy methyl uracil, 5 - methoxy-uracil 2 - methylthio-N6-isopentyl-adenine, uracil -5 - oxo acetic acid (v), wybutoxosine, pseudo uracil, queosme, 2 - thio cytosine and 5 - methyl-2 - thio uracil Pyridine 2 - thiouracil 4 - thiouracil, 5 - methyluracil, uracil -5 - oxo acetic acid methyl Ester, uracil -5 - oxo acetic acid (v), 5 - methyl-2 - thiouracil, 3 - (3 - amino-3-N-2- Carboxypropyl) uracil, (acp3) w, and 2,6 - diamino-purine. In addition, the antisense nucleic acid can be Biosynthesis using an expression vector wherein the nucleic acid is subcloned in an antisense orientation into the expression vector (I.e., transcribed from the inserted nucleic acid RNA, relative to the insertion of the nucleic acid is in an antisense orientation, There are further described in the following section). ...

The antisense nucleic acid molecule of the invention is typically applied to a cell or generated in situ from MP and their encoded proteins with cellular mRNA and / or genomic DNA hybridization or Junction Together, thereby inhibiting the expression of proteins, for example, inhibit transcription and / or translation. Hybridization can be Conventional nucleotide complementarity to form a stable duplex, or, for example, when the antisense nucleic acid molecule When combined with the DNA double helix, it happens the major groove of the double helix specific interactions. Antisense molecules can be As to be modified, so that the molecule with a specific cell surface receptors or antigens expressed Specific binding, for example, antisense nucleic acid molecules binding to the polypeptide or antibody, the antibody and fine Cell surface receptor or antigen. Antisense nucleic acid molecule can also be used here to describe a carrier delivery To the cells. In order to obtain sufficient concentration of intracellular antisense molecules, such vectors are preferred, In the support that the antisense nucleic acid molecule is placed in a prokaryotic, eukaryotic or viral promoter control Beneath. ...

And in another embodiment, the present invention is a nucleic acid molecule is antisense ribozyme. Ribozyme is a reminder Type of RNA molecules with ribonuclease activity, can cut single-stranded nucleic acids, such as mRNA, It has a single-stranded nucleic acid complementary to a region. Thus, ribozymes (eg, hammerhead ribozymes (described in Haselhoff and Gerlach (1988) Nature 334:585-591)) can be used for catalytic cleavage MP mRNA transcript, thereby inhibiting the MP mRNA translation. For MP nucleic acid molecules encoding Is specific ribozyme, may be based on published herein MP DNA nucleotide sequence (i.e., SEQ ID NO: 1 (RAX 02229)) to design. For example, you can build genus Tetrahymena L-19 IVS RNA The derivative of the nucleotide sequence of the active site of the cut and the MP-encoding mRNA nucleoside Acid sequence is complementary. See, e.g., Cech et al.USPatent No.4, 987,071, and Cech et al.U.S.Patent No.5, 116,742. In addition, MP mRNA can be used for RNA points Sub-library was screened with specific catalytic activity of the ribozyme RNA. See, e.g., Bartel, D.and Szostak, J.W. (1993) Science 261:1411-1418. ...

B. recombinant expression vectors and host cells

Another aspect of the present invention, the carrier is preferably a protein containing the coding MP (or its Part) a nucleic acid expression vector. As used herein, the term "vector" refers to the ability even Then the other nucleic acid, and its nucleic acid molecule for transport. One type of vector is a "plasmid", Plasmids are circular double stranded DNA loop means, which is connected to an additional DNA fragment. Another type of The vector is a viral vector, wherein additional DNA segments can be connected to the viral genome. Some These vectors can be introduced in their host cells for autonomous replication (e.g., a bacterial Bacterial origin of replication vectors, and episomal mammalian vectors). Other vectors (e.g., Non-episomal mammalian vectors) will host cell upon introduction into the host cell's genes The group, thereby replicated along with the host genome. In addition, some carriers can be connected to guide Then the expression of genes. These vectors are referred to herein as "expression vectors." In short, recombinant DNA technology Technique is often used in form of a plasmid expression vector. In this description, "plasmid" and "vector" Can be used interchangeably, as the plasmid is the most commonly used form of vector. However, the present invention intends to package These include other forms of expression vectors, such as viral vectors (eg, replication defective reverse Viruses, adenoviruses and adeno-associated viruses), which have the same function. ...

Recombinant expression vectors of the invention comprise a nucleic acid of the invention, the nucleic acid in a host cell to fit Co-expressed in the form of nucleic acids, which means that the recombinant expression vectors contain one or multiple tone Section sequences that are based on the host cell selected for expression, and they are viable even Received a nucleic acid sequence to be expressed. In the recombinant expression vector, "workable connection" mean Refers to a nucleotide sequence of interest and regulatory sequences to allow expression of the nucleotide sequence of the way Connection (for example, in vitro transcription / translation system, or in a vector is introduced into a host cell In). The term "regulatory sequence" is meant to include promoters, enhancers and other expression control Elements (e.g., polyadenylation signal). Such regulatory sequences in, for example, Goeddel; Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, CA (1990) is described. Regulatory sequences, including those in many types of host cells, Constitutive expression of a nucleotide sequence of instructions in sequence, and that in some host cells, guide The expression of the nucleotide sequence of sequence. Preferred regulatory sequences are, for example, such as cos-, tac-, trp-, tet-, trp-tet-, lpp-, lac-, lpp-lac-, lacI ...^q-，T7-，T5-，T3-，gal-，trc-，ara-，SP6-，arny-， SPO2-，λ-P _RRecombinant expression vectors of the invention comprise a nucleic acid of the invention, the nucleic acid in a host cell to fit Co-expressed in the form of nucleic acids, which means that the recombinant expression vectors contain one or multiple tone Section sequences that are based on the host cell selected for expression, and they are viable even Received a nucleic acid sequence to be expressed. In the recombinant expression vector, "workable connection" mean Refers to a nucleotide sequence of interest and regulatory sequences to allow expression of the nucleotide sequence of the way Connection (for example, in vitro transcription / translation system, or in a vector is introduced into a host cell In). The term "regulatory sequence" is meant to include promoters, enhancers and other expression control Elements (e.g., polyadenylation signal). Such regulatory sequences in, for example, Goeddel; Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, CA (1990) is described. Regulatory sequences, including those in many types of host cells, Constitutive expression of a nucleotide sequence of instructions in sequence, and that in some host cells, guide The expression of the nucleotide sequence of sequence. Preferred regulatory sequences are, for example, such as cos-, tac-, trp-, tet-, trp-tet-, lpp-, lac-, lpp-lac-, lacI ..._LSuch promoters, these promoters is preferred for use in bacteria. Additional regulatory sequences are, for example, promoters of yeast and fungi, such as ADC1, MFα, AC, P-60, CYC1, GAPDH, TEF, rp28, ADH, promoters of plants, for example, CaMV/35S, SSU, OCS, lib4, usp, STLS1, B33, nos or ubiquitin-or phaseolin-promoter. Synthetic promoters can also be used. For those skilled in the art is aware that The design of the expression vector depends on these factors: for the selection of transformed host cells, the desired protein The expression level of the white matter. The expression vector of the present invention can be introduced into a host cell, thereby generating the Described at the protein encoded by the nucleic acid or polypeptides, including fusion proteins or peptides (for example, If, MP protein, MP mutant forms of protein, fusion proteins, etc.). ...

Can design a recombinant expression vector of the present invention, for in prokaryotic or eukaryotic cells expressing MP proteins. For example, MP gene can be expressed in the following cells, such as Corynebacterium glutamicum which Like bacterial cells, insect cells (using baculovirus expression vectors), yeast and other fungi Cells (see Romanos, MAet al. (1992) "Foreign gene expression in yeast: a review ", Yeast 8:423-488; van den Hondel, CAMJJet al. (1991)" Heterologous gene expression in filamentous fungi "in: More Gene Manipulations in Fungi, JW Bennet & L.L.Lasure, eds., P.396-428: Academic Press: San Diego; and van den Hondel, CAMJJ & Punt, PJ (1991) "Gene transfer systems and vector development for filamentous fungi, in: Applied Molecular Genetics of Fungi, Peberdy, JFet al., Eds., P.1-28, Cambridge University Press: Cambridge) or multicellular algae Plant cells (see Schmidt, R.and Willmitzer, L. (1998) High efficiency Agrobacterium tumefaciens-mediated transformation of Arabidopsis thaliana leafand cotyledon explants "Plant Cell Rep. :583-586), or breast-feeding Animal cells. Appropriate host cell in Goeddel, Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, CA (1990) in progress Step exposition. In addition, recombinant expression vectors can be transcribed and translated in vitro, for example using T7 start Promoter regulatory sequences and T7 polymerase. ...

Prokaryotic expression of proteins in cells, is often containing constitutive or inducible promoters The carrier, the fusion protein of these promoters instructions or a non-fusion protein expression. Protein fusion vector encoding a certain number of amino acids added, usually in the recombinant protein The amino terminus, but may also be the C-terminus, or the appropriate area of the fusion protein. The financial Typical co-carrier has three purposes: 1) to increase expression of recombinant protein; 2) increase the recombinant protein White matter solubility; and 3) as ligand affinity purified, fusion protein purification help. In the fusion Expression vector, the protein cleavage site is often introduced into the recombinant fusion protein portion and Junction, so that the purification of the fusion protein, the fusion protein can be part of the recombinant Separated. Such enzymes, and their cognate recognition sequences, include Factor Xa, thrombin and intestinal Kinases. ...

Typical fusion expression vectors include pGEX (Pharmacia Biotech Inc; Smith, DB and Johnson, KS (1988) Gene 67:31-40), pMAL (New England Biolabs, Beverly, MA) and pRIT5 (Pharmacia, Piscataway, NJ), respectively with the target Recombinant fusion protein glutathione S-transferase (GST), maltose E binding protein, or Protein A. In one embodiment, MP protein coding sequence is cloned into an expression vector pGEX The body, resulting in a fusion protein encoded vector which from N-terminus to C-terminus, including, GST-thrombin cleavage site-X protein. Fusion protein can be used glutathione - agarose tree Fat, by affinity chromatography. Separated with GST protein recombinant MP, can be prepared by coagulation Cleavage of fusion proteins of blood obtained. ...

Suitable inducible non-fusion E. coli expression vectors include, pTrc (Amann et al., (1988) Gene 69:301-315), pLG338, pACYC184, pBR322, pUC18, pUC19, pKC30, pRep4, pSH1, pSH2, pPLc236, pMBL24, pLG200, pUR290, pIN-III 113-B1, λgt11, pBdC1, and pET 11d (Studier et al., Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, California (1990) 60-89; and Pouwels et al., Eds. (1985) Cloning Vectors. Elsevier: New York IBSN 0 444 904018). pTrc vector target gene expression, by Depends on the hybrid trp-lac fusion promoter host RNA polymerase transcription. pET 11d vector Target gene expression, depending on the co-expression of viral RNA polymerase (T7 gn1) T7-mediated gn10-lac fusion promoter transcription. The viral polymerase from the host strain BL21 (DE3) or By HMS174 (DE3) reside λ phage provides the phage containing the lacUV5 Kai Mover T7 gn1 under the transcriptional control genes. For other types of bacterial transformation, can be selected together A suitable carrier. For example, it is known plasmid pIJ101, pIJ364, pIJ702 and pIJ361 transformed Streptomyces Is valid, and the plasmid pUB110, pC194, or pBD214 rod species for transformation. Have Contribute to the genetic information into Corynebacterium include several plasmids pHM1519, pBL1, pSA77 Or pAJ667 (Pouwels et al., Eds. (1985) Cloning Vectors, Elsevier: New York IBSN 0 444 904018). ...

In another embodiment, MP protein expression vector is a yeast expression vector. Yeast S. cerivisae for expression vectors include, pYepSec1 (Baldari, et al., (1987) Embo J.6 :229-234), 2μ, pAG-1, Yep6, Yep13, pEMBK Ye23, pMFa (Kurjan and Herskowitz, (1982) Cell 30:933-943), pJRY88 (Schultz et al., (1987) Gene 54:113-123), and pYES2 (Invitrogen Corporation, San Diego, CA). Suitable for building in other fungi, such as filamentous fungi, the use of a carrier vector And methods, including those described in the following literature: van den Hondel, CAMJJ & Punt, PJ (1991) "Gene transfer systems and vector development for filamentous fungi, in: Applied Molecular Genetics of Fungi, JFPeberdy, et al., eds., p.1-28, Cambridge University Press: Cambridge, and Pouwels et al., eds. (1985) Cloning Vectors, Elsevier: New York IBSN 0 444 904018). ...

In another embodiment, the present invention is in the single cell protein MP plant cells (for example, Such as algae) expression, or higher plants (e.g., seed plants, such as crop plants) Plant cells. Examples of plant expression vectors include those described in the following literature: Becker, D., Kemper, E., Schell, J.and Masterson, R. (1992) "New plant binary vectors with selectable markers located proximal to the left border ", Plant Mol.Biol. 20:1195-1197; and Bevan, MW (1984) "Binary Agrobacterium vectors for plant transformation ", Nucl.Acid.Res.12 :8711-8721, including pLGV23, pGHlac, pBIN19, pAK2004, and pDH51 (Pouwels et al., eds. (1985) Cloning Vectors. Elsevier: New York IBSN 0 444 904018).

Also in another embodiment, the present invention is a nucleic acid used in the feeding mammalian expression vector Mammalian cells. Examples of mammalian expression vectors include pCDM8 (Seed, B. (1987) Nature 329:840) and pMT2PC (Kaufman et al. (1987) EMBO J.6: 187-195). The control expression vector was used in mammals, often by the disease Regulating toxic elements to offer. For example, commonly used promoters derived from polyoma, Adenovirus 2, cytomegalovirus Cytomegalovirus and simian virus 40. For other prokaryotic cells and eukaryotic cells are suitable for expression of the body System, see Sambrook, J., Fritsh, EF, and Maniatis, T.Molecular Cloning: A Laboratory Manual.2nd, ed.Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989, 16 chapters and 17 chapters. ...

In another embodiment, the recombinant mammalian expression vector capable of directing specific details Preferred cell types in the expression of the nucleic acid (e.g., tissue specific regulatory elements are used to express the nuclear Acid). Tissue specific regulatory elements are known in the. Suitable tissue-specific promoter Examples of the child including but not limited, the albumin promoter (liver-specific; Pinkert et al. (1987) Gene Dev.1 :268-277), lymphoid-specific promoters (Calame and Eaton (1988) Adv. Immunol.43 :235-275), T-cell receptor specific promoter (Winoto and Baltimore (1989) EMBO J.8 :729-933) and immunoglobulins specific promoter (Banerji et al. (1983) Cell 33:729-740; Queen and Baltimore (1983) Cell 33:741-748), Neuron-specific promoters (e.g., neurofilament promoter; Byrne and Ruddle (1989) PANS 86:5473-5477), pancreas-specific promoters (Edlund et al. (1985) Science 230: 912-916), and mammary gland-specific promoters (e.g., milk whey promoter; USPatent No. 4,873,316 and European Application Publication No.264, 166). Also include hair Sterile regulated promoters, such as murine hox promoters (Kessel and Gruss (1990) Science 249:374-379) and α-fetoprotein promoter (Campes and Tilghman (1989) Genes Dev. 3:537-546). ...

The present invention also provides a DNA molecule of the invention comprising a recombinant expression vector, the DNA Molecular cloning antisense orientation in the expression vector. In other words, DNA molecules can operability The regulator connected to the following sequence, which allows the MP mRNA antisense RNA molecule Expression (by transcription of the DNA molecule) of the method. Can be selected in a variety of cell types that Guiding a continuous expression of the antisense RNA molecule, a regulatory sequence, for example, viral promoters and / or enhanced Son, or you can choose guided continuous, tissue-specific or cell type-specific antisense RNA Expression control sequence, a regulatory sequence. Antisense expression vector can be recombinant plasmid phage Grain form or attenuated virus in which antisense nucleic acids under the control of regulatory regions and efficient Generation, its activity by introducing the carrier to determine the type of cell. On the use of antisense gene Regulation of gene expression, can see Weintraub, H.et al., Antisense RNA as a molecular tool for genetics analysis, Review-Trends in Genetics, Vol.1 (1) 1986. ...

The host cell can be any prokaryotic or eukaryotic cell. For example, MP protein may like Corynebacterium glutamicum such as bacterial cells, insect cells, yeast or mammalian cells Animal cells (e.g. Chinese rat ovary cells (CHO) or COS cells) cells. Other Suitable host cells for the skilled staff is well known. The present invention can be used Nucleic acid and protein molecules of the host cell affinity Corynebacterium glutamicum microorganism listed in Table 3 Out.

Vector DNA by conventional transformation or transfection techniques, the introduction of a prokaryotic or eukaryotic Cells. As used herein, the term "transformation" and "transfection" means that there is a variety of skills Domain is well known, the exogenous nucleic acid (e.g., linear DNA or RNA (e.g., linear vector Genes alone or without the vehicle structure)) or in the form of a nucleic acid vector (e.g., Plasmid, phage, phagemid, phagemids, transposons or other DNA) into a host cell Technology, including calcium phosphate or calcium chloride coprecipitation, DEAE-dextran mediated transfection, Lipofection, or fax holes. Host cells transformed or transfected with the appropriate way, you can Sambrook, et al. (Molecular Cloning: A Laboratory Manual.2nd, ed .., Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989), and other laboratory manuals found. ...

Known for stable transfection of mammalian cells, rely on the use of expression vectors and transfection Technology, only a small portion can be exogenous DNA into its own genome. In order to identify And filter these integrants, coding selection marker (eg, antibiotic resistance) gene through Often associated with the gene of interest is introduced into the host cells. The preferred selection markers include those that Fu To the drug resistance marker, such as G418, hygromycin and methotrexate. Encoding a selection marker nuclear Acid, the MP protein may be the same vector is introduced into a host cell, or contained in a separate Body into a host cell. The nucleic acid is introduced into the stably transfected cells, drug screening can be used Identification (for example, combined with the selection marker gene can survive cells, while other cells die). ...

To create a homologous recombinant microorganism comprising at least part of the MP gene preparation of the carrier, the Gene has a deletion, addition or substitution, thereby changing, for example, functional damage, MP gene. Preferably the MP gene is Corynebacterium glutamicum, but it can also be derived from the same bacterial phylogenetic Source material, even from mammalian, yeast or insect. In a preferred embodiment, Design vector, such that homologous recombination according to the endogenous gene is functional MP destroyed (i.e., not Encoding functional proteins; also called "knockout" vector). In addition, the carrier can be designed such that According to homologous recombination, the endogenous gene is mutated or MP change, but still encoding protein White matter (for example, changing the upstream regulatory region, thereby changing the expression of endogenous genes MP). In Homologous recombination vector, the altered portion of the MP gene, at its 5 'and 3' ends are connected to the side MP nucleic excess, so that homologous recombination can occur in the vectors carrying the foreign gene and the MP MP endogenous microbial genes. MP connected to the side of excess nucleic acids long enough Degrees endogenous gene can be successful homologous recombination. Typically, the vector containing thousands Base side chain DNA (5 'and 3' ends) (see, e.g., Thomas, KR, and Capecchi, MR (1987) Cell 51:503 for a description of homologous recombination vectors). Introduction of micro-organisms (such as fax hole) and cell carriers, selecting those which lead The MP gene into the endogenous MP gene, using techniques known in the art for homologous recombination. ...

In another embodiment, the host cell is destroyed endogenous MP gene (e.g., By homologous recombination or other genetic means known in the art) such that the protein product Expression can not occur. In another embodiment, the host cells or introduced in the endogenous MP gene, via one or more point mutations, deletion or inversion and change, but still coding MP protein function. And in another embodiment, the microorganism one or more genes MP A regulatory region (eg, promoter, repressor or inducer) is changed (for example, by Missing, cut, inverted or point mutation) such that gene expression is regulated MP. Ability Field technicians can recognize that contain more than one of the MP gene and protein modification places Primary cells, the use of the present invention can be easily produced, these cells are also included in the present Ming. ...

C. Separation of MP protein

Aspect of the invention relates to an isolated part of MP and biological activity of the protein. "Sub- From "or" purified "protein or biologically active portion which, when the use of recombinant DNA technology Production technique basically no cellular material, while chemical synthesis of chemical precursors or substantially no Other chemicals. The term "substantially free of cellular material" includes such MP protein preparation, Wherein the protein is produced from natural or recombinant cells of the protein fraction isolated Out. In one embodiment, the term "substantially free of cellular material" includes preparation containing at Least about 30% (dry weight) of MP protein (herein also referred to as "pollution protein") of the MP Protein, more preferably contains less than about 20% of non-MP protein, even more preferably containing less Approximately 10% of non-MP protein, and most preferably contains less than about 5% of non-MP proteins. When the MP protein or a biologically active portion of rescheduled generated, is preferably substantially free of culture medium, Producing a protein that is less than the volume of the medium approximately 20%, preferably less than 10%, most preferably Less than about 5%. The term "substantially free of chemical precursors or other chemicals" includes such The MP protein preparation, wherein the protein is involved in protein synthesis from chemical precursors or other Isolated chemicals. In one embodiment, the term "substantially free of chemical precursors or By other chemicals "includes a solution containing at least about 30% (dry weight) of chemical precursors or non- MP MP protein chemicals, more preferably contains less than about 20% chemical precursors or non- MP chemical substances, even more preferably containing less than about 10% chemical precursors or non-MP of Chemicals, and most preferably contains less than about 5% chemical precursors or non-MP chemicals. In A preferred embodiment, the isolated protein, or part of its biological activity, does not contain to MP protein obtained from the same organism contamination proteins. This protein is typically from heavy Group was produced, for example, such as a microorganism of Corynebacterium glutamicum Corynebacterium glutamicum MP eggs White recombinant expression. ...

MP isolated protein of the invention or a part of its biological activity, capable of catalyzing the amino acids, Vitamins, nutritional factors, nucleotides, nucleosides, or trehalose in the metabolic pathway or enzymatic reactions Having one or more listed in Table 1 activity. In a preferred embodiment, the protein Containing such substance or part of the amino acid sequence, the sequence of the amino acid sequence of the present invention (Example For example, the sequence SEQ ID sequence even in a sequence) there is sufficient homology such that the egg White matter or part of its biological activity, capable of catalyzing the amino acids, vitamins, cofactors, nutrition because Son, nucleotides, nucleosides, or trehalose in the enzymatic reaction pathway. Part of the protein, preferably Select refers to biologically active portion as described herein. In another preferred embodiment, the present invention is The MP protein has an even number in the sequence table lists the amino acid sequence SEQ ID NO. In another In a preferred embodiment, MP from the nucleotide sequence encodes a protein having the amino acid sequence of the nuclear Nucleotide sequence of the nucleotide sequence of the present invention (e.g., SEQ ID sequence number in a sequence Sequences) hybridize under stringent conditions such as hybridization. In another preferred embodiment, MP protein has a nucleotide sequence from such an amino acid sequence encoded by the nucleotide sequence of the A nucleic acid sequence of the invention or parts thereof, at least about 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, or 60% homology, preferably at least about 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69% or 70% homology, more preferably There are at least about 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79% or 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 88%, 89% or 90%, or 91%, 92%, 93%, 94%, and even more preferably at least about 95%, 96%, 97%, 98%, 99% or higher homology. Between the values quoted above range, or the consistency between the values (for example, If the consistency of 70 to 90%, or 80-95% identity), but also intentionally included in the present invention. For example, intentionally containing such a consistency range, the range is the upper limit of the above-cited And / or the lower limit value combination. MP Preferred proteins of the present invention is also preferred that at least one here has MP activity is described. For example, one preferred MP protein of the invention contains such nucleotides The amino acid sequence encoded by the sequence, the nucleotide sequence of the nucleotide sequence of the present invention, hybridization, for example, Hybridizes under stringent conditions as, and to catalyze the sequence of amino acids, vitamins, cofactors, Nutritional factors, nucleotides, nucleosides, or trehalose, the enzymatic reaction in the metabolic pathway, or having a Species listed in Table 1 or more of the active. ...

In other embodiments, MP protein with the amino acid sequence of the present invention (e.g., Sequence Table Even a sequence SEQ ID sequence) has sufficient homology with the present invention and the amino group Acid sequence of the functional activity of the protein, as described in detail above, as Part I, its amino acid Sequence of natural change or mutation varies. Thus, in another embodiment, MP protein is such that the protein, which has the amino acid sequence of the complete amino acid sequence of the present invention. Column, at least about 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, or In 60% homology, preferably at least about 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69% or 70% homology, more preferably at least about 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79% or 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 88%, 89% or 90%, or 91%, 92%, 93%, 94%, and even More preferably to at least about 95%, 96%, 97%, 98%, 99% or higher homology, and And having at least one MP activity as described herein. Ranged between the values quoted above range, or By consistency value (for example, 70-90% identity or 80-95% identity), but also intentional Included in the present invention. For example, intentionally containing such a consistency range, these ranges The above-cited upper and / or lower limit value combination. In another embodiment, the present invention relates to Such length of the Corynebacterium glutamicum protein, the protein amino acid sequence of the present invention. Have sufficient homology. ...

MP portion of the biological activity of proteins containing such a polypeptide, the polypeptide comprising protein derived from MP Amino acid sequence, for example, the even sequence or amino acid sequence of SEQ ID MP homology with the amino acid sequence of the protein portion contains more than length or MP Full-length protein homologous proteins MP fewer amino acids, and exhibit at least one MP protein Activity. Typical biologically active portions (peptides, e.g., amino acids in length such as 5,10,15,20, 30,35,36,37,38,39,40,50,100 or more peptides) having at least one comprises a MP domain of protein activity or primitives. Alternatively, other biologically active portion, wherein the protein Other parts have been removed, can be prepared by recombinant techniques and identification of the one described herein Kind or variety of activity. Preferred biological active portion MP protein containing one or more The selection of the biologically active domains / base element or part thereof. ...

The present invention also provides a chimeric protein or a fusion protein MP. As used herein, MP "inlay Co-protein "or" fusion protein "containing polypeptide operatively linked to a non-MP MP on the multi- Peptides. "MP polypeptide" refers to an amino acid sequence containing a MP related polypeptide, and "non-MP protein" Refers to a protein containing such a polypeptide related to the amino acid sequence of the protein with the MP protein not A basic homology, for example, from the same or different organisms and different proteins MP Proteins. In the fusion protein, the term "operatively linked" is meant, MP protein And between each of MP protein in-frame fusion. Non MP polypeptide can be fused to the MP N-terminus of the polypeptide or the C-terminus. For example, in one embodiment, the fusion protein is DST-MP fusion protein, wherein the MP sequence fused to the C-terminal GST sequence. The fusion protein White matter helps recombinant protein purification MP. In another embodiment, the fusion protein is In the N-terminal signal sequence with heterologous proteins MP. In certain host cells (e.g. mammalian Was the host cell), by using a heterologous signal sequence can increase the MP protein expression and / or Secretion. ...

Preferably, the chimeric protein of the invention or a fusion protein produced by standard recombinant DNA techniques Health. For example, according to conventional techniques, a DNA sequence encoding a different polypeptide fragments are connected in-frame Together, for example, the use of blunt ends or staggered ends the end of the connection, the use of restrictive Enzyme digestion to provide appropriate end use of cohesive ends blunted ends as appropriate, so that Treated with alkaline phosphatase to avoid undesirable connections and the use of enzymatic connection. In another In one embodiment, using conventional techniques including automated DNA synthesizer fusion gene were synthesized. Further, the anchor primer can be used for PCR amplification of gene fragments, can be increased two anchor primer Between consecutive gene fragments complementary protruding end, a continuous gene can subsequently be annealed and re-expansion Increase resulting chimeric gene sequence (see, for example, Current Protocols in Molecular Biology, eds.Ausubel et al.John Wiley & Sons: 1992). In addition, many have been compiled Code fusion part (e.g. GST polypeptide) expression vectors are commercially available. MP-encoding nucleic acid Can be cloned into such an expression vector such that part of the in-frame fusion to connect to the MP protein White on. ...

MP homologues proteins produced by mutation, such as a discontinuity point MP mutant protein Or cut. As used herein, the term "homologue" refers to a variant form of MP protein, They can be used MP protein agonists or antagonist activity. MP protein agonists MP protein having substantially the same or part of the biological activity. MP protein antagonists may MP inhibition of protein naturally present in the form of one or more of the active, for example, by containing the MP MP proteins downstream or upstream member system competitive binding. Accordingly, the present invention is glutamic Corynebacterium MP acid protein and its homologues, you can adjust one or more pieces Carbohydrate transport pathways Activity, or to adjust the microorganism MP proteins play a role in intracellular signal transduction pathways Activity. ...

In other embodiments, MP protein homologue mutant protein can be screened MP The combinatorial library of the body, such as cut mutants to identify agonists or antagonists of protein activity MP Sex. In one embodiment, MP library diversity variant nucleic acid level by a combination of Mutations generated by the diversity of the gene library encoding. MP library diversity of variants can pass However, for example, the mixture of synthetic oligonucleotides into gene sequences enzymatic connected such that the latent The MP set of degenerate sequences as a single polypeptide sequence set MP or which contains more Large fusion proteins (eg for phage display) of the collection, can be expressed. Are each Methods can be used to be a simple and oligonucleotide sequences potentially MP library congeners. Can With automatic DNA synthesizer using degenerate gene sequence for chemical synthesis, and the synthetic gene is connected Receiving a suitable expression vector. The use of degenerate set of genes, allowing to provide encoding mixed Potential sequence set to be the entire sequence of MP. Degenerate oligonucleotide synthesis method is technically Are known (see, for example, Narang, SA (1983) Tetrahedron 39:3; Itakura et al. (1984) Annu.Rev.Biochem.53: 323; Itakura et al. (1984) Science 198:1056; Ike et al. (1983) Nucleic Acid Res.11: 477). ...

In addition, protein fragments encoding a library MP, MP can be used to produce fragments diversity group Body, the group used to screen and select MP protein homologue. In one embodiment, the series Library code sequence fragments can be generated that produce only about one cut per molecule strip Conditions, with a nucleic acid sequence encoding the enzyme treatment MP double-stranded PCR fragment of denatured double-stranded DNA, complex To form a double-stranded DNA of the DNA, the double-stranded DNA may include a cutout from a different shape of the product Into the sense / antisense pairs, re-formation of the double helix by S1 nuclease treatment to remove single-stranded Portion, and the resulting fragment library into an expression vector to connect. By this method, may be To give the N-terminal coding, C-terminus and different sizes MP protein middle fragment expression library. ...

Filter or cut from the point mutation resulting in a gene product of a combinatorial library of many techniques And the screening selected cDNA library having the technical characteristics of the gene product, is technically Known. These techniques are suitable combinations of mutated homologue by the MP gene library obtained by fast Speed filtering. Screening large gene pool of the most widely used technology that can be used for high yield analysis, Including the genomic library cloned into replicable expression vectors, transformed with the resulting library of vectors containing Body library, as well as in combination gene expressed under certain conditions, under conditions in which detection of the desired activity Code is detected helps carrier of the gene product of the separation. Regression ensemble mutation (REM), a Kind of library functions to increase the mutant frequency of new technologies, along with screening assays for the identification of MP homologues (Arkin and Yourvan (1992) PANS 89:7811-7815; Delgrave et al. (1993) Protein Engineering 6 (3) :327-331). ...

D. Application of the present invention and methods

Nucleic acid molecules described herein, proteins, protein homologues, fusion proteins, primers, Vectors and host cells that can be applied to one or more of the following: identification of Corynebacterium glutamicum Coli and affinity microorganisms; draw phylogenetic organism Corynebacterium glutamicum genome; identification And positioning the sequence of interest Corynebacterium glutamicum; Evolution; determine protein function must MP Area required; MP protein activity regulation; MP activity regulation pathway; desired compound, for example, fine Chemical substances produced by cells of the regulation.

MP nucleic acid molecule of the present invention has various uses. First, they can be used to identify a Whether the organism Corynebacterium glutamicum organism or its close relatives. They can also be used to identify mixed Total microbial population or the relatives Corynebacterium glutamicum organisms exists. The present invention provides Many gene of Corynebacterium glutamicum nucleic acid sequence; under stringent conditions, with over glutamate Corynebacterium specific gene probes, the probe from a single or mixed microbial culture of the base was extracted Genomic DNA, can determine the existence of the organism. Although Corynebacterium glutamicum is a non-induced Disease resistance, but it is associated with the pathogenic species such as Corynebacterium diphtheria. Corynebacterium diphtheria diphtheria Pathogenic source, Diphtheria is a rapid development of acute, fever, infections, it involves local symptoms And system pathologies. Have this disease, localized lesions of the upper respiratory tract, including epithelial cells and Necrotic cell injury; bacteria secrete toxins, toxins from the lesions spread to the end of the body vulnerable Shoot tissue. These organizations include the heart, muscles, peripheral nerves, adrenal gland, kidney, liver and Spleen, in which protein synthesis is inhibited due caused by degenerative changes which would lead to the disease System disease symptoms. Diphtheria in many parts of the world to maintain a high incidence of these regions including Africa, Asia, Eastern Europe and the former Soviet Union, independent state. From 1990 onwards, in the latter two regions diphtheria-holding Continued popularity, leading to at least 5,000 people were killed. ...

The nucleic acid and protein molecules of the present invention can also be used in a specific area of genomic markers. This Genome only in the drawing useful, and can be used for protein function Corynebacterium glutamicum Research. For example, Corynebacterium glutamicum in order to identify specific DNA-binding proteins bind to the gene Groups area, can digest Corynebacterium glutamicum genome, the fragment was incubated with the DNA-binding proteins. Binding protein fragments can be further treated with a nucleic acid molecule probe of the present invention, it is preferable to use and easy Detectable label; these nucleic acid molecules combined with genomic fragments, fragments can be positioned in glutamate Corynebacterium location on the genome, and, when performed multiple times with different enzymes operation, Helps to quickly identify the proteins bind to the nucleic acid sequence. Further, the present invention is a nucleic acid molecule With the phylogenetic species have sufficient homology to the nucleic acid molecules that can be used as building Genetic fine Bacterial genome marker, such as Brevibacterium. ...

MP nucleic acid molecules of the present invention and can be used to study protein structure and evolution. The present invention The carbohydrate molecules involved pickup system is used for a variety of bacteria; the present invention by comparison Nucleic acid molecule and those in other organisms sequence encoding nucleic acid molecule sequence similarity, may Organisms have evolved estimated correlation. Similarly, this comparison allows the estimation of conserved sequence regions and Non-conserved sequence regions, which could help to identify the protein in the region required for enzyme function. This determination of the type of value for protein engineering, and may indicate the eggs White matter can be tolerated without loss of function mutations. ...

The present invention also provides methods of screening protein activity can be adjusted MP molecular methods, these molecules Either by itself or with the protein substrate interactions, or the spouse of the MP protein body junction Together, or by adjusting the nucleic acid molecule of the present invention, MP to regulate transcription or translation of proteins MP Activity. In this method, the expression of one or more MP proteins of microorganisms, with one or Contact with a variety of test compounds and to assess each test compound or protein activity for MP The role of expression.

When from Corynebacterium glutamicum culture in large scale fermentation desired fine chemical separation Materials are amino acids, vitamins, cofactors, nutritional factors, nucleotides, nucleosides, or trehalose, By recombinant genetic mechanisms regulating one or more of the present invention, the efficiency or activity of the protein activity can be straight These directly affect a fine chemicals. For example, the desired amino acid biosynthetic pathway The enzyme, the enzyme activity or efficiency (including the presence of multiple copies of the gene) should Production or lead to the production of amino acids required for increased efficiency. For the synthesis of amino acids required for students with Synthesis of competition in the amino acid biosynthetic pathway, the enzyme activity or the reduction in efficiency (including Including gene deletion) amino acids required for production or causes an increase in productivity is due to the intermediate Donor compound and / or energy less competition. For the desired amino acid degradation pathway enzymes, the activity Or can cause reduced efficiency or yield of the desired product higher productivity, which is due to reduced Xie reduced. Finally, the desired amino acid biosynthetic enzyme mutation makes the enzyme is no longer By feedback inhibition, which causes the desired amino acid yield or production efficiency. The invention Vitamins, cofactors, nutritional factors, nucleotides, nucleosides and biosynthesis of trehalose metabolism Sing-degrading enzymes, the same is true. ...

Similarly, when the desired fine chemical substance is not one of the above compounds, the invention is a Regulating the activity of proteins of Corynebacterium glutamicum may still affect the production of large-scale cultivation of the compound The efficiency and / or production. Any organism's metabolic pathways are closely related, a way to make Intermediates used often supplied by different ways. Enzyme expression and function can vary according to metabolize too Cheng compound to regulate the cellular level, the basic necessary for production of molecules such as amino acids and nucleotides Cellular level for large-scale cultivation of microorganisms activity has a significant impact. Thus, adjusting one kinds Amino acid biosynthetic enzyme so that it no longer respond to the feedback inhibition, or increase efficiency, or change, Of one or more amino acids can cause increased cell level. Result, the increase in the supply of amino acids is not However, increasing the supply of molecules necessary for protein synthesis, also increases for a variety of other biological synthesis Pathway intermediates and precursors molecules supply. If a specific amino acid intracellular limited, increasing Increase its production will increase cell capacity of various other metabolic reactions, and make cells more efficiently To produce a variety of proteins, may increase large-scale cultivation of cells or survival of the total energy production rate Force. Increased activity was improved in the fermentation culture to produce the desired fine chemical cells Number, thereby increasing the yield of the compound. By adjusting the degradation activity of the present invention so that the No enzyme catalyzed biosynthesis of the desired compound is important, or the mass of the cell growth culture Length and cell proliferation more effectively the degradation of the compound or catalytic efficiency, there is a similar situation will Condition. It should be emphasized that the present invention optimize the degradation activity of certain molecules or lower biosynthetic activity Corynebacterium glutamicum sex will also produce some fine chemicals have a positive effect. For example, by Reducing the biosynthesis pathway of the desired compound of one or more intermediate competition in biosynthetic pathways The efficiency of the enzyme activity into more intermediates can be used for transformation of the desired material. Similar circumstances Need to improve one or more of degradation of the protein of the invention or efficiency. ...

The aforementioned compound causes increased production of the desired protein mutagenesis program list MP, This does not mean that only limited; these programs change mutagenesis of ordinary skill in the art Is very plain. After these mechanisms, the present invention can be used for nucleic acid and protein molecules MP expressing mutant nucleic acids and produces a protein molecule or a genetic Corynebacterium glutamicum strain, The desired compound thus increasing the yield, production and / or production efficiency. The desired compound can be Corynebacterium glutamicum any natural products, including the final product of the biosynthetic pathway and natural The metabolic pathway intermediates, and not in the metabolism of Corynebacterium glutamicum exists but naturally Of the present invention Corynebacterium glutamicum strains molecules. ...

Table 1: genes include

Lysine biosynthesis

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02229 RXS02970 FRXA01009 RXC02390 RXC01796 RXC01207 RXC00657 RXC00552	GR00653 GR00287	2793 4714	3617 5943	DIAMINOPIMELATE EPIMERASE(EC 5.1.1.7) ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11) ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11) MEMBRANE SPANNING PROTEIN INVOLVED IN LYSINE METABOLISM MEMBRANE ASSOCIATED PROTEIN INVOLVED IN LYSINE METABOLISM CYTOSOLIC PROTEIN INVOLVED IN METABOLISM OF LYSINE AND THREONINE TRANSCRIPTIONAL REGULATOR INVOLVED IN LYSINE METABOLISM CYTOSOLIC PROTEIN INVOLVED IN LYSINE METABOLISM
1 3 5 7 9 11 13 15	2 4 6 8 10 12 14 16						SEQ ID NO	SEQ ID NO

Trehalose

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN00351 FRXA00351 RXA00873 RXA00891	VV0135 GR00066 GR00241 GR00243	37078 1486 3 100	38532 2931 758 4	ALPHA.ALPHA-TREHALOSE-PHOSPHATE SYNTHASE(UDP-FORMING)56 KD SUBUNIT(EC 2.4.1.15) ALPHA.ALPHA-TREHALOSE-PHOSPHATE SYNTHASE(UDP-FORMING)56 KD SUBUNIT(EC 2.4.1.15) trehalose synthase(EC 2.4.1.-) trehalose synthase(EC 2.4.1.-)
17 19 21 23	18 20 22 24						SEQ ID NO	SEQ ID NO

Lysine biosynthesis

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA00534 RXA00533 RXA02843 RXA02022 RXA00044 RXA00863 RXA00864 RXA02843 RXN00355 FRXA00352	GR00137 GR00137 GR00842 GR00613 GR00007 GR00236 GR00236 GR00842 VV0135 GR00068	4758 3469 543 2063 3458 896 1694 543 31980 861	3496 2438 4 3169 4393 1639 2443 4 30961 4	ASPARTOKINASE ALPHA AND BETA SUBUNITS(EC 2.7.2.4) ASPARTATE-SEMIALDEHYDE DEHYDROGENASE(EC 1.2.1.11) 2，3，4，5-TRAHYDROPYRIDINE-2-CARBOXYLATE N-SUCCINYLTRANSFERASE (EC 2.3.1.117) SUCCINYL-DIAMINOPIMELATE DESUCCINYLASE(EC 3.5.1.18) DIHYDRODIPICOLINATE SYNTHASE(EC 4.2.1.52) DIHYDRODIPICOLINATE REDUCTASE(EC 1.3.1.26) probable 2，3-dihydrodipicolinate N-C6-lyase(cyclizing)(EC 4.3.3.-)- Corynebacterium glutamicum 2，3，4，5-TETRAHYDROPYRIDINE-2-CARBOXYLATE N-SUCCINYLTRANSFERASE (EC 2.3.1.117) MESO-DIAMINOPIMELATE D-DEHYDROGENASE MESO-DIAMINOPIMELATE D-DEHYDROGENASE(EC1.4.1.16)
25 27 29 31 33 35 37 39 41 43	26 28 30 32 34 36 38 40 42 44						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA00972 RXA02653 RXA01393 RXA00241 RXA01394 RXA00865 RXS02021 RXS02157 RXC00733 RXC00861 RXC00866 RXC02095 RXC03185	GR00274 GR00752 GR00408 GR00036 GR00408 GR00236	3 5237 4249 5443 4320 2647	1379 7234 3380 6945 5018 3549	DIAMINOPIMELATE DECARBOXYLASE(EC 4.1.1.20) DIAMINOPIMELATE DECARBOXYLASE(EC 4.1.1.20) LYSINE EXPORT REGULATOR PROTEIN L-LYSINE TRANSPORT PROTEIN LYSINE EXPORTER PROTEIN DIHYDRODIPICOLINATE SYNTHASE(EC 4.2.1.52) 2，3，4，5-TETRAHYDROPYRIDINE-2-CARBOXYLATE N-SUCCINYLTRANSFERASE (EC 2.3.1.117) ACETYLORNITHINE AMINOTRANSFERASE(EC2.6.1.11) ABC TRANSPORTER ATP-BINDING PROTEIN INVOLVED IN LYSINE METABOLISM PROTEIN INVOLVED IN LYSINE METABOLISM ZN-DEPENDENT HYDROLASE INVOLVED IN LYSINE METABOLISM ABC TRANSPORTER ATP-BINDING PROTEIN INVOLVED IN LYSINE METABOLISM PROTEIN INVOLVED IN LYSINE METABOLISM
45 47 49 51 53 55 57 59 61 63 65 67 69	46 48 50 52 54 56 58 60 62 64 66 68 70						SEQ ID NO	SEQ ID NO

Glutamate and glutamine metabolism in

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN00367 FRXA00007 FRXA00364 FRXA00367 RXN00076 FRXA00075 RXN00198 FRXA00198 RXN00365 FRXA00365 RXA00366 RXA02072 RXA00323 RXA00335 RXA00324 RXN03176 FRXA02879 RXA00278 RXA00727	VV0196 GR00001 GR00074 GR00075 VV154 GR00012 VV0181 GR00031 VV0196 GR00075 GR00075 GR00628 GR00057 GR00057 GR00057 VV0332 GR10017 GR00043 GR00193	9744 7107 1296 1806 2752 2757 7916 2 14607 630 961 1259 3855 19180 5262 2 2 2612 614	14273 8912 4 964 4122 3419 7368 283 15233 4 605 2599 5192 17750 8396 862 862 1581 1525	GLUTAMATE SYNTHASE[NADH]PRECURSOR(EC 1.4.1.14) GLUTAMATE SYNTHASE(NADPH)LARGE CHAIN PRECURSOR(EC 1.4.1.13) GLUTAMATE SYNTHASE(NADPH)LARGE CHAIN PRECURSOR(EC 1.4.1.13) GLUTAMATE SYNTHASE(NADPH)LARGE CHAIN PRECURSOR(EC 1.4.1.13) GLUTAMATE SYNTHASE(NADPH)SMALL CHAIN(EC 1.4.1.13) GLUTAMATE SYNTHASE(NADPH)SMALL CHAIN(EC 1.4.1.13) GLUTAMATE SYNTHASE(NADPH)SMALL CHAIN(EC 1.4.1.13) GLUTAMATE SYNTHASE(NADPH)SMALL CHAIN(EC 1.4.1.13) GLUTAMATE SYNTHASE[NADPH]SMALL CHAIN(EC 1.4.1.13) GLUTAMATE SYNTHASE(NADPH)SMALL CHAIN(EC 1.4.1.13) GLUTAMATE SYNTHASE(NADPH)SMALL CHAIN(EC 1.4.1.13) NADP-SPECIFIC GLUTAMATE DEHYDROGENASE(EC 1.4.1.4) GLUTAMINE SYNTHETASE(EC 6.3.1.2) GLUTAMINE SYNTHETASE(EC 6.3.1.2) GLUTAMATE-AMMONIA-LIGASE ADENYLYLTRANSFERASE(EC 2.7.7.42) GLUTAMINASE(EC 3.5.1.2) GLUTAMINASE(EC 3.5.1.2) GLUTAMINE-BINDING PROTEIN PRECURSOR GLUTAMINE-BINDING PERIPLASMIC PROTEIN PRECURSOR
71 73 75 77 79 81 83 85 87 89 91 93 95 97 99 101 103 105 107	72 74 76 78 80 82 84 86 88 90 92 94 96 98 100 102 104 106 108						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Alanine and aspartic acid and asparagine metabolic

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02139 RXN00116 FRXA00116 RXN00618 FRXA00618 FRXA00627 RXA02550 RXA02193 RXA02432 RXN03003 RXN00508 RXN00636	GR00639 VV0100 GR00018 VV0135 GR00163 GR00164 GR00729 GR00645 GR00708 VV0138 VV0086 VV0135	6739 26974 510 10288 213 854 1585 1942 2669 680 4701 20972	4901 25814 4 9182 746 1138 275 365 1695 6 5783 19944	ASPARAGINE SYNTHETASE(GLUTAMINE-HYDROLYZING)(EC 6.3.5.4) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ASPARTATE AMMONIA-LYASE(EC 4.3.1.1) L-ASPARAGINASE(EC 3.5.1.1) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ALANINE RACEMASE(EC 5.1.1.1) ALANINE RACEMASE，BIOSYNTHETIC(EC 5.1.1.1)
109 111 113 115 117 119 121 123 125 127 129 131	110 112 114 116 118 120 122 124 126 128 130 132						SEQ ID NO	SEQ ID NO

β-alanine metabolic

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02536 RXS00870 RXS02299	GR00726	8581	7826	BETA-UREIDOPROPIONASE(EC 3.5.1.6) METHYLMALONATE-SEMIALDEHYDE DEHYDROGENASE(ACYLATING)(EC 1.2.1.27) ASPARTATE 1-DECARBOXYLASE PRECURSOR(EC 4.1.1.11)
133 135 137	134 136 138						SEQ ID NO	SEQ ID NO

Glycine and serine metabolic

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA01561 RXA01850 RXA00580 RXA01821 RXN02263 FRXA02263 RXA02176 RXN02758 FRXA02479 FRXA02758 FRXA02759 RXA02501 RXN03105 RXS01130 RXS03112	GR00435 GR00525 GR00156 GR00515 VV0202 GR00654 GR00641 GR00766 GR00717 GR00766 GR00766 GR00720 VV0074	1113 481 7343 10253 11783 33454 11454 5082 393 5082 5330 15041 15857	2042 1827 6042 9876 12160 33813 12581 4648 4 4648 5220 13977 15423	L-SERINE DEHYDRATASE(EC 4.2.1.13) L-SERINE DEHYDRATASE(EC 4.2.1.13) SERINE HYDROXYMETHYLTRANSFERASE(EC 2.1.2.1) SARCOSINE OXIDASE(EC 1.5.3.1) SARCOSINE OXIDASE(EC 1.5.3.1) SARCOSINE OXIDASE(EC 1.5.3.1) PHOSPHOSERINE AMINOTRANSFERASE(EC 2.6.1.52) PHOSPHOSERINE PHOSPHATASE(EC 3.1.3.3) PHOSPHOSERINE PHOSPHATASE(EC 3.1.3.3) PHOSPHOSERINE PHOSPHATASE(EC 3.1.3.3) PHOSPHOSERINE PHOSPHATASE(EC 3.1.3.3) PHOSPHOSERINE PHOSPHATASE(EC 3.1.3.3) SARCOSINE OXIDASE(EC 1.5.3.1) D-3-PHOSPHOGLYCERATE DEHYDROGENASE(EC 1.1.1.95) D-3-PHOSPHOGLYCERATE DEHYDROGENASE(EC 1.1.1.95)
139 141 143 145 147 149 151 153 155 157 159 161 163 165 167	140 142 144 146 148 150 152 154 156 158 160 162 164 166 168						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Threonine metabolic

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN00969 FRXA00974 RXA00970 RXA00330 RXN00403 FRXA00403 RXC01207 RXC00152	VV0149 GR00274 GR00273 GR00057 VV0086 GR00088	12053 2623 161 12968 70041 723	13387 3015 1087 14410 68911 1832	HOMOSERINE DEHYDROGENASE(EC 1.1.1.3) HOMOSERINE DEHYDROGENASE(EC 1.1.1.3) HOMOSERINE KINASE(EC 2.7.1.39) THREONINE SYNTHASE(EC 4.2.99.2) HOMOSERINE O-ACETYLTRANSFERASE HOMOSERINE O-ACETYLTRANSFERASE(EC 2.3.1.11) CYTOSOLIC PROTEIN INVOLVED IN METABOLISM OF LYSINE AND THREONINE MEMBRANE ASSOCIATED PROTEIN INVOLYED IN THREONINE METABOLISM
169 171 173 175 177 179 181 183	170 172 174 176 178 180 182 184						SEQ ID NO	SEQ ID NO

Methionine and S-adenosylmethionine metabolic

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA00115 RXN00403 FRXA00403 RXS03158 FRXA00254 RXA02532 RXS03159 FRXA02768 RXA00216 RXN00402 FRXA00402 RXA00405 RXA02197 RXN02198 FRXA02198 RXN03074 FRXA02906 RXND0132 FRXA00132	GR00017 VV0086 GR00088 GR00038 GR00726 GR00770 GR00032 VV0086 GR00088 GR00089 GR00645 VV0302 GR00646 VV0042 GR10044 VV0124 GR00020	5359 70041 723 2404 3085 1919 16286 70787 1 3289 4552 9228 2483 2238 1142 3612 7728	4313 68911 1832 1811 2039 2521 15297 70188 576 3801 4025 11726 6 1741 645 5045 7624	HOMOSERINE O-ACETYLTRANSFERASE(EC 2.3.1.31) HOMOSERINE O-ACETYLTRANSFERASE HOMOSERINE O-ACETYLTRANSFERASE(EC 2.3.1.11) CYSTATHIONINE GAMMA-SYNTHASE(EC 4.2.99.9) CYSTATHIONINE GAMMA-SYNTHASE(EC 4.2.99.9) CYSTATHIONINE GAMMA-SYNTHASE(EC 4.2.99.9) CYSTATHIONINE GAMMA-SYNTHASE(EC 4.2.99.9) CYSTATHIONINE GAMMA-SYNTHASE(EC 4.2.99.9) 5-methyltetrahydrofolate-homocysteine methyltransferase(methionine synthetase) O-ACETYLHOMOSERINE SULFHYDRYLASE(EC 4.2.99.10)/O-ACETYLSERINE SULFHYDRYLASE(EC 4.2.99.8) O-ACETYLHOMOSERINE SULFHYDRYLASE(EC 4.2.99.10)/O-ACETYLSERINE SULFHYDRYLASE(EC 4.2.99.8) O-ACETYLHOMOSERINE SULFHYDRYLASE(EC 4.2.99.10)/O-ACETYLSERINE SULFHYDRYLASE(EC 4.2.99.8) 5-METHYLTETRAHYDROFOLATE-HOMOCYSTEINE METHYLTRANSFERASE (EC 2.1.1.13) 5-METHYLTETRAHYDROFOLATE-HOMOCYSTEINE METHYLTRANSFERASE (EC2.1.1.13) 5-METHYLTETRAHYDROFOLATE-HOMOCYSTEINE METHYLTRANSFERASE (EC 2.1.1.13) S-ADENOSYLMETHIONINE：2-DEMETHYLMENAQUINONE METHYLTRANSFERASE(EC 2.1.-.-) S-ADENOSYLMETHIONINE：2-DEMETHYLMENAQUINONE METHYLTRANSFERASE(EC 2.1.-.-) ADENOSYLHOMOCYSTEINASE(EC 3.3.1.1) ADENOSYLHOMOCYSTEINASE(EC 3.3.1.1)
185 187 189 191 193 195 197 199 201 203 205 207 209 211 213 215 217 219 221	186 188 190 192 194 196 198 200 202 204 206 208 210 212 214 216 218 220 222						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	FRXA01371 RXN02085 FRXA02085 FRXA02086 RXN02648 FRXA02648 FFXA02658 RXC02238 RXC00128	GR00398 GR00629 GR00629 GR00751 GR00752	2339 3496 5252 5254 14764	3631 5295 5731 4730 15447	ADENOSYLHOMOCYSTEINASE(EC 3.3.1.1) S-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLJRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) PROTEIN INVOLVED IN METABOLISM OF S-ADENOSYUMETHI0NINE，PURINES AND PANTOTHENATE EXPORTED PROTEIN INVOLVED IN METABOLISM OF PYRIDIMES AND ADENOSYLHOMOCYSTEINE
223 225 227 229 231 233 235 237 239	224 226 228 230 232 234 236 238 240						SEQ ID NO	SEQ ID NO

S-adenosylmethionine (SAM) biosynthesis

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RKA02240	GR00654	7160	8380	S-ADENOSYLMETHIONINE SYNTHEIASE(EC 2.5.1.6)
241	242						SEQ ID NO	SEQ ID NO

Cysteine Metabolism

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA00780 RXA00779 RXN00402 FRXA00402 RXS00405 RXC00164 RXC01191	GR00206 GR00206 VV0086 GR00088	1689 550 70787 1	2234 1482 70188 576	SERINE ACETYLTRANSFERASE(EC 2.3.1.30) CYSTEINE SYNTHASE(EC 4.2.99.8) O-ACETYLHOMOSERINE SULFHYDRYLASE(EC 4.2.99.10)/O-ACETYLSERINE SULFHYDRYLASE(EC 4.2.99.8) O-ACETYLHOMOSERINE SULFHYDRYLASE(EC 4.2.99.10)/O-ACETYLSERINE SULFHYDRYLASE(EC 4.2.99.8) O-ACETYLHOMOSERINE SULFHYDRYLASE(EC 4.2.99.10)/O-ACETYLSERINE SULFHYDRYLASE(EC 4.2.99.8) ABC TRANSPORTER ATP-BINDING PROTEIN INVOLVED IN CYSTEINE METABOLISM ABC TRANSPORTER ATP-BINDING PROTEIN INVOLVED IN CYSTEINE METABOLISM
243 245 247 249 251 253 255	244 246 248 250 252 254 256						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Valine, leucine and isoleucine

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02646 RXA00766 RXN01690 FRXA01690 RXN01026 FRXA01026 RXN01127 FRXA01132 RXN00536 FRXA00536 RXN02965 RXN01929 FRXA01929 RXN01420 RXS01145 FRXA01145	GR00751 GR00204 VV0246 GR00473 VV0143 GR00294 VV0157 GR00315 VV0219 GR00137 VV0143 VV0127 GR00555 VV0122 GR00321	3856 5091 1296 1248 9171 1 4491 1349 6128 6128 7711 47590 2766 15584 1075	2588 4249 196 196 7513 1602 3472 1651 7498 7360 7121 48402 1960 14643 1530	THREONINE DEHYDRATASE BIOSYNTHETIC(EC 4.2.1.16) BRANCHED-CHAIN AMINO ACID AMINOTRANSFERASE(EC 2.6.1.42) BRANCHED-CHAIN AMINO ACID AMINOTRANSFERASE(EC 2.6.1.42) BRANCHED-CHAIN AMINO ACID AMINOTRANSFERASE(EC 2.6.1.42) 3-ISOPROPYLMALATE DEHYDRATASE LARGE SUBUNIT(EC 4.2.1.33) 3-ISOPROPYLMALATE DEHYDRATASE LARGE SUBUNIT(EC 4.2.1.33) 3-ISOPROPYLMALATE DEHYDROGENASE(EC 1.1.1.85) 3-ISOPROPYLMALATE DEHYDROGENASE(EC 1.1.1.85) 2-ISOPROPYLMALATE SYNTHASE(EC 4.1.3.12) 2-ISOPROPYLMALATE SYNTHASE(EC 4.1.3.1) 3-ISOPROPYLMALATE DEHYDRATASE SMALL SUBUNIT(EC 4.2.1.33) 3-METHYL-2-OXOBUTANOATE HYDROXYMETHYLTRANSFERASE(EC 2.1.2.11) /DECARBOXYLASE(EC 4.1.1.44) 3-METHYL-2-OXOBUTANOATE HYDROXYMETHYLTRANSFERASE(EC 2.1.2.11) 4”-MYCAROSYL ISOVALERYL-COA TRANSFERASE(EC 2.-.-.-) KETOL-ACID REDUCTOISOMERASE(EC 1.1.1.86) KETOL-ACID REDUCTOISOMERASE(EC 1.1.1.86)
257 259 261 263 265 267 269 271 273 275 277 279 281 283 285 287	258 260 262 264 266 268 270 272 274 276 278 280 282 284 286 288						SEQ ID NO	SEQ ID NO

Arginine and proline metabolism in

Proline biosynthetic enzyme:

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02375 RXN02382 FRXA02378 FRXA02382 RXA02499 RXS02157 RXS02262 RXS02970 FRXA01009	GR00689 VV0213 GR00690 GR00691 GR00720 GR00287	1449 5162 624 2493 11883 4714	223 3867 16 1894 12692 5943	GLUTAMATE5-KINASE(EC 2.7.2.11) GAMMA-GLUTAMYL PHOSPHATE REDUCTASE(GPR)(EC 1.2.1.41) GAMMA-GLUTAMYL PHOSPHATE REDUCTASE(GPR)(EC 1.2.1.41) GAMMA-GLUTAMYL PHOSPHATE REDUCTASE(GPR)(EC 1.2.1.41) PYRROLINE-5-CARBOXYLATE REDUCTASE(EC 1.5.1.2) ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11) ORNITHINE CYCLODEAMINASE(EC 4.3.1.12) ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11) ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11)
289 291 293 295 297 299 301 303 305	290 292 294 296 298 300 302 304 306						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Proline-degrading enzymes:

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN00023 FRXA00023 FRXA02284 RXC02498	VV0127 GR00003 GR00660	68158 2 3028	64703 454 5	PROLINE DEHYDROGENASE(EC 1.5.99.8)/DELTA-1-PYRROLINE-5- CARBOXYLATEDEHYDROGENASE (EC 1.5.1.12) PROLINE DEHYDROGENASE(EC 1.5.99.8)/DELTA-1-PYRROLINE-5- CARBOXYLATE DEHYDROGENASE (EC 1.5.1.12) PROLINE DEHYDROGENASE(EC 1.5.99.8)/DELTA-1-PYRROLINE-5- CARBOXYLATE DEHYDROGENASE (EC 1.5.1.12) PROTEIN INVOLVED IN PROLINE METABOLISM
307 309 311 313	308 310 312 314						SEQ ID NO	SEQ ID NO

3 - hydroxy - proline synthesis:

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA01491	GR00423	5337	4687	DNA FOR L-PROLINE 3-HYDROXYLASE，COMPLETE CDS
315	316						SEQ ID NO	SEQ ID NO

Ornithine, arginine and spermidine metabolic enzymes:

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02155 RXA02156 RXN02153 FRXA02153 RXA02154 RXA02157 RXS02970 FRXA01009 RXA02158 RXA02160 RXN02162 FRXA02161 FRXA02162 RXA02262 RXA00219 RXA01508 RXA01757 RXA02159 RXN02154 RXS00147 RXS00905 RXS00906	GR00640 GR00640 VV0122 GR00640 GR00640 GR00640 GR00287 GR00640 GR00640 VV0122 GR00640 GR00640 GR00654 GR00032 GR00424 GR00498 GR00640 VV0122	1913 3125 14106 757 1536 4079 4714 5268 6914 6683 8180 8949 32291 19289 12652 2942 6231 13327	3076 4075 13327 1536 1826 5251 5943 6224 8116 5253 8962 9611 33436 20230 14190 2142 6743 13037	GLUTAMATE N-ACETYLTRANSFERASE(EC 2.3.1.35)/AMINO-ACID ACETYLTRANSFERASE(EC 2.3.1.1) ACETYLGLUTAMATE KINASE(EC 2.7.2.8) N-ACETYL-GAMMA-GLUTAMYL-PHOSPHATE REDUCTASE(EC 1.2.1.38) N-ACETYLGLUTAMATE-5-SEMIALDEHYDE DEHYDROGENASE N-ACETYLGLUTAMATE-5-SEMIALDEHYDE DEHYDROGENASE ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11) ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11) ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11) ORNITHINE CARBAMOYLTRANSFERASE(EC 2.1.3.3) ARGININOSUCCINATE SYNTHASE(EC 6.3.4.5) ARGININOSUCCINATE LYASE(EC 4.3.2.1) ARGININOSUCCINATE LYASE(EC 4.3.2.1) ARGININOSUCCINATELYASE(EC 4.3.2.1) ORNITHINE CYCLODEAMINASE(EC 4.3.1.12) SPERMIDINE SYNTHASE(EC 2.5.1.16) SPERMIDINE SYNTHASE(EC 2.5.1.16) PUTRESCINE OXIDASE(EC 1.4.3.10) ARGININE HYDROXIMATE RESISTANCE PROTEIN N-ACETYL-GAMMA-GLUTAMYL-PHOSPHATEREDUCTASE(EC 1.2.1.38) CARBAMOYL-PHOSPHATE SYNTHASE SMALL CHAIN(EC 6.3.5.5) N-ACYL-L-AMINO ACIDAMIDOHYDROLASE(EC 3.5.1.14) N-ACYL-L-AMINO ACIDAMIDOHYDROLASE(EC 3.5.1.14)
317 319 321 323 325 327 329 331 333 335 337 339 341 343 345 347 349 351 353 355 357 359	318 320 322 324 326 328 330 332 334 336 338 340 342 344 346 348 350 352 354 356 358 360						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXS00907 RXS02001 RXS02101 RXS02234 FRXA02234 RXS02565 RXS02937	GT00654	1	3198	N-ACYL-L-AMINO ACID AMIDOHYDROLASE(EC 3.5.1.14) N-ACYL-L-AMIMO ACID AMIDOHYDROLASE(EC 3.5.1.14) N-ACYL-L-AMINO ACID AMIDOHYDROLASE(EC 3.5.1.14) CARBAMOYL-PHOSPHATE SYNTHASE LARGE CHAIN(EC 6.3.5.5) CARBAMOYL-PHOSPHATE SYNTHASE SYNGE CHAIN(EC 6.3.5.5) N-ACYL-L-AMINO ACID AMIDOHYDROLASE(EC 3.5.1.14) N-ACYL-L-AMINO ACID AMIDOHYDROLASE(EC 3.5.1.14)
361 363 365 367 369 371 373	362 364 366 368 370 372 374						SEQ ID NO	SEQ ID NO

Histidine metabolic

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02194 RXA02195 RXA01097 RXA01100 RXA01101 RXN01657 FRXA01657 RXA01098 RXN01104 FRXA01104 RXN00446 FRXA00446 RXA01105 RXA01106 RXC00930 RXC01096 RXC01056 RXC01158	GR00645 GR00645 GR00306 GR00306 GR00306 VV0010 GR00460 GR00306 VV0059 GR00306 VV0112 GR00108 GR00306 GR00306	2897 3186 4726 7072 7726 39950 2444 5499 7037 10927 24181 4 12044 13378	2055 2917 4373 6335 7094 39351 2944 4726 6432 10322 23318 525 10947 12053	ATP PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.17) PHOSPHORIBOSYL-ATP PYROPHOSPHOHYDROLASE(EC 3.6.1.31) PHOSPHORIBOSYL-AMP CYCLOHYDROLASE(EC 3.5.4.19) PHOSPHORIBOSYLFORMIMINO-5-AMINOIMIDAZOLE CARBOXAMIDE RIBOTIDE ISOMERASE(EC 5.3.1.16) AMIDOTRANSFERASE HISH(EC 2.4.2.-) AMIDOTRANSFERASE HISH(EC 2.4.2.-) AMIDOTRANSFERASE HISH(EC 2.4.2.-) HISF PROTEIN IMIDAZOLEGLYCEROL-PHOSPHATE DEHYDRATASE(EC 4.2.1.19) IMIDAZOLEGLYCEROL-PHOSPHATE DEHYDRATASE(EC 4.2.1.19)/ HISTIDINOL-PHOSPHATASE(EC 3.1.3.15) HISTIDINOL-PHOSPHATE AMINOTRANSFERASE(EC 2.6.1.9) HISTIDINOL-PHOSPHATE AMINOTRANSFERASE(EC 2.6.1.9) HISTIDINOL-PHOSPHATE AMINOTRANSFERASE(EC 2.6.1.9) HISTIDINOL DEHYDROGENASE(EC 1.1.1.23) PROTEIN INVOLVED IN HISTIDINE METABOLISM PROTEIN INVOLVED IN HISTIDINE METABOLISM PROTEIN INVOLVED IN HISTIDINE METABOLISM MEMBRANE SPANING PROTEIN INVOLVED IN HISTIDINE METABOLISM
375 377 379 381 383 385 387 389 391 393 389 397 399 401 403 405 407 409	376 378 380 382 384 366 388 390 392 394 396 398 400 402 404 406 408 410						SEQ ID NO	SEQ ID NO

Aromatic amino acid metabolism

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02458 RXA02790 RXN00954 FRXA00954 RXN00957 FRXA00957	GR00712 GR00777 VV0247 GR00263 VV0208 GR00264	3056 5806 3197 3 1211 3	4345 6948 2577 590 2764 1130	3-PHOSPHOSHIKIMATE1-CARBOXYPINYLTRANSFERASE(EC 2.5.1.19) 4-AMINO-4-DEOXYCHORISMATE LYASE(EC 4.-.-.-) ANTHRANILATE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.18) ANTHRANILATE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.18) ANTHRANILATE SYNTHASE COMPONENT I(EC 4.1.3.27) ANTHRANILATE SYNTHASE COMPONENT I(EC 4.1.3.27)
411 413 415 417 419 421	412 414 416 418 420 422						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02687 RXN01698 FRXA01698 RXA01095 RXA00955 RXA02814 RXA00229 RXA02093 RXA02791 RXA01699 RXA00952 RXN00956 FRXA00956 RXA0064 RXN00448 FRXA00448 FRXA00452 RXA00584 RXA00579 RXA00958 RXN03007 RXN02918 RXN01116 RXN01115 RXS00116 FRXA00116 RXS00391 RXS00393 FRXA00393 RXS00446 FRXA00446 RS00618 FRXA00618 FRXA00327 RXS01105 RXS02315 RXS02550 RXS02319 RXS02908 RXS03003 RXS03026	GR00754 VV0134 GR00477 GR00306 GR00263 GR00795 GR00033 GR00629 GR00777 GR00477 GR00262 VV0247 GR00263 GR00010 VV0112 GR00109 GR00110 GR00156 GR00156 GR00264 VV0208 VV0086 VV0182 VV0182 GR00018 GR00086 GR00108 GR00163 GR00164	11306 11507 2 3603 586 598 1715 12444 6968 984 97 1140 2027 2499 33959 3 854 11384 5946 1130 3410 25447 7497 10347 510 4030 4 213 854	12250 12736 991 2821 2007 128 936 13247 7795 1553 936 4 3157 3776 32940 668 1099 10260 4087 1753 3778 25887 6886 11099 4 4911 525 746 1138	CHORISMATE MUTASE(EC 5.4.99.5)/PREPHENATE DEHYDRATASE(EC 4.2.1.51) CHORISMATE SYNTHASE(EC 4.6.1.4) CHORISMATE SYNTHASE(EC 4.6.1.4) INDOLE-3-GLYCEROL PHOSPHATE SYNTHASE(EC 4.1.1.48) INDOLE-3-GLYCEROL PHOSPHATE SYNTHASE(EC 4.1.1.48)/N-(5’-PHOSPHO- RIBOSYL)ANTHRANILATE ISOMERASE(EC 5.3.1.24) ISOCHORISMATE MUTASE SHIKIMATE 5-DEHYDROGENASE(EC 1.1.1.25) SHIKIMATE 5-DEHYDROGENASE(EC 1.1.1.25) SHIKIMATE 5-DEHYDROGENASE(EC 1.1.1.25) SHIKIMATE KINASE(EC 2.7.1.71) TRYPtopHAN SYNTHASE ALPHA CHAIN(EC 4.2.1.20) TRYPtopHAN SYNTHASE BETA CHAIN(EC 4.2.1.20) TRYPtopHAN SYNTHASE BETA CHAIN(EC 4.2.1.20) TYROSINE AMINOTRANSFERASE(EC 2.6.1.5) PREPHENATE DEHYDROGENASE(EC 1.3.1.12) PREPHENATE DEHYDROGENASE(EC 1.3.1.12) PREPHENATE DEHYDROGENASE(EC 1.3.1.12) PHOSPHO-2-DEHYDRO-3-DEOXYHEPTONATE ALDOLASE(EC 4.1.2.15) PARA-AMINOBENZOATE SYNTHASE COMPONENT I(EC 4.1.3.-) PARA-AMINOBENZOATE SYNTHASE GLUTAMINE AMIDOTRANSFERASE COMPONENT II(EC 4.1.3.-)/ANTHRANILATE SYNTHASE COMPONENT II(EC 4.1.3.27) ANTHRANILATE SYNTHASE COMPONENT II(EC 4.1.3.27) TRYPtopHAN SYNTHASE BETA CHAIN(EC 4.2.1.20) 3-OXOADIPATE COA-TRANSFERASE SUBUNIT B(EC 2.8.3.6) 3-OXOADIPATE ENOL-LACTONE HYDROLASE(EC 3.1.1.24)/4- CARBOXYMUCONOLACTONE ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) O-SUCCINYUBENZOICACID-COALIGASE(EC 6.2.1.26) 1，4-DIHYDROXY-2-NAPHTHOATE OCTAPRENYLTRANSFERASE(EC 2.5.-.-) 1，4-DIHYDROXY-2-NAPHTHOATE OCTAPRENYLTRANSFERASE(EC 2.5.-.-) HISTIDINOL-PHOSPHATE AMINOTRANSFERASE(EC 2.6.1.9) HISTIDINOL-PHOSPHATE AMINOTRANSFERASE(EC 2.6.1.9) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) HISTIDINOL-PHOSPHATE AMINOTRANSFERASE(EC 2.6.1.9) 2-SUCCINYL-6-HYDROXY-2.4(YCLOHEXADIENE-1-CARBOXYLATE SYNTHASE/2-OXOGLUTARATE DECARBOXYLASE(EC 4.1.1.71) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) NAPHTHOATE SYNTHASE(EC 4.1.3.36) O-SUCCINYLBENZOIC ACID-COA LIGASE(EC 6.2.1.26) ASPARTATE AMINOTRANSFERASE(EC 2.6.1.1) 3-DEHYDROQUINATE DEHYDRATASE(EC 4.2.1.10)
423 425 427 429 431 433 435 437 439 441 443 445 447 449 451 453 455 457 459 46 463 465 467 469 471 473 475 477 479 481 483 485 487 489 491 493 495 497 499 501 503	424 426 428 430 432 434 436 438 440 442 444 446 448 450 452 454 456 458 460 462 464 466 468 470 472 474 476 478 480 482 484 486 488 490 492 494 496 498 500 502 504						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXS03074 RXC01434 RXC02080 RXC02789 RXC02295	S-ADENOSYLMETHIONINE：2-DEMETHYLMENAQUINONE METHYLTRANSFERASE(EC 2.1.-.-) MEMBRANE SPANNING PROTEIN INVOLVED IN METABOLISM OF AROMA TIC AMINO ACIDS AND RIBOFLAVIN MEMBRANE SPANNING PROTEIN INVOLVED IN METABOLISM OF AROMA TIC AMINO ACIDS CYTOSOLIC PROTEIN INVOLVED IN METABOLISM OF AROMATIC AMINO ACIDS MEMBRANE SPANNING PROTEIN INVOLVED IN METABOLISM OF AROMATIC AMINO ACIDS
505 507 509 511 513	506 508 510 512 514						SEQ ID NO	SEQ ID NO

Aminobutyric acid metabolic

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN03063 RXN02970 FRXA01009	VV0035 VV0021 GR00287	666 4714 4714	1697 6081 5943	4-aminobutyrate aminotransferase(EC 2.6.1.19) ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11) ACETYLORNITHINE AMINOTRANSFERASE(EC 2.6.1.11)
515 517 519	516 518 520						SEQ ID NO	SEQ ID NO

Vitamins, vitamin-like substance (cofactor), nutritional factors

Thiamine Metabolism

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA01551 RXA01019 RXA01352 RXA01381 RXA01360 RXA01361 RXA01208 RXA00838 RXA02400 RXN01209 FRXA01209 RXN01413 RXN01617 FRXA01617 RXS01807 RXC01021	GR00431 GR00291 GR00393 GR00403 GR00394 GR00394 GR00348 GR00227 GR000699 VV0270 GR00348 VV0050 VV0050 GR00451	2945 6 609 3206 162 983 229 1532 1988 1019 1019 27306 22187 2	4819 995 4 2286 4 378 1032 633 2557 2446 2446 27905 22858 616	THIAMINBIOSYNTHESIS PROTEIN THIC THIAMIN-MONOPHOSPHATE KINASE(EC 2.7.4.16) THIAMIN-PHOSPHATE PYROPHOSPHORYLASE(EC 2.5.1.3) THIF PROTEIN THIG PROTEIN THIG PROTEIN HYDROXYETHYLTHIAZOLE KINASE(EC 2.7.1.50) APBA PROTEIN THIAMIN BIOSYNTHESIS PROTEIN X PHOSPHOMETHYLPYRIMIDINE KINASE(EC 2.7.4.7) PHOSPHOMETHYLPYRIMLDINE KINASE(EC 2.7.4.7) PHOSPHOMETHYLPYRIMIDINE KINASE(EC 2.7.4.7) PHOSPHOMETHYLPYRIMIDINE KINASE(EC 2.7.4.7) PHOSPHOMETHYLPYRIMIDINE KINASE(EC 2.7.4.7) PYRIDOXINE KINASE(EC 2.7.1.35) CYTOSOLIC KINASE INVOLVED IN METABOLISM OF SUGARS AND THIAMIN
521 523 525 527 529 531 533 535 537 539 541 543 545 547 549 551	522 524 526 528 530 532 534 536 538 540 542 544 546 548 550 552						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Riboflavin Metabolism

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN02246 FRXA02246 RXA02247 RXN02248 FRXA02248 RXN02249 FRXA02249 RXA02250 RXA01489 RXA02135 RXA01489 RXN01712 FRXA01712 RXN02384 RXN01560 RXN00667 RXC01711 RXC02380 FRXA02380 RXC02921 RXC01434	VV0130 GR00654 GR00654 VV0130 GR00654 VV0130 GR00654 GR00654 GR00423 GR00639 GR00423 VV0191 GR00484 VV0213 VV0319 VV0109 GR00691	4388 14299 15286 6021 15932 7301 17212 17778 3410 2809 3410 8993 2652 1386 767 1363 709	5371 15282 15918 7286 17197 7777 17688 18356 2388 1736 2388 8298 2152 679 438 350 56	diaminohydroxyphosphoribosylaminopyrimiditne deaminase(EC 3.5.4.26)/5-amino- 6-(5-phosphoribosylamino)uracil reductare(EC 1.1.1.1.93) RIBG PROTEIN riboflavin-specific deaminase[EC：3.5.4.-] RIBOFLAYIN SYNTHASE ALPHA CHAIN(EC 2.5.1.9) GTP CYCLOHYDROLASE II(EC 3.5.4.25)/3，4-DIHYDROXY-2-BUTANONE 4- PHOSPHATE SYNTHASE RIBA PROTEIN-GTP cyclohydrolase II[EC：3.5.4.25] 6，7-DIMETHYL-8-RIBITYLLUMAZINE SYNTHASE(EC 2.5.1.9) RIBH PROTEIN-6，7-dimethyl-8-ribityllumazine synthase(dmrl synthase，lumazine synthase，riboflavin synthase beta chain)[EC：2.5.1.9] RIBX PROTEIN RIBOFLAVIN KINASE(EC 2.7.1.26)/FMN ADENYLYLTRANSFERASE(EC 2.7.7.2) NICOTINATE-NUCLEOTIDE-DIMETHYLBENZIMIDAZOLE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.21) RIBOFLAVIN KINASE(EC 2.7.1.26)/FMN ADENYLYLTRANSFERASE(EC 2.7.7.2) RIBOFLAVIN-SPECIFIC DEAMINASE(EC 3.5.4.-) RIBOFLAVIN-SPECIFIC DEAMINASE(EC 3.5.4.-) ALPHA-RIBAZOLE-5’-PHOSPHATE PHOSPHATASE(EC 3.1.3.-) RIBOFLAVIN-SPECIFIC DEAMINASE(EC 3.5.4.-) DRAP DEAMINASE MEMBRANE SPANNING PROTEIN INVOLVED IN RIBOFLAVIN METABOLISM PROTEIN INVOLVED IN RIBOFLAVIN METABOUSM Predicted nucleotidyltransferases CYIOSOLIC PROTEIN INVOLVED IN METABOLISM OF RIBOFLAVIN AND LIPIDS MEMBRANE SPANNING PROTEIN INVOLVED IN METABOLISM OF AROMATIC AMINO ACIDS AND RIBOFLAVIN
553 555 557 559 561 563 565 567 569 571 573 575 577 579 581 583 585 587 589 591 593	554 556 558 560 562 564 566 568 570 572 574 576 578 580 582 584 586 588 590 592 594						SEQ ID NO	SEQ ID NO

Vitamin B6 Metabolism

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA01807	GR00509	7868	7077	PYRIDOXINE KINASE(EC 2.7.1.35)，pyridoxal/pyridoxine/pyridoxamine kinase
595	596						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Niacin, nicotinamide, NAD and NADP

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN02754 FRXA02405 FRXA02754 RXA02112 RXA02111	VV0084 GR00701 GR00766 GR00632 GR00632	22564 774 3 5600 4310	23901 4 488 6436 5593	NICOTINATE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.11) NICOTINATE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.11) NICOTINATE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.11) NICOTINATE-NUCLEOTIDE PYROPHOSPHORYLASE(CARBOXYLATING)(EC 2.4.2.19) QUINOLINATE SYNTHETASE A
597 599 601 B03 605	598 600 602 604 606						SEQ ID NO	SEQ ID NO

NAD biosynthesis

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID N0	SEQ ID NO	RXA01073 RXN02754	GR00300 VV0084	1274 22564	2104 23901	NH(3)-DEPENDENTNAD(+)SYNTHEYASE(EC 6.3.5.1) NICOTINSTE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.11)
607 609	608 610						SEQ ID N0	SEQ ID NO

Pantothenic acid and coenzyme A (CoA) biosynthesis

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02299 RXA01928 RXN01929 FRXA01929 RXA01521 RXS01145 FRXA01145 RXA02239 RXA00581 RXS00838 RXC02238	GR00662 GR00555 VV0127 GR00555 GR00424 GR00321 GR00654 GR00156	10452 1957 47590 2766 25167 1075 5784 7572	10859 1121 48402 1960 25964 1530 7049 8540	ASPARTATE 1-DECARBOXYLASE PRECURSOR(EC 4.1.1.1) PANTOATE-BETA-ALANINE LIGASE(EC 6.3.2.1) 3-METHYL-2-OXOBUTANOATE HYDROXYMETHYLTRANSFERASE(EC 2.1.2.11) /DECARBOXYLASE(EC 4.1.1.44) 3-METHYL-2-OXOBUTANQATE HYDROXYMETHYLTRANSFERASE(EC 2.1.2.11) PANTOATE-BETA-ALANINE LIGASE(EC 6.3.2.11) KETOL-ACID REDUCTOISOMERASE(EC 1.1.1.86) KETOL-ACID REDUCTOISOMERASE(EC 1.1.1.86) DNA/PANTOTHENATE METABOLISM FLAVOPROTEIN PANTOTHENATE KINASE(EC 2.7.1.33) 2-DEHYDROPANTOATE 2-REDUCTASE(EC 1.1.1.1.69) PROTEIN INVOLVED IN METABOLISM OF S-ADENOSYLMETHIONINE，PURINES AND PANTOTHENATE
511 613 615 617 619 621 623 625 627 629 631	612 614 616 618 620 622 624 626 628 630 632						SEQ ID NO	SEQ ID NO

Biotin Metabolism

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN03058	VV0028	8272	8754	BIOTIN SYNTHESIS PROTEIN BIOC
633	634						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	FRXA02903 RXA00166 RXA00633 RXA00632 RXA00295 RXA00223 RXN00262 FRXA00262 RXN00435 FRXA00435 FRXA02801 RXA02516 RXA02517	GR10040 GR00025 GR00166 GR00166 GR00047 GR00032 VV0123 GR00040 VV0112 GR00100 GR00782 GR00723 GR00723	11532 3650 3556 2281 3407 23967 16681 79 10037 3563 438 1724 2989	12014 4309 2288 1610 4408 22879 15608 897 11209 2949 4 2986 3435	BIOTIN SYNTHESIS PROTEIN BIOC BIOTIN SYNTHESIS PROTEIN BIOC ADENOSYLMETHIONINE-8-AMINO-7-OXONONANOATE AMINOTRANSFERASE (EC 2.6.1.62) DETHIOBIOTIN SYNTHETASE(EC 6.3.3.3) BIOTIN SYNTHASE(EC 2.8.1.6) NIFS PROTEIN NIFS PROTEIN NIFS PROTEIN NIFS PROTEIN NIFS PROTEIN NIFS PROTEIN NIFS PROTEIN NIFU PROTEIN
635 637 639 641 643 645 647 649 651 653 655 657 659	636 638 640 642 644 646 648 650 652 654 656 658 660						SEQ ID NO	SEQ ID NO

Lipoic acid

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA01747 RXA01746 RXA02106 RXS01183 RXS01260 RXS01261	GR00495 GR00495 GR00632	2506 1614 472	3549 2366 1527	LIPOIC ACID SYNTHETASE LIPOATE-PROTEIN LIGASE B(EC 6.-.-.-) LIPOATE-PROTEIN LIGASE A(EC 6.-.-.-) DIHYDROLIPOAMIDE SUCCINYLTRANSFERASE COMPONENT(E2)OF 2- OXOGLUTARATE DEHYDROGENASE COMPLEX(EC 2.3.1.61) LIPOAMIDE DEHYDROGENASE COMPONENT(E3)OFBRANCHED-CHAIN ALPHA-KETO ACID DEHYDROGENASE COMPLEX(EC 1.8.1.4) LIPOAMIDE DEHYDROGENASE COMPONENT(E3)OFBRANCHED-CHAIN ALPHA-KETO ACID DEHYDROGENASE COMPLEX(EC 1.8.1.4)
661 663 665 667 669 671	662 654 666 668 670 672						SEQ ID NO	SEQ ID NO

Folic acid biosynthesis

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02717 RXN02027 FRXA02027 RXA00106 RXN01321 FRXA01321 RXA00461 RXA01514 RXA01516	GR00758 VV0296 GR00616 GR00014 VV0082 GR00384 GR00116 GR00424 GR00424	18281 503 500 17469 8868 23 428 20922 22360	17400 1003 6 17924 9788 559 1279 21509 22749	5，10-METHYLENETETRAHYDROFOLATE REDUCTASE(EC 1.7.99.5) 5-FORMYLTETRAHYDROFOLATE CYCLO-LIGASE(EC 6.3.3.2) 5-FORMYLTETRAHYDROFOLATE CYCLO-LIGASE(EC 6.3.3.2) DIHYDROFOLATE REDUCTASE(EC 1.5.1.3) FORMYLTETRAHYDROFOLATE DEFORMYLASE(EC 3.5.1.10) FORMYLTETRAHYDROFOLATE DEFORMYLASE(EC 3.5.1.10) METHYLENETETRAHYDROFOLATE DEHYDROGENASE(EC 1.5.1.5) METHENYLTETRAHYDROFOLATE CYCLOHYDROLASE(EC 3.5.4.9) GTP CYCLOHYDROLASE I(EC 3.5.4.16) DIHYDRONEOPTERIN ALDOLASE(EC 4.1.2.25)
673 675 677 679 681 683 685 687 689	674 676 67B 680 682 684 686 688 690						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA01515 RXA02024 RXA00106 RXA00989 RXA01517 RXA00579 RXA00958 RXA02790 RXA00106 RXN02198 FRXA02198 RXN02085 FRXA02085 FRXA02086 RXN02648 FRXA02648 FRXA0265B RXS02197 RXC00988 RXC01518 RXC01942	GR00424 GR00613 GR00014 GR00280 GR00424 GR00156 GR00264 GR00777 GR00014 VV0302 GR00646 VV0126 GR00629 GR00629 GR00751 GR00752	21513 4026 17469 2903 22752 5946 1130 5806 17469 9228 2483 8483 3496 5252 5254 14764	22364 4784 17924 1371 23228 4087 1753 6948 17924 11726 6 10717 5295 5731 4730 15447	DIHYDROPTEROATE SYNTHASE(EC 2.5.1.15) DIHYDROPTEROATE SYNTHASE(EC 2.5.1.15) DIHYDROFOLATE REDUCTASE(EC 1.5.1.3) FOLYLPOLYGLUTAMATE SYNTHASE(EC 6.3.2.17) 2-AMINO-4-HYDROXY-6-HYDROXYMETHYLDIHYDROPTERIDINE PYROPHOSPHOKINASE(EC 2.7.6.3) PARA-AMINOBENZOATE SYNTHASE COMPONENT I(EC 4.1.3.-) PARA-AMINOBENZOATE SYNTHASE GLUTAMINE AMIDOTRANSFERASE COMPONENT II(EC 4.1.3.-)/ANTHRANILATE SYNTHASE COMPONENT II(EC 4.1.3.27) 4-AMINO-4-DEOXYCHORISMATE LYASE(EC 4.-.-.-) DIHYDROFOLATE REDUCTASE(EC 1.5.1.3) 5-METHYLTETRAHYDROFOLATE-HOMOCYSTEINE METHYLTRANSFERASE (EC 2.1.1.13) 5-METHYLTETRAHYDROFOLATE-HOMOCYSTEINE METHYLTRANSFERASE (EC 2.1.1.13) 5-METHYLTETRAHYDROPTEROYLTTIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRABSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROFOLATE-HOMOCYSTEINE METHYLTRANSFERASE (EC 2.1.1.13) PROTEIN INVOLVED IN FOLATE METABOLISM MEMBRANE SPANNING PROTEIN INVOLVED IN FOLATE METABOLISM ATP-BINDING PROTEIN INVOLVED IN FOLATE METABOLISM
691 693 695 697 699 701 703 705 707 709 711 713 715 717 719 721 723 725 727 729 731	692 694 696 69B 700 702 704 706 708 710 712 714 716 718 720 722 724 726 728 730 732						SEQ ID NO	SEQ ID NO

Molybdenum butterflies make metabolic

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN02802 FRXA02802 FRXA00438 RXN00437 FRXA00437 RXN00439 FRXA00439 FRXA00442	VV0112 GR00783 GR00103 VV0112 GR00103 VV0112 GR00104 GR00105	17369 7 362 178224 3 18742 2 830	16299 474 796 17369 362 18275 196 1087	MOLYBDOPTERIN BIOSYNTHESIS MOEB PROTEIN MOLYBDOPTERIN BIOSYNTHESIS MOEB PROTEIN MOLYBDOPPTEIN BIOSYNTHESIS MOEB PROTEIN MOLYBDOPTERIN(MPT)CONVERTING FACTOR，SUBUNIT2 MOLYBDOPTERIN(MPT)CONVERTING FACTOR，SUBUNIT2 MOLYBDOPTERIN CO-FACTOR SYNTHESIS PROEIIN MOLYBDOPTERIN CO-FACTOR SYNTHESIS PROTEIN MOLYBDOPTERIN CO-FACTOR SYNTHESIS PROTEIN
733 735 737 39 741 743 745 747	734 736 738 740 742 744 746 748						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA00440 RXN00441 FRXA00441 RXN02085 FRXA02085 FRXA02086 RXN02648 FRXA02648 FRXA02658 RXA01516 RXA01515 RXA02024 RXA01719 RXA01720 RXS03223 FRXA01970 RXA02629 RXA02318 RXA01517 RXN01304 RXS02556 RXS02560	GR00104 VV0112 GR00105 GR00629 GR00629 GR00751 GR00752 GR00424 GR00424 GR00613 GR00488 GR00488 GR00568 GR00748 GR00665 GR00424 VV0148	196 19942 2 3496 5252 5254 14764 22360 21513 4026 1264 2476 2 1274 9684 22752 4449	654 18779 793 5295 5731 4730 15447 22749 22364 4784 704 1268 1207 690 9962 23228 4934	MOLYBDENUM COFACTOR BIOSYNTHESIS PROTEIN CB MOLYBDOPTERIN CO-FACTOR SYNTHESIS PROTEIN MOLYBDOPTERIN CO-FACTOR SYNTHESIS PROTEIN 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) 5-METHYLTETRAHYDROPTEROYLTRIGLUTAMATE-HOMOCYSTEINE METHYLTRANSFERASE(EC 2.1.1.14) DIHYDRONEOPTERIN ALDOLASE(EC 4.1.2.25) DIHYDROPTEROATE SYNTHASE(EC 2.5.1.15) DIHYDROPTEROATE SYNTHASE(EC 2.5.1.15) MOLYBDOPTERIN-GUANINE DINUCLEOTIDE BIOSYNTHESIS PROTEIN A MOLYBDOPTERIN BIOSYNTHESIS MOEA PROTEIN MOLYBDOPTERIN BIOSYNTHESIS MOEA PROTEIN MOLYBDOPTERIN BIOSYNTHESIS MOEA PROTEIN MOLYBDOPTERIN BIOSYNTHESIS CNX1 PROTEIN (D90909)pterin-4a-carbinolamine dehydratase[Synechocystis sp.) 2-AMINO-4-HYDROXY-6-HYDROXYMETHYLDIHYDROPTERIDINE PYROPHOSPHOKINASE(EC 2.7.6.3) MOLYBDOPTERIN BIOSYNTHESIS MOG PROTEIN FLAVOHEMOPROTEIN/DIHYDROPTERIDINE REDUCTASE (EC 1.6.99.7) OXYGEN-INSENSITIVE NAD(P)H NITROREDUCTASE(EC 1.-.-.-)/ DIHYDROPTERIDINE REDUCTASE(EC1.6.99.7)
749 751 753 755 757 759 761 763 765 767 769 771 773 775 777 779 781 783 785 787 789 791	750 752 754 756 758 760 762 764 756 768 770 772 774 776 778 780 782 784 786 788 790 792						SEQ ID NO	SEQ ID NO

Vitamin B₁₂Porphyrin and heme metabolism in

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA00382 RXA00156 RXA00624 RXA00306 RXA00884 RXN02503 FRXA02503 RXA00377 RXN02504 FRXA02504	GR00082 GR00023 GR00163 GR00051 GR00242 VV0007 GR00720 GR00081 VV0007 GR00720	2752 10509 7910 2206 10137 22456 16906 1427 22805 17379	1451 9400 8596 1274 11276 22854 17340 306 23362 17816	GLUTAMATE-1-SEMIALDEHYDE 2.1-AMINOMUTASE(EC 5.4.3.8) FERROCHELATASE(EC 4.99.1.1) FERROCHELATASE(EC 4.99.1.1) HEMK PROTEIN OXYGEN-INDEPENDENT COPROPORPHYRINOGEN III OXIDASE(EC 1.-.-.-) PORPHOBILINOGEN DEAMINASE(EC 4.3.1.8) PORPHOBILINOGEN DEAMINASE(EC 4.3.1.8) UROPORPHYRINOGEN DECARBOXYLASE(EC 4.1.1.37) PORPHOBILINOGEN DEAMINASE(EC 4 3.1.8) PORPHOBILINOGEN DEAMINASE(EC 4 3.1.8)
793 795 797 799 801 803 805 807 809 811	794 796 798 800 802 804 806 808 810 812						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN01162 FRXA01162 RXA01692 RXN00371 FRXA00371 FRXA00374 RXN00383 FRXA00376 FRXA00383 RXA01253 RXA02134 RXA02135 RXA02136 RXN03114 RXN01810 RXS03205 FRXA00306 RXC01715	VV0088 GR00330 GR00474 VV0226 GR00078 GR00079 VV0223 GR00081 GR00082 GR00365 GR00639 GR00639 GR00639 VV0088 VV0082	1849 1248 1498 4180 929 1102 4206 287 3876 2536 1721 2809 3362 1 1739	524 4 749 5973 6 371 2863 6 2863 1787 801 1736 2841 552 663	PRECORRIN-6Y METHYLASE(EC 2.1.1.-) PRECORRIN-6Y METHYLASE(EC 2.1.1.-) UROPORPHYRIN-III C-METHYLTRANSFERASE(EC 2.1.1.107) UROPORPHYRIN-III C-METHYLTRANSFERASE(EC 2.1.1.107)/ UROPORPHYRINOGEN-III SYNTHASE(EC 4.2.1.75) UROPORPHYRIN-III C-METHYLTRANSFERASE(EC 2.1.1.107)/ UROPORPHYRINOGEN-III SYNTHASE(EC 4.2.1.75) UROPORPHYRIN-III C-METHYLTRANSFERASE(EC 2.1.1.107)/ UROPORPHYRINOGEN-III SYNTHASE(EC 4.2 1.75) PROtopORPHYRINOGEN OXIDASE(EC 1.3.3.4) PROtopORPHYRINOGEN OXIDASE(EC 1.3.3 4) PROtopORPHYRINOGEN OXIDASE(EC 1.3.3.4) COBYRIC ACID SYNTHASE COBALAMIN(5′-PHOSPHATE)SYNTHASE NICOTINATE-NUCLEOTIDE--DIMETHYLBENZIMIDAZOLE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.21) COBINAMIDE KINASE/COBINAMIDE PHOSPHATE GUANYLYLTRANSFERASE COBG PROTEIN(EC 1.-.-.-) HEMIN-BINDING PERIPLASMIC PROTEIN HMUT PRECURSOR HEMK PROTEIN HEMK PROTEIN CYTOSOLIC PROTEIN INVOLVED IN PORPHYRIN METABOLISM
813 815 817 819 821 823 825 827 829 831 833 835 837 839 841 843 845 847	814 816 818 820 822 824 826 828 830 832 834 836 838 840 842 844 846 848						SEQ ID NO	SEQ ID NO

Precursor of vitamin C

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN00420 FRXA00420 FRXA00426 RXN00708 FRXA00708 RXA02373 RXS00389 RXS00419 RXC00416 RXC02206	VV0112 GR00096 GR00097 VV0005 GR00185 GR00688	2511 2 1737 4678 2030 1540	1048 541 2258 3872 1359 626	L-GULONOLACTONE OXIDASE(EC 1.1.3.8) L-GULONOLACTONE OXIDASE(EC 1.1.3.8) L-GULONOLACTONE OXIDASE(EC 1.1.3.8) 2，5-DIKETO-D-GLUCONIC ACID REDUCTASE(EC 1.1.1.-) 2，5-DIKETO-D-GLUCONIC ACID REDUCTASE(EC 1.1.1.-) 2，5-DIKETO-D-GLUCONIC ACID REDUCTASE(EC 1.1.1.-) oxoglutarate semialdehyde dehydrogenase(EC 1.2.1.-) ACETOACETYL-COA REDUCTASE(EC 1.1.1.36) MEMBRANE SPANNING PROTEIN INVOLVED IN METABOLISM OF VITAMIN C PRECURSORS OXIDOREDUCTASE INVOLVED IN METABOLISM OF VITAMIN C PTECURSORS
849 851 853 855 857 859 861 863 865 867	850 852 854 856 858 860 862 864 866 868						SEQ ID NO	SEQ ID NO

Vitamin K2

Nucleic acid	Amino acids	Numbers starting NT NT contig termination
Nucleic acid	Amino acids	Numbers starting NT NT contig termination	SEQ ID NO	SEQ ID NO	RXS03074 S-ADENOSYLMETHIONINE：2-DEMETHYLMENAQUINONE METHYLTRANSFERASE(EC 2.1.-.-)
869	870		SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	FRXA02906 RXA02315 RXA02319 RXS00393 FRXA00393 RXA00391 RXS02908	GR10044 GR00665 GR00665 GR00086 GR00086	1142 8011 9977 4030 2031	645 6383 10933 4911 2750	S-ADENOSYLMETHIONINE：2-DEMETHYLMENAQUINONE METHYLTRANSFERASE(EC 2.1.-.-) 2-SUCCINYL-6-HYDROXY-2，4-CYCLOHEXADIENE-1-CARBOXYLATE SYNTHASE/2-OXOGLUTARATEDECARBOXYLASE(EC 4 1.1.71) NAPHTHOATE SYNTHASE(EC 4.1.3-36) 1，4-DIHYDROXY-2-NAPHTHOATE OCTAPRENYLTRANSFERASE(EC 2.5.-.-) 1，4-DIHYDROXY-2-NAPHTHOATE OCTAPRENYLTRANSFERASE(EC 2 5.-.-) O-SUCCINYLBENZOIC ACID-COA LIGASE(EC 6.2.1.26) O-SUCCINYLBENZOIC ACID-COA LIGASE(EC 6.2.1.26)
871 873 875 877 879 881 883	872 874 876 878 880 882 884						SEQ ID NO	SEQ ID NO

Ubiquinone biosynthesis

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA00997 RXA02189 RXA02311 RXN02912 RXS00998	GR00283 GR00642 GR00665 VV0135	2389 986 3073 13299	1808 249 2384 12547	3-DEMETHYLUBIQUINONE-9 3-METHYLTRANSFERASE(EC 2.1 1.64) 3-DEMETHYLUBIQUINONE-9 3-METHYLTRANSFERASE(EC 2 1.1.64) 3-DEMETHYLUBIQUINONE-9 3-METHYLTRANSFERASE(EC 2.1.1.64) UBIQUINONE/MENAQUINONE BIOSYNTHESIS METHLYTRANSFERASE UBIE (EC 2.1.1.-) COMA OPERON PROTEIN 2
885 887 889 891 893	886 888 890 892 894						SEQ ID NO	SEQ ID NO

Purine, pyrimidine, and other nucleotides

Purine, pyrimidine biosynthetic pathway regulation

Purine Metabolism

Purine biosynthesis

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA01215 RXN00558 FRXA00558 RXN00626 FRXA00629 FRXA00626 RXA02623 RXA01442	GR00352 VV0103 GR00148 VV0135 GR00165 GR00164 GR00746 GR00418	1187 8235 61 11624 1450 1 4875 10277	213 9581 501 10362 1713 780 4285 9054	RIBOSE-PHOSPHATE PYROPHOSPHOKINASE，PRPP synthetase(EC 2.7.6.1) AMIDOPHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.14) AMIDOPHOSPHORIBOSYLTRANSFERASE(EC 2.4 2.14) PHOSPHORIBOSYLAMINE-GLYCINE LIGASE(EC 6 3.4.13) PHOSPHORIBOSYLAMINE-GLYCINE LIGASE(EC 6.3.4.13) PHOSPHORIBOSYLAMINE-GLYCINE LIGASE.GARS(EC 6.3.4.13) PHOSPHORIBOSYLAMINE-GLYCINE LIGASE(EC 6.3.4.13)/ PHOSPHORIBOSYLFORMYLGLYCINAMIDINE CYCLO-LIGASE(EC 6.3 31)/ PHOSPHORIBOSYLGLYCINAMIDE FORMYLTRANSFERASE(EC 2.1.2.2) PHOSPHORIBOSYLGLYCINAMIDE FORMYLTRANSFERASE 2(EC 2.1.2-)
895 897 899 901 903 905 907 909	896 898 900 902 904 906 908 910						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO 911 913 915 917 919 921 923 925 927 929 931 933 935 937 939 941 943	SEQ ID NO 912 914 916 918 920 922 924 926 928 930 932 934 936 938 940 942 944	RXN00537 FRXA02805 FRXA00537 FRXA00561 RXA00541 RXA00620 RXN00770 FRXA00557 FRXA00770 RXN02345 FRXA02345 RXN02350 FRXA02346 FRXA02350 RXA01087 RXA00619 RXA02622	VV0103 GR00786 GR00138 GR00150 GR00139 GR00163 VV0103 GR00147 GR00204 VV0078 GR00676 VV0078 GR00677 GR00678 GR00304 GR00163 GR00746	3351 54 23 2 2269 3049 9614 15 7809 4788 1534 8369 127 1120 498 793 4274	5636 638 697 280 2937 3939 10783 818 7495 5984 725 8863 5 911 1373 2220 2715	PHOSPHORIBOSYLFORMYLGLYCINAMIDINE SYNTHASE(EC 6.3.5.3) PHOSPHORIBOSYLFORMYLGLYCINAMIDINE SYNTHASE(EC 6.3.5.3) PHOSPHORIBOSYLFORMYLGLYCINAMIDINE SYNTHASE(EC 6.3.5.3) PHOSPHORIBOSYLFORMYLGLYCINAMIDINE SYNTHASE(EC 6.3.5.3) PHOSPHORIBOSYLFORMYLGLYCINAMIDINE SYNTHASE(EC 6.3.5.3) PHOSPHORIBOSYLAMIDOIMIDAZOLE-SUCCINOCARBOXAMIDE SYNTHASE (EC 6.3.2.6) PHOSPHORIBOSYLFORMYLGLYCINAMIDINE CYCLO-LIGASE(EC 6.3.3.1) PHOSPHORIBOSYLFORMYLGLYCINAMIDINE CYCLO-LIGASE(EC 6.3.3.1) PHOSPHORIBOSYLFORMYLGLYCINAMIDINE CYCLO-LIGASE(EC 6.3.3.1) PHOSPHORIBOSYLAMINOIMIDAZOLE CARBOXYLASE ATPASE SUBUNIT(EC 4.1.1.21) PHOSPHORIBOSYLAMINOIMIDAZOLE CARBOXYLASE ATPASE SUBUNIT(EC 4.1.1.21) PHOSPHORIBOSYLAMINOIMIDAZOLE CARBOXYLASE CATALYTIC SUBUNIT (EC 4.1.1.21) PHOSPHORIBOSYLAMINOIMIDAZOLE CARBOXYLASE CATALYTIC SUBUNIT (EC4.1.1.21) PHOSPHORIBOSYLAMINOIMIDAZOLE CARBOXYLASE CATALYTIC SUBUNIT (EC 4.1.1.21) PHOSPHORIBOSYLAMINOIMIDAZOLE CARBOXYLASE(EC 4.1.1.21) ADENYLOSUCCINATE LYASE(EC 4.3.2.2) PHOSPHORIBOSYLAMINOIMIDAZOLECARBOXAMIDE FORMYLTRANSFERASE (EC 2.1.2.3)/IMP CYCLOHYDROLASE(EC 3.5.4.10)

GMP, GDP, AMP and ADP synthesis, starting from inosine-5'-monophosphate (IMP)

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO 945 947 949 951 953 955 957 959 961 963 965	SEQ ID NO 946 948 950 952 954 956 958 960 962 964 966	RXN00488 FRXA00492 FRXA00488 RXA02469 RXN00487 FRXA00487 RXA02237 RXA01446 RXA00619 RXA00688 RXA00266	VV0086 GR00122 GR00121 GR00715 VV0086 GR00120 GR00654 GR00418 GR00163 GR00179 GR00040	19066 1171 1 1927 23734 712 4577 17765 793 10443 3769	20583 1644 534 497 25302 2097 5146 16476 2220 10985 3362	INOSINE-5′-MONOPHOSPHATE DEHYDROGENASE(EC 1.1.1.205) INOSINE-5′-MONOPHOSPHATE DEHYDROGENASE(EC 1.1.1 205) INOSINE-5′-MONOPHOSPHATE DEHYDROGENASE(EC 1.1.1.205) INOSINE-5′-MONOPHOSPHATE DEHYDROGENASE(EC 1.1.1.205) GMP SYNTHASE[GLUTAMINE-HYDROLYZING](EC 6.3.5.2) GMP SYNTHASE(EC 6.3.4.1) GUANYLATE KINASE(EC 2.7.4.8) ADENYLOSUCCINATE SYNTHETASE(EC 6.3.4.4) ADENYLOSUCCINATE LYASE(EC 4.3.2.2) ADENYLATE KINASE(EC 2.7.4.3) NUCLEOSIDE DIPHOSPHATE KINASE(EC 2.7-4.6)

Table 1 (continued)

GMP / AMP degradation activity

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO 967 969 971	SEQ ID NO 968 970 972	RXA00489 RXN02281 FRXA02281	GR00121 VV0152 GR00659	654 1893 1101	1775 3323 34	GMP REDUCTASE(EC 1.6.6.8) AMP NUCLEOSIDASE(EC 3.2.2.4) AMP NUCLEOSIDASE(EC 3.2.2.4)

Pyrimidine metabolic

De novo pyrimidine biosynthesis:

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA00147 RXA00145 RXA00146 RXA02208 RXA01660 RXA02235 RXN01892 FRXA01892 RXA00105 RXA00131 RXA00266 RXA00718 RXA01599 RXN02234 FRXA02234 RXN00450 FRXA00450 RXN02272 FRXA02272 RXN03004 RXN03137 RXN03171 FRXA02857	GR00022 GR00022 GR00022 GR00647 GR00462 GR00654 VV0150 GR00542 GR00014 GR00020 GR00040 GR00188 GR00447 VV0134 GR00654 VV0112 GR00110 VV0020 GR00655 VV0237 VV0129 VV0328 GR10003	9722 7258 8249 2 591 3207 3020 47 16672 7621 3769 4576 8780 24708 1 34491 322 15566 6691 1862 9680 568 570	10900 8193 9589 1003 1142 4040 3748 775 17346 7013 3362 5283 10441 28046 3198 34814 5 16810 7935 2341 9579 1080 1082	CARBAMOYL-PHOSPHATE SYNTHASE SMALL CHAIN(EC 6.3.5.5) ASPARTATE CARBAMOYLTRANSFERASE CATALYTIC CHAIN(EC 2.1.3.2) DIHYDROOROTASE(EC 3.5.2.3) DIHYDROOROTATE DEHYDROGENASE(EC 1.3.3.1) OROTATE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.10) OROTIDINE 5′-PHOSPHATE DECARBOXYLASE(EC 4.1.1.23) URIDYLATE KINASE(EC 2.7.4.-) URIDYLATE KINASE(EC 2.7.4.-) THYMIDYLATE SYNTHASE(EC 2.1.1.45) THYMIDYLATE KINASE(EC 2.7.4.9) NUCLEOSIDE DIPHOSPHATE KINASE(EC 2.7.4.6) CYTIDYLATE KINASE(EC 2.7.4.14) CTP SYNTASE(EC 6.3.4.2) CARBAMOYL-PHOSPHATE SYNTASE LARGE CHAIN(EC 6.3.5.5) CARBAMOYL-PHOSPHATE SYNTASE LARGE CHAIN(EC 6.3.5.S) CYTOSINE DFAMINASE(EC 3 5.4.1) CYTOSINE DEAMINASE(EC 3.5.4.1) CYTOSINE DEAMINASE(EC 3.5.4.1) CREATININE DEAMINASE(EC 3.5.4.21) DEOXYCYTIDINE TRIPHOSPHATE DEAMINASE(EC 3.5.4.13) THYMIDYLATE SYNTHASE(EC 2.1.1.45) URACIL PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.9) URACIL PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.9)
973 975 977 979 981 983 985 987 989 991 993 995 997 999 1001 1003 1005 1007 1009 1011 1013 1015 1017	974 976 978 980 982 984 986 988 990 992 994 996 998 1000 1002 1004 1006 1008 1010 1012 1014 1016 1018						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Purine and pyrimidine bases, nucleotides and nucleosides remedy mutual transformation, reduction and degradation:

Purine:

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA02771 RXA01512 RXA02031 RXA00981 RXN02772 FRXA02772 FRXA02773 RXA01835 RXA01483 RXN01027 FRXA01024 FRXA01027 RXA01528 RXA00072 RXA01878 RXN02281 FRXA02281 RXN01240 RXN02008	GR00772 GR00424 GR00618 GR00276 VV0171 GR00772 GR00772 GR00517 GR00422 VV0143 GR00293 GR00294 GR00425 GR00012 GR00537 VV0152 GR00659 VV0090 VV0171	1329 17633 3820 3388 2045 1962 2741 3147 19511 5761 661 2580 5653 446 1239 1893 1101 30442 1138	1883 18232 3347 4017 1011 2741 2902 3677 18240 6768 5 2347 5126 6 2117 3323 34 29420 5	ADENINE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.7) HYPOXANTHINE-GUANINE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.8) XANTHINE-GUANINE PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.22) GTP PYROPHOSPHOKINASE(EC 2.7.6.5) GUANOSINE-3′，5′-BIS(DIPHOSPHATE)3′-PYROPHOSPHOHYDROLASE(EC 3.1.7.2) GUANOSINE-3′，5′-BIS(DIPHOSPHATE)3′-PYROPHOSPHOHYDROLASE(EC 3.1.7.2) GUANOSINE-3′，5′-BIS(DIPHOSPHATE)3′-PYROPHOSPHOHYDROLASE(EC 3.1.7.2) GUANOSINE-3′，5′-BIS(DIPHOSPHATE)3′-PYROPHOSPHOHYDROLASE(EC 3.1.7.2) DEOXYGUANOSINETRIPHOSPHATE TRIPHOSPHOHYDROLASE(EC 3.1.5.1) DIADENOSINE 5′，5_-P1，P4-TETRAPHOSPHATE HYDROLASE(EC 3.6.1.17) DIADENOSINE 5′，5_-P1，P4-TETRAPHOSPHATE HYDROLASE(EC 3.6 1.17) DIADENOSINE 5′，5_-P1，P4-TETRAPHOSPHATE HYDROLASE(EC 3.6.1.17) DIADENOSINE 5′，5_-P1，P4-TETRAPHOSPHATE HYDROLASE(EC 3.6.1.17) PHOSPHOADENOSINE PHOSPHOSULFATE REDUCTASE(EC 1.8.99.4) DIMETHYLADENOSINE TRANSFERASE(EC 2.1.1.-) AMP NUCLEOSIDASE(EC 3.2.2.4) AMP NUCLEOSIDASE(EC 3.2.2.4) GTP PYROPHOSPHOKINASE(EC 2.7.6.5) GUANOSINE-3′，5′-BIS(DIPHOSPHATE)3′-PYROPHOSPHOHYDROLASE(EC 3.1.7.2)
1019 1021 1023 1025 1027 1029 1031 1033 1035 1037 1039 1041 1043 1045 1047 1049 1051 1053 1055	1020 1022 1024 1026 1028 1030 1032 1034 1036 1038 1040 1042 1044 1046 1048 1050 1052 1054 1056						SEQ ID NO	SEQ ID NO

Pyrimidine and purine metabolism:

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN01940 FRXA01940 RXA02559 RXA02497 RXN01079 FRXA01079 FRXA01084 RXN01920 FRXA01920 RXA01080 RXA00867 RXA01416 RXA01486	VV0120 GR00557 GR00731 GR00720 VV0084 GR00301 GR00302 VV0084 GR00550 GR00301 GR00237 GR00413 GR00423	10268 3 5418 10059 38084 693 3402 32843 1321 1240 1 2 660	9333 581 6320 10685 35982 4 2062 31842 908 797 627 631 4	INOSINE-URIDINE PREFERRING NUCLEOSIDE HYDROLASE(EC 3.2.2.1) INOSINE-URIDINE PREFERRING NUCLEOSIDE HYDROLASE(EC 3.2 2.1) INOSINE-URIDINE PREFERRING NUCLEOSIDE HYDROLASE(EC 3.2.2.1) EXOPOLYPHOSPHATASE(EC 3.6.1.11) RIBONUCLEOSIDE-DIPHOSPHATE REDUCTASE ALPHA CHAIN(EC 1.1.7.4.1) RIBONUCLEOSIDE-DIPHOSPHATE REDUCTASE ALPHA CHAIN(EC 1.1.7.4.1) RIBONUCLEOSIDE-DIPHOSPHATE REDUCTASE ALPHA CHAIN(EC 1.1.7.4.1) RIBONUCLEOSIDE-DIPHOSPHATE REDUCTASE 2BETA CHAIN(EC 1.1.7 4.1) RIBONUCLEOTIDE REDUCTASE SUBUNITR2F NRDI PROTEIN POLYRIBONUCLEOTIDE NUCLEOTIDYLTRANSFERASE(EC 2.7.7.8) POLYRIBONUCLEOTIDE NUCLEOTIDYLTRANSFERASE(EC 2.7.7.8) POLYRIBONUCLEOTIDE NUCLEOTIDYLTRANSFERASE(EC 2.7.7.8)
1057 1059 1061 1063 1065 1067 1069 1071 1073 1075 1077 1079 1081	1058 1060 1062 1064 1066 1068 1070 1072 1074 1076 1078 1080 1082						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Nucleic acid	Amino acids	Number	Contigs starting NT NT termination
Nucleic acid	Amino acids	Number	Contigs starting NT NT termination	SEQ ID NO	SEQ ID NO	RXA01678 RXA01679 RXN01488 RXC00540 RXC00560 RXC01088 FXC02624 RXC02665 RXC02770 FRXC02238 RXC01946	GR00467 7162 7689 2′，3′-CYCLIC-NUCLEOTIDE 2′-PHOSPHODIESTERASE(EC 3.1 4.16) GR00467 7729 8964 2′，3′-CYCLIC-NUCLEOTIDE 2′-PHOSPHODIESTERASE(EC 3.1.4.16) VV0139 39842 40789 INOSINE-URIDINE PREFERRING NUCLEOSIDE HYDROLASE(EC 3.2.2.1) CYTOSOLIC PROTEIN INVOLVED IN PURINE METABOLISM PROTEIN INVOLVED IN PURINE METABOLISM CYTOSOLIC PROTEIN INVOLVED IN PURINE METABOLISM MEMBRANE SPANNING PROTEIN INVOLVED IN PURINE METABOLISM PROTEIN INVOLVED IN PURINE METABOLISM LIPOPROTEIN INVOLVED IN PURINE METABOLISM PROTEIN INVOLVED IN METABOLISM OF S-ADENOSYLMETHIONINE，PURINES AND PANTOTHENATE ABC TRANSPORTER ATP-BINDING PROTEIN INVOLVED IN PURINE METABOLISM
1083 1085 1087 1089 1091 1093 1095 1097 1099 1101 1103	1084 1086 1088 1090 1092 1094 1096 1098 1100 1102 1104			SEQ ID NO	SEQ ID NO

Pyrimidine:

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXN03171 FRXA02857 RXN00450 FRXA00450 RXA00465 RXA00717 RXA01894 RXA02536 RXN01209 FRXA01209 RXN01617 FRXA01617 RXC01600 RXC01622 RXC00128 RXC01709 RXC00207	VV0328 GR10003 VV0112 GR00110 GR00117 GR00188 GR00542 GR00726 VV0270 GR00348 VV0050 GR00451	568 570 34491 322 337 3617 1622 8581 1019 1019 22187 2	1080 1082 34814 5 828 4576 2476 7826 2446 2446 22858 616	URACIL PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.9) URACIL PHOSPHORIBOSYLTRANSFERASE(EC 2.4.2.9) CYTOSINE DEAMINASE(EC 3.5.4.1) CYTOSINE DEAMINASE(EC 3.5.4.1) CYTOSINE DEAMINASE(EC 3.5.4.1) RIBOSOMAL LARGE SUBUNIT PSEUDOURIDINE SYNTHASE B(EC 4.2.1.70) PHOSPHATIDATE CYTIDYLYLTRANSFERASE(EC 2.7.7.41) BETA-UREIDOPROPIONASE(EC 3.5.1.6) PHOSPHOMETHYLPYRIMIDINE KINASE(EC 2.7.4.7) PHOSPHOMETHYLPYRIMIDINE KINASE(EC 2.7.4.7) PHOSPHOMETHYLPYRIMIDINE KINASE(EC 2.7.4.7) PHOSPHOMETHYLPYRIMIDINE KINASE(EC 2.7.4.7) CYTOSOLIC PROTEIN INVOLVED IN PYRIMIDINE METABOLISM CYTOSOLIC PROTEIN INVOLVED IN PYRIMIDINE METABOLISM EXPORTED PROTEIN INVOLVED IN METABOLISM OF PYRIDIMES AND ADENOSYLHOMOCYSTEINE CYTOSOLIC PROTEIN INVOLVED IN PYRIMIDINE METABOLISM EXPORTED PROTEIN INVOLVED IN PYRIMIDINE METABOLISM
1105 1107 1109 1111 1113 1115 1117 1119 1121 1123 1125 1127 1129 1131 1133 1135 1137	1106 1108 1110 1112 1114 1116 1118 1120 1122 1124 1126 1128 1130 1132 1134 1136 1138						SEQ ID NO	SEQ ID NO

Table 1 (continued)

Sugar

Trehalose

Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function
Nucleic acid	Amino acids	Number	Contigs	NT Start	NT termination	Function	SEQ ID NO	SEQ ID NO	RXA00347 RXN01239 FRXA01239 RXA02645 RXN02355 RXN02909 RXS00349 RXS03183 RXC00874	GR00065 VV0090 GR00358 GR00751 VV0051 VV0135	246 32921 5147 714 735 38532	1013 30489 7579 2543 4 39017	TREHALOSE-PHOSPHATASE(EC 3.1.3.12) maltooligosyltrehalose synthase maltooligosyltrehalose synthase maltooligosyltrehalose trehalohydrolase TREHALOSE/MAL TOSE BINDING PROTEIN Hypothetical Trehalose-Binding Protein Hypothetical TrehaIose Transport Protein TREHALOSE/MALTOSE BINDING PROTEIN TRANSMEBRANE PROTEIN INVOLVED IN TREHALOSE METABOLISM
1139 1141 1143 1145 1147 1149 1151 1153 1155	1140 1142 1144 1146 1148 1150 1152 1154 1156						SEQ ID NO	SEQ ID NO

Table 2 - exclude genes
Table 2 - exclude genes				GenBank ^TMAccession number	Gene Name	Gene function	Literature
A09073	ppg	Phosphoenol pyruvate carboxylase	Bachmann，B.et al.“DNA fragment coding for phosphoenolpyruvat corboxylase，recombinant DNA carrying said fragment，strains carrying the recombinant DNA and method for producing L-aminino acids using said stains，”Patent：EP 0358940-A 3 03/21/90	GenBank ^TMAccession number	Gene Name	Gene function	Literature
A09073	ppg	Phosphoenol pyruvate carboxylase		A45579， A4558I， A45583， A45585 A45587		Threonine dehydratase	Moeckel，B.et al.“Production of L-isoleucine by means of recombinant micro-organisms with deregulated threonine dehydratase，”Patent：WO 9519442-A 5 07/20/95
AB003132	murC；ftsQ；ftsZ		Kobayashi，M.et al.“Cloning，sequencing，and characterization of the ftsZ gene from coryneform bacteria，” Biochem.Biophys.Res.Commun.， 236(2)：383-388(1997)	A45579， A4558I， A45583， A45585 A45587		Threonine dehydratase
AB003132	murC；ftsQ；ftsZ			AB015023	murC；ftsQ		Wachi，M.et al.“A murC gene from Coryneform bacteria，”Appl.Microbiol. Biotechnol，51(2)：223-228(1999)
AB018530	dtsR		Kimura，E.et al.“Molecular cloning of a novel gene，dtsR，which rescues the detergent sensitivity of a mutant derived from Brevibacterium lactofermentum，”Biosci.Biotechnol.Biochem.，60(10)：1565-1570(1996)	AB015023	murC；ftsQ
AB018530	dtsR			AB018531	dtsR1；dtsR2
AB020624	murl	D-glutamate racemase		AB018531	dtsR1；dtsR2
AB020624	murl	D-glutamate racemase		AB023377	tkt	transketolase
AB024708	gltB；gltD	GIutamine 2-oxoglutatate aminotransferase large and small subunits		AB023377	tkt	transketolase
AB024708	gltB；gltD			AB025424	acn	aconitase
AB027714	rep	Replication protein		AB025424	acn	aconitase
AB027714	rep	Replication protein		AB027715	rep；aad	Replication protein；aminoglycoside adenyltransferase
AF005242	argC	N-acetylglutamate-5-semialdehyde dehydrogenase		AB027715	rep；aad	Replication protein；aminoglycoside adenyltransferase
AF005242	argC	N-acetylglutamate-5-semialdehyde dehydrogenase		AF005635	glnA	Glutamine synthetase
AF030405	hisF	cyclase		AF005635	glnA	Glutamine synthetase
AF030405	hisF	cyclase		AF030520	argG	Argininosuccinate synthetase
AF031518	argF	Omithine carbamolytransferase		AF030520	argG	Argininosuccinate synthetase
AF031518	argF	Omithine carbamolytransferase		AF036932	aroD	3-dehydroquinate dehydratase
AF038548	pyc	Pyruvate carboxylase		AF036932	aroD	3-dehydroquinate dehydratase
AF038548	pyc	Pyruvate carboxylase

Table 2 (continued)
Table 2 (continued)				AF038651	dciAE；apt；rel	Dipeptide-binding protein；adenine phosphoribosyltransferase；GTP pyrophosphokinase	Wehmeier，L.et al.“The role of the Corynebacterium glutamicum rel gene in (p)ppGpp metabolism，”Microbiology，144：1853-1862(1998)
AF041436	argR	Arginine repressor		AF038651	dciAE；apt；rel
AF041436	argR	Arginine repressor		AF045998	impA	Inositol monophosphate phosphatase
AF048764	argH	Argininosuccinate lyase		AF045998	impA	Inositol monophosphate phosphatase
AF048764	argH	Argininosuccinate lyase		AF049897	argC；argJ；argB； argD；argF；argR； argG；argH	N-acetylglutamylphosphate reductase； omithine acetyltransferase；N- acetylglutamate kinase；acetylomithine transminase；omithine carbamoyltransferase；arginine repressor； argininosuccinate synthase； argininosuccinate lyase
AF050109	inhA	Enoyl-acyl carrier protein reductase		AF049897	argC；argJ；argB； argD；argF；argR； argG；argH
AF050109	inhA	Enoyl-acyl carrier protein reductase		AF050166	hisG	ATP phosphoribosyltransferase
AF051846	hisA	Phosphoribosylformimino-5-amino-1- phosphoribosyl-4-imidazolecarboxamide isomerase		AF050166	hisG	ATP phosphoribosyltransferase
AF051846	hisA			AF052652	metA	Homoserine O-acetyltransferase	Park，S.et al.“Isolation and analysis of metA，a methionine biosynthetic gene encoding homoserine acetyltransferase in Corynebacterium glutamicum，”Mol. Cells.，8(3)：286-294(1998)
AF053071	aroB	Dehydroquinate synthetase		AF052652	metA	Homoserine O-acetyltransferase
AF053071	aroB	Dehydroquinate synthetase		AF060558	hisH	Glutamine amidotransferase
AF086704	hisE	Phosphoribosyl-ATP- pyrophosphohydrolase		AF060558	hisH	Glutamine amidotransferase
AF086704	hisE	Phosphoribosyl-ATP- pyrophosphohydrolase		AF114233	aroA	5-enolpyruvylshikimate 3-phosphate synthase
AF116184	panD	L-aspartate-alpha-decarboxylase precursor	Dusch，N.et al.“Expression of the Corymebacterium glutamicum pamD gene encoding L-aspartate-alpha-decarboxylase leads to pantothenate overproduction in Escherichia coli，”Appl.Environ.Microbiol，65(4)1530- 1539(1999)	AF114233	aroA	5-enolpyruvylshikimate 3-phosphate synthase
AF116184	panD	L-aspartate-alpha-decarboxylase precursor		AF124518	aroD；aroE	3-dehydroquiniase；shikimate dehydrogenase
AF124600	aroC；aroK；aroB； pepQ	Chorismate synthase；shikimate kinase；3- dehydroquinate synthase；putative cytoplasmic peptidase		AF124518	aroD；aroE	3-dehydroquiniase；shikimate dehydrogenase
AF124600	aroC；aroK；aroB； pepQ			AF145897	inhA
AF145898	inhA			AF145897	inhA

Table 2 (continued)
Table 2 (continued)				AJ001436	ectP	Transport of ectoine，glycine betaine， proline	Peter，H.et al.“Corynebacterium glutamicum is equipped with four secondary carriers for compatible solutes：Identification，sequencing，and characterization of the proline/ectoine uptake system，ProP，and the ectoine/proline/glycine betaine carrier，EctP，”J.Bacteriol.，180(22)：6005-6012(1998)
AJ004934	dapD	Tetrahydrodipicolinate succinylase (incompletei)	Wehrmann，A.et al.“Different modes of diaminopimelate synthesis and their role in cell wall integrity：A study with Corynebacterium glutamicum，”J. Bacteriol.，180(12)：3159-3165(1998)	AJ001436	ectP	Transport of ectoine，glycine betaine， proline
AJ004934	dapD	Tetrahydrodipicolinate succinylase (incompletei)		AJ007732	ppc；secG；amt；ocd； soxA	Phosphoenolpyruvate-carboxylase；？；high affinity ammonium uptake protein；putative omithine-cyclodecarboxylase；sarcosine oxidase
AJ010319	ftsY，glnB，glnD；srp； amtP	Involved in cell division；Pll protein； uridylyltransferase(uridylyl-removing enzmye)；signal recognition particle；low affinity ammonium uptake protein	Jakoby，M.et al.“Nitrogen regulation in Corynebacterium glutamicum； Isolation of genes involved in biochemical characterization of corresponding proteins，”FEMS Microbiol.，173(2)：303-310(1999)	AJ007732	ppc；secG；amt；ocd； soxA
AJ010319	ftsY，glnB，glnD；srp； amtP			AJ132968	cat	Chloramphenicol aceteyl transferase
AJ224946	mqo	L-malate：quinone oxidoreductase	Molenaar，D.et al.“Biochemical and genetic characterization of the membrane-associated malate dehydrogenase(acceptor)from Corynebacterium glutamicum，”Eur.J.Biochem.，254(2)：395-403(1998)	AJ132968	cat	Chloramphenicol aceteyl transferase
AJ224946	mqo	L-malate：quinone oxidoreductase		AJ238250	ndh	NADH dehydrogenase
AJ238703	potA	Porin	Lichtinger，T.et al.“Biochemical and biophysical characterization of the cell wall porin of Corynebacterium glutamicum：The channel is formed by a low molecular mass polypeptide，”Biochemistry，37(43)：15024-15032(1998)	AJ238250	ndh	NADH dehydrogenase
AJ238703	potA	Porin		D17429		Transposable element IS31831	Vertes et al.“Isolation and characterization of IS31831，a transposable element from Corynebacterium glutamicum，”Mol.Microbiol.，11(4)：739-746(1994)
D84102	odhA	2-oxoglutarate dehydrogenase	Usuda，Y.et al.“Molecular cloning of the Corynebacterium glutamicum (Brevibacterium lactofermentum AJ12036)odhA gene encoding a novel type of 2-oxoglutarate dehydrogenase，”Microbiology，142：3347-3354(1996)	D17429		Transposable element IS31831
D84102	odhA	2-oxoglutarate dehydrogenase		E01358	hdh；hk	Homoserine dehydrogenase；homoserine kinase	Katsumata，R.et al.“Production of L-thereonine and L-isoleucine，”Patenl：JP 1987232392-A 1 10/12/87
E01359		Upstream of the start codon of homoserine kinase gene	Katsumata，R.et al.“Production of L-thereonine and L-isoleucine，”Patent：JP 1987232392-A 2 10/12/87	E01358	hdh；hk	Homoserine dehydrogenase；homoserine kinase
E01359		Upstream of the start codon of homoserine kinase gene		E01375		Tryptophan operon
E01376	trpL；trpE	Leader peptide；anthranilate synthase	Matsui，K.et al.“Tryptophan operon，peptide and protein coded thereby， utilization of tryptophan operon gene expression and production of tryptophan，”Patent：JP 1987244382-A 1 10/24/87	E01375		Tryptophan operon
E01376	trpL；trpE	Leader peptide；anthranilate synthase

Table 2 (continued)
Table 2 (continued)			E01377	Promoter and operator regions of tryplophan operon	Matsui，K.et al.“Tryptophan operon，peptide and protein coded thereby， utilization of tryptophan operon gene expression and production of tryptophan，”Patent：JP 1987244382-A 1 10/24/87
E03937	Biotin-synthase	Hatakeyama，K.et al.“DNA fragment containing gene capable of coding biotin synthetase and its utilization，”Patent：JP 1992278088-A 1 10/02/92	E01377	Promoter and operator regions of tryplophan operon
E03937	Biotin-synthase		E04040	Diamino pelargonic acid aminotransferase	Kohama，K.et al.“Gene coding diaminopelargonic acid aminotransferase and desthiobiotin synthetase and its utilization，”Patent：JP 1992330284-A 1 11/18/92
E04041	Desthiobiotinsynthetase	Kohama，K.et al.“Gene coding diaminopelargonic acid aminotransferase and desthiobiotin synthetase and its utilization，”Patent：JP 1992330284-A 1 11/18/92	E04040	Diamino pelargonic acid aminotransferase
E04041	Desthiobiotinsynthetase		E04307	Flavum aspartase	Kurusu，Y.et al.“Gene DNA coding aspartase and utilization thereof，”Patent： JP 1993030977-A 1 02/09/93
E04376	Isocitric acid lyase	Katsumata，R.et al.“Gene manifestation controlling DNA，”Patent：JP 1993056782-A 3 03/09/93	E04307	Flavum aspartase
E04376	Isocitric acid lyase		E04377	Isocitric acid lyase N-terminal fragment	Katsumata，R.et al.“Gene manifestation controlling DNA，”Patent：JP 1993056782-A 3 03/09/93
E04484	Prephenate dehydratase	Sotouchi，N.et al.“Production of L-phenylalanine by fermentation，”Patent：JP 1993076352-A 2 03/30/93	E04377	Isocitric acid lyase N-terminal fragment
E04484	Prephenate dehydratase		E05108	Aspartokinase	Fugono，N.et al.“Gene DNA coding Aspartokinase and its use，”Patent：JP 1993184366-A 1 07/27/93
E05112	Dihydro-dipichorinate synthetase	Hatakeyama，K.et al.“Gene DNA coding dihydrodipicolinic acid synthetase and its use，”Patent：JP 1993184371-A 1 07/27/93	E05108	Aspartokinase
E05112	Dihydro-dipichorinate synthetase		E05776	Diaminopimelic acid dehydrogenase	Kobayashi，M.et al.“Gene DNA coding Diaminopimelic acid dehydrogenase and its use，”Patent：JP 1993284970-A 1 11/02/93
E05779	Threonine synthase	Kohama，K.et al.“Gene DNA coding threonine synthase and its use，”Patent： JP 1993284972-A 1 11/02/93	E05776	Diaminopimelic acid dehydrogenase
E05779	Threonine synthase		E06110	Prephenate dehydratase	Kikuchi，T.et al.“Production of L-phenylalanine by fermentation method，” Patent：JP 1993344881-A 1 12/27/93
E06111	Mutated Prephenate dehydratase	Kikuchi，T.et al.“Production of L-phenylalanine by fermentation method，” Patent：JP 1993344881-A1 12/27/93	E06110	Prephenate dehydratase
E06111	Mutated Prephenate dehydratase		E06146	Acetohydroxy acid synthetase	Inui，M.et al.“Gene capable of coding Acetohydroxy acid synthetase and its use，”Patent：JP 1993344893-A 1 12/27/93
E06825	Aspartokinase	Sugimoto，M.et al.“Mutant aspartokinase gene，”patent：JP 1994062866-A 1 03/08/94	E06146	Acetohydroxy acid synthetase
E06825	Aspartokinase		E06826	Mutated aspartokinase alpha subunit	Sugimoto，M.et al.“Mutant aspartokinase gene，”patent：JP 1994062866-A 1 03/08/94
			E06826	Mutated aspartokinase alpha subunit

Table 2 (continued)
Table 2 (continued)				E06827	Mutated aspartokinase alpha subunit	Sugimoto，M.et al.“Mutant aspartokinase gene，”patent：JP 1994062866-A 1 03/08/94
E07701	secY		Honno，N.et al.“Gene DNA participating in integration of membraneous protein to membrane，”Patent：JP 1994169780-A 1 06/21/94	E06827	Mutated aspartokinase alpha subunit
E07701	secY			E08177	Aspartokinase	Sato，Y.et al.“Genetic DNA capable of coding Aspartokinase released from feedback inhibition and its utilization，”Patent：JP 199426 1766-A 1 09/20/94
E08178， E08179， E08180， E08181， E08182		Feedback inhibition-released Aspartokinase	Sato，Y.et al.“Genetic DNA capable of coding Aspartokinase released from feedback inhibition and its utilization，”Patent：JP 1994261766-A 1 09/20/94	E08177	Aspartokinase
E08178， E08179， E08180， E08181， E08182		Feedback inhibition-released Aspartokinase		E08232	Acetohydroxy-acid isomeroreductase	Inui，M.et al、“Gene DNA coding acetohydroxy acid isomeroreductase，” Patent：JP 1994277067-A 1 10/04/94
E08234	secE		Asai，Y.et al.“Gene DNA coding for translocation machinery of protein，” Patent：JP 1994277073-A 1 10/04/94	E08232	Acetohydroxy-acid isomeroreductase
E08234	secE			E08643	FT aminotransferase and desthiobiotin synthetase promoter region	Hatakeyama，K.et al.“DNA fragment having promoter function in coryneform bacterium，”Patent：JP 1 99503 1 476-A 1 02/03/95
E08646		Biotin synthetase	Hatakeyama，K.et al.“DNA fragment having promoter function in coryneform bacterium，”Patent：JP 1995031476-A 1 02/03/95	E08643
E08646		Biotin synthetase		E08649	Aspartase	Kohama，K.et al“DNA fragment having promoter function in coryneform bacterium，”Patent：JP 1995031478-A 1 02/03/95
E08900		Dihydrodipicolinate reductase	Madori，M.et al.“DNA fragment containing gene coding Dihydrodipicolinate acid reductase and utilization thereof，”Patent：JP 1995075578-A 1 03/20/95	E08649	Aspartase
E08900		Dihydrodipicolinate reductase		E08901	Diaminopimelic acid decarboxylase	Madori，M.et al.“DNA fragment containing gene coding Diaminopimelic acid decarboxylase and utilization thereof，”Patent：JP 1995075579-A1 03/20/95
E12594		Serine hydroxymethyltransferase	Hatakeyama，K.et al.“Production of L-trypophan，”Patent：JP 1997028391-A 1 02/04/97	E08901	Diaminopimelic acid decarboxylase
E12594		Serine hydroxymethyltransferase		E12760， E12759， E12758	transposase	Moriya，M.et al.“Amplification of gene using artificial transposon，”Patent： JP 199707029l-A 03/18/97
E12764		Arginyl-tRNA synthetase；diaminopimelic acid decarboxylase	Moriya，M.et al.“Amplification of gene using artificial transposon，”Patent： JP 199707029l-A 03/18/97	E12760， E12759， E12758	transposase
E12764		Arginyl-tRNA synthetase；diaminopimelic acid decarboxylase		E12767	Dihydrodipicolinic acid synthetase	Moriya，M.et al.“Amplification of gene using artificial transposon，”Patent： JP 199707029l-A 03/18/97
E12770		aspartokinase	Moriya，M.et al.“Amplification of gene using artificial transposon，”Patent： JP 1997070291-A 03/18/97	E12767	Dihydrodipicolinic acid synthetase
E12770		aspartokinase		E12773	Dihydrodipicolinic acid reductase	Moriya，M.et al.“Amplification of gene using artificial transposon，”Patent： JP 1997070291-A 03/18/97

Table 2 (continued)
Table 2 (continued)				E13655		Glucose-6-phosphate dehydrogenase	Hatakeyama，K.et al.“Glucose-6-phosphate dehydrogenase and DNA capable of coding the same，”Patent：JP 199722461-A 1 09/02/97
L01508	IlvA	Threonine dehydratase	Moeckel，B.et al.“Functional and strctural analysis of the threonine dehydratsae of Corynebacterium glutamicum，”J.Bacteriol.，174：8065-8072 (1992)	E13655		Glucose-6-phosphate dehydrogenase
L01508	IlvA	Threonine dehydratase		L07603	EC4.2.1.15	3-deoxy-D-arabinoheptulosonate-7- phosphate synthase	Chen，C.et al.“The cloning and nucleotide sequence of Corynebacterium glutamicum 3-deoxy-D-arabinoheptulosonatee-7-phosphate synthase gene，” FEMS Microbiol.Len.，107：223-230(1993)
L09232	IlvB；ilvN；ilvC	Acetohydroxy acid synthase large subunit； Acetohydroxy acid synthase small subunit； Acetohydroxy acid isomeroreductase	Keilhauer，C.et al.“lsoleucine synthesis in Corynebacterium glutamicum： molecular analysis of the ilvB-ilvN-ilvC operon，”J.Bacteriol.175(17)：5595- 5603(1993)	L07603	EC4.2.1.15	3-deoxy-D-arabinoheptulosonate-7- phosphate synthase
L09232	IlvB；ilvN；ilvC			L18874	PtsM	Phosphoenolpyruvate sugar phosphotransferase	Fouet，A et al.“Bacillus subtilis sucrose-specific enzyme II of the phosphotransferase system：expression in Escherichia coli and homology to enzymes II from enteric bacteria，”PNAS USA，84(24)：8773-8777(1987)；Lee， J.K.et al.“Nucleotide sequence of the gene encoding the Corynebacterium glutamicum mannose enzyme II and analyses of the deduced protein sequence，”FEMS Microbiol.，119(1-2)：137-145(1994)
L27123	aceB	Malate synthase	Lee，H-S.et al.“Molecular characterization of aceB，a gene encoding malate synthase in Corynebacterium glutamicum，”J.Microbiol.Bioiechnol.， 4(4)：256-263(1994)	L18874	PtsM	Phosphoenolpyruvate sugar phosphotransferase
L27123	aceB	Malate synthase		L27126		Pyruvate kinase	Jetten，M.S.et al.“Structural and functional analysis of pyruvate kinase from Corynebacterium glutamicum，”Appl.Environ.Microbiol.，60(7)：2501-2507 (1994)
L28760	aceA	Isocitrate lyase		L27126		Pyruvate kinase
L28760	aceA	Isocitrate lyase		L35906	dtxr	Diphtheria toxin repressor	Oguiza，J.A.et al.“Molecular cloning，DNA sequence analysis，and characterization ofthe Corynebacterium diphtheriae dtxR from Brevibacterium lactofermentum，”J.Bacteriol，177(2)：465-467(1995)
M13774		Prephenate dehydratase	Follettie，M.T.et al.“Molecular cloning and nucleotide sequence of the Corynebacterium glutamicum pheA gene，”J.Bacteriol.，167：695-702(1986)	L35906	dtxr	Diphtheria toxin repressor
M13774		Prephenate dehydratase		M16175	5SrRNA		Park，Y-H.et al.“Phylogenetic analysis of the coryneform bacteria by 56 rRNA sequences，”J.Bacteriol.，169：1801-1806(1987)
M16663	trpE	Anthranilate synthase，5’end	Sano，K.et al.“Structure and function of the trp operon control regions of Brevibacterium lactofermentum，a glutamic-acid-producing bacterium，”Gene， 52：191-200(1987)	M16175	5SrRNA
M16663	trpE	Anthranilate synthase，5’end		M16664	trpA	Tryptophan synthase，3’end	Sano，K.et al.“Structure and function of the trp operon control regions of Brevibacterium lactofermentum，a glutamic-acid-producing bacterium，”Gene 52：191-200(1987)
				M16664	trpA	Tryptophan synthase，3’end

Table 2 (continued)
Table 2 (continued)				M25819		Phosphoenolpyruvate carboxylase	O’Regan，M.et al.“Cloning and nucleotide sequence of the Phosphoenolpyruvate carboxylase-coding gene of Corynebacterium glutamicum ATCC13032，”Gene，77(2)：237-251(1989)
M85106		23S rRN A gene insertion sequence	Roller，C.et al.“Gram-positive bacteria with a high DNA G+C content are characterized by a common insertion within their 23S rRNA genes，”J.Gen. Microbiol.，138：1167-1175(1992)	M25819		Phosphoenolpyruvate carboxylase
M85106		23S rRN A gene insertion sequence		M85107， M85108		23S rRNA gene insertion sequence	Roller，C.et al.“Gram-positive bacteria with a high DNA G+C content are characterized by a common insertion within their 23S rRNA genes，”J.Gen. Microbiol.，138：1167-1175(1992)
M89931	aecD；brnQ；yhbw	Beta C-S lyase；branched-chain amino acid uptake carrier；hypothetical protein yhbw	Rossol，I.et al.“The Corynebacterium glutamicum aecD gene encodes a C-S lyase with alpha，beta-elimination activity that degrades aminoethylcysteine，” J.Bacteriol.，174(9)：2968-2977(1992)；Tauch，A.et al.“Isoleucine uptake in Corynebacterium glutamicum ATCC 13032 is directed by the brnQ gene product，”Arch.Microbiol.，169(4)：303-312(1998)	M85107， M85108		23S rRNA gene insertion sequence
M89931	aecD；brnQ；yhbw			S59299	trp	Leader gene(promoter)	Herry，D.M.et al.“Cloning of the trp gene cluster from a tryptophan- hyperproducing strain of Corynebacterium glutamicum：identification of a mutation in the teader sequence，”Appl.Environ.Microbiol.，59(3)：791-799 (1993)
U11545	trpD	Anthranilate phosphoribosyltransferase	O’Gara，J.P.and Dunican，L.K.(1994)Complete nucleotide sequence of the Corynebacterium glutamicum ATCC 21850 tpD gene.”Thesis，Microbiology Department，University College Galway，Ireland.	S59299	trp	Leader gene(promoter)
U11545	trpD	Anthranilate phosphoribosyltransferase		U13922	cglIM；cglIR；clgIIR	Putative type II 5-cytosoine methyltransferase；putative type II restriction endonuclease；putative type I or type III restriction endonuclease	Schafer，A.et al.“Cloning and characterization of a DNA region encoding a stress-sensitive restriction system from Corynebacterium glutamicum ATCC 13032 and analysis of its role in intergeneric conjugation with Escherichia coli，”J.Bacteriol，176(23)：7309-7319(1994)；Schafer，A.et al.“The Corynebacterium glutamicum cglIM gene encoding a 5-cytosine in an McrBC- deficient Escherichia coli strain，”Gene，203(2)：95-101(1997)
U14965	recA			U13922	cglIM；cglIR；clgIIR
U14965	recA			U31224	ppx		Ankri，S.et al.“Mutations in the Corynebacterium glutamicumproline biosynthetic pathway：A natural bypass of the proA step，”J.Bacteriol.， 178(15)：4412-4419(1996)
U31225	proC	L-proline：NADP+5-oxidoreductase	Ankri，S.et al.“Mutations in the Corynebacterium glutamicumproline biosynthetic pathway：A natural bypass of the proA step，”J.Bacteriol.， 178(15)：4412-4419(1996)	U31224	ppx
U31225	proC	L-proline：NADP+5-oxidoreductase		U31230	obg；proB；unkdh	？；gamma glutamyl kinase；similar to D- isomer specific 2-hydroxyacid dehydrogenases	Ankri，S.et al.“Mutations in the Corynebacterium glutamicumproline biosynthetic pathway：A natural bypass of the proA step，”J.Bacteriol.， 178(15)：4412-4419(1996)
				U31230	obg；proB；unkdh

Table 2 (continued)
Table 2 (continued)				U31281	bioB	B iotin synthase	Serebriiskii，l.G.，“Two new members of the bio B superfamily：Cloning， sequencing and expression of bio B genes of Methylobacillus flagellatum and Corymebacterium glutamicum，”Gene，175：15-22(1996)
U35023	thtR；accBC	Thiosulfate sulfurtransferase；acyl CoA carboxylase	Jager，W.et al.“A Corynebacterium glutamicum gene encoding a two-domain protein simlar to biotin carboxylases and biotin-carboxyl-carrier proteins，” Arch.Microbiol.，166(2)；76-82(1996)	U31281	bioB	B iotin synthase
U35023	thtR；accBC	Thiosulfate sulfurtransferase；acyl CoA carboxylase		U43535	cmr	Multidrug resistance protein	Jager，W.et al.A“Corynebacterium glutamicum gene conferring multidrug resistance in the heterologous host Escherichia coli，”J.Bacteriol.， 179(7)：2449-2451(1997)
U43536	clpB	Heat shock ATP-binding protein		U43535	cmr	Multidrug resistance protein
U43536	clpB	Heat shock ATP-binding protein		U53587	aphA-3	3’5”-aminoglycoside phosphotransferase
U89648		Corynebacterium glutamicum unidentified sequence involved in histidine biosynthesis， partial sequence		U53587	aphA-3	3’5”-aminoglycoside phosphotransferase
U89648				X04960	trpA；trpB；trpC；trpD； trpE；trpG；trpL	Tryptophan operon	Matsui，K.et al.“Complete nuleotide and deduced amino acid sequences of the Brevibacterium lactofermentum tryptophan operon，”Nucleic Acids Res.， 14(24)：10113-10114(1986)
X07563	lys A	DAP decarboxylase(meso-diaminopimelate decarboxylase，EC 4.1.1.20)	Yeh，P.et al.”Nucleic sequence of the lysA gene of Corynebacterium glutamicum，and possible mechanisms for modulation of its expression，”Mol. Gen.Genet.，212(1)：112-119(1988)	X04960	trpA；trpB；trpC；trpD； trpE；trpG；trpL	Tryptophan operon
X07563	lys A			X14234	EC4.1.1.31	Phosphooenolpyruvate carboxylase	Eikmanns，B.J.et al.“The Phosphoenolpyruvate carboxylase gene of Corynebacterium glutamicum：Molecular cloning，nucleotide sequence，and expression，”Mol. Gen.Genet.，218(2)：330-339(1989)；Lepiniec，L.et al. “Sorghum Phosphoenolpyruvate carboxylase gene family：structure，function and molecular evolution，”Plant.Mol.Biol.，21(3)：487-502(1993)
X17313	fda	Fructose-bisphosphate aldolase	Von der Osten，C.H.et al.“Molecular cloning，nucleotide sequence and fine- structural analysis of the Corynebacterium glutamicum fda gene：structural comparison of C.glutamicum fructose-1，6-biphosphate aldolase to class I and class II aldolases，”Mol. Microbiol.，	X14234	EC4.1.1.31	Phosphooenolpyruvate carboxylase
X17313	fda	Fructose-bisphosphate aldolase		X53993	dapA	L-2，3-dihydrodipicolinate synthetase(EC 4.2.1.52)	Bonnassie，S.et al.“Nucleic sequence of the dapA gene from Corynebacterium glutamicum，”Nucleic Acids Res.，18(21)：6421(1990)
X54223		AttB-related site	Cianciotto，N.et al“DNA sequence homology between att B-related sites of Corynebacterium diphtheriae，Corynebacterium ulcerans，Corynebacterium glutamicum，and the attP site of lambdacorynephage，”FEMS.Microbiol， Lett.，66：299-302(1990)	X53993	dapA	L-2，3-dihydrodipicolinate synthetase(EC 4.2.1.52)
X54223		AttB-related site		X54740	argS；lysA	Arginyl-tRNA synthetase；Diam inopimelate decarboxylase	Marcel，T.et al.“Nucleotide sequence and organization of the upstream region of the Corynebactetium glutamicum lysA gene，”Mol.Microbiol.，4(11)：1819- 1830(1990)

Table 2 (continued)
Table 2 (continued)				X55994	trpL；trpE	Putative leader peptide；anthranilate synthase component I	Heery，D.M.et al.“Nucleotide sequence of the Corynebacterium glutamicum trpE gene，”Nucleic Acids Res.，18(23)：7138(1990)
X56037	thrC	Threonine synthase	Han，K.S.et al.“The molecular structure of the Corynebacterium glutamicum threonine synthase gene，”Mol.Microbiol.，4(10)：1693-1702(1990)	X55994	trpL；trpE	Putative leader peptide；anthranilate synthase component I
X56037	thrC	Threonine synthase		X56075	attB-related site	Attachment site	Cianciotto，N.et al.“DNA sequence homology between att B-related sites of Corynebacterium diphtheriae，Corynebacterium ulcerans，Corynebacterium glutamicum，and the attP site of lambdacorynephage，”FEMS.Microbiol. Lett.，66：299-302(1990)
X57226	lysC-alpha；lysC-beta； asd	Aspartokinase-alpha subunit； Aspartokinase-beta subunit；aspartate beta semialdehyde dehydrogenase	Kalinowski，J.et al.“Genetic and biochemical analysis of the Aspartokinase from Corynebacterium glutamicum，”Mol.Microbiol.，5(5)：1197-1204(1991)； Kalinowski，J.et al.“Aspartokinase genes lysC alpha and lysC beta overlap and are adjacent to the asperate beta-semialdehyde dehydrogenase gene asd in Corynebacterium glutamicum，”Mol.Gen.Genet.，224(3)：317-324(1990)	X56075	attB-related site	Attachment site
X57226	lysC-alpha；lysC-beta； asd			X59403	gap；pgk；tpi	Glyceraldehyde-3-phosphate； phosphoglycerate kinase；triosephosphate isomerase	Eikmanns，B.J.“Identitication，sequence analysis，and expression of a Corynebacterium glutamicum gene cluster encoding the three glycolytic enzymes glyceraldehyde-3-phosphate dehydrogenase，3-phosphoglycerate kinase，and triosephosphate isomeras，”J.Bacteriol.，174(19)：6076-6086 (1992)
X59404	gdh	Glutamate dehydrogenase	Bormann，E.R.et al.“Molecular analysis of the Corynebacterium glutamicum gdh gene encoding glutamate dehydrogenase，”Mol.Microbiol.，6(3)：317-326 (1992)	X59403	gap；pgk；tpi
X59404	gdh	Glutamate dehydrogenase		X60312	lys1	L-lysine permase	Seep-Feldhaus，A.H.et al.“Molecular analysis of he Corynebacterium glutamicum lysl gene involved in lysine uptake，”Mol.Microbiol.5(12)：2995- 3005(1991)
X66078	cop1	Psl protein	Joliff，G.et al.“Cloning and nucleotide sequence of the csp1 gene encoding PS1，one ofthe two major secreted proteins of Corynebacterium glutamicum： The deduced N-terminal region of PS1 is similar to the Mycobacterium antigen 85 complex，”Mol.Microbiol.，6(16)：2349-2362(1992)	X60312	lys1	L-lysine permase
X66078	cop1	Psl protein		X66112	glt	Citrate synthase	Eikmanns，B.J.et al.“Cloning sequence，expression and transcriptional analysis of the Corynebacterium glutamicum gltA gene encoding citrate synthase，”Microbiol.，140：1817-1828(1994)
X67737	dapB	Dihydrodipicolinate reductase		X66112	glt	Citrate synthase
X67737	dapB	Dihydrodipicolinate reductase		X69103	csp2	Surface layer protein PS2	Peyret，J.L.et al.“Characterization of the cspB gene encoding PS2，an ordered surface-layer protein in Corynebacterium glutamicum，”Mol.Microbiol.， 9(1)：97-109(1993)
X69104		IS3 related insertion element	Bonamy，C.et al.“Identification of IS1206，a Corynebacterium glutamicum IS3-related insertion sequence and phylogenetic analysis，”Mol.Microbiol.， 14(3)：571-581(1994)	X69103	csp2	Surface layer protein PS2

Table 2 (continued)
Table 2 (continued)				X70959	leuA	Isopropylmalate synthase	Patek，M.et al.“Leucine synthesis in Corynebacterium glutamicum：enzyme activities，structure of leuA，and effect of leuA inactivation on lysine synthesis，”Appl.Environ.Microbiol.，60(1)：133-140(1994)
X71489	icd	Isocitrate dehydrogenase(NADP+)	Eikmanns，B.J.et al.“Cloning sequence analysis，expression，and inactivation of the Corynebacterium glutamicum icd gene encoding isocitrate dehydrogenase and biochemical characterization of the enzyme，”J.Bacteriol.， 177(3)：774-782(1995)	X70959	leuA	Isopropylmalate synthase
X71489	icd	Isocitrate dehydrogenase(NADP+)		X72855	GDHA	Glutamate dehydrogenase(NADP+)
X75083， X70584	mtrA	5-methyltryptophan resistance	Heery，D.M.et al.“A sequence from a tryptophan-hyperproducing strain of Corynebacterium glutamicum encoding resistance to 5-methyltryptophan，” Biochem.Biophys.Res.Commun.，201(3)：1255-1262(1994)	X72855	GDHA	Glutamate dehydrogenase(NADP+)
X75083， X70584	mtrA	5-methyltryptophan resistance		X75085	recA		Fitzpatrick，R.et al.“Construction and characterization of recA mutant strains of Corynebacterium glutamicum and Brevibacterium lactofermentum，”Appl. Microbiol.Biotechnol.，42(4)：575-580(1994)
X75504	aceA；thiX	Partial Isocitrate lyase；？	Reinscheid，D.J.et al.“Characterization of the isocitrate lyase gene from Corynebacterium glutamicum and biochemical analysis of the enzyme，”J. Bacteriol.，176(12)：3474-3483(1994)	X75085	recA
X75504	aceA；thiX	Partial Isocitrate lyase；？		X76875		ATPase beta-subunit	Ludwig，W.et al.“Phylogenetic relationships of bacteria based on comparative sequence analysis of elongation factor Tu and ATP-synthase beta-subunit genes，”Antonie Van Leeuwenhoek，64：285-305(1993)
X77034	tuf	Elongation factor Tu	Ludwig，W.et al.“Phylogenetic relationships of bacteria based on comparative sequence analysis of elongation factor Tu and ATP-synthase beta-subunit genes，”Antonie Van Leeuwenhoek，64：285-305(1993)	X76875		ATPase beta-subunit
X77034	tuf	Elongation factor Tu		X77384	recA		Billman-Jacobe，H.“Nucleotide sequence ofa recA gene from Corynebacterium glutamicum，”DNA Seq.，4(6)：403-404(1994)
X78491	aceB	Malate synthase	Reinscheid，D.J.et al.“Malate synthase from Corynebacterium glutamicum pta-ack operon encoding phosphotransacetylase：sequence analysis，” Microbiology，140：3099-3108(1994)	X77384	recA
X78491	aceB	Malate synthase		X80629	16S rDNA	16S ribosomal RNA	Rainey，F.A.et al.“Phylogenetic analysis of the genera Rhodococcus and Norcardia and evidence for the evolutionary origin of the genus Norcardia from within the radiation of Rhodococcus species，”Microbiol.，141：523-528 (1995)
X81191	gluA；gluB；gluC； gluD	Glutamate uptake system	Kronemeyer，W.et al.“Structure of the gluABCD cluster encoding the glutamate uptake system of Corynebacterium glutamicum，”J.Bacteriol.， 177(5)：1152-1158(1995)	X80629	16S rDNA	16S ribosomal RNA
X81191	gluA；gluB；gluC； gluD	Glutamate uptake system		X81379	dapE	Succinyldiaminopimelate desuccinylase	Wehrmann，A.et al.“Analysis of different DNA fragments of Corynebacterium glutamicum complementing dapE of Escherichia coli，” Microbiology，40：3349-56(1994)

Table 2 (continued)
Table 2 (continued)				X82061	16S rDNA	16S ribosomal RNA	Ruimy，R.et al.“Phylogeny of the genus Corynebacterium deduced from alyses of small-subunit ribosomal DNA sequences，”Int.J.Syst.Bacteriol.， 45(4)：740-746(1995)
X82928	asd；lysC	Aspartate-semialdehyde dehydrogenase；？	Serebrijski，I.et al.“Multicopy suppression by asd gene and osmotic stress- dependent complementation by heterologous proA in proA mutants，”J. Bacteriol.，177(24)：7255-7260(1995)	X82061	16S rDNA	16S ribosomal RNA
X82928	asd；lysC	Aspartate-semialdehyde dehydrogenase；？		X82929	proA	Gamma-glutamyl phosphate reductase	Serebrijski，1.et al.“Multicopy，suppression by asd gene and osmotic stress- dependent complementation by heterologous proA in proA mutants，”J. Bacteriol.，177(24)：7255-7260(1995)
X84257	16S rDNA	16S ribosomal RNA	Pascual，C.et al.”Phylogenetic analysis of the genus Corynebacterium based on 16S rRNA gene sequences，”Int.J.Syst.Bacteriol.，45(4)：724-728(1995)	X82929	proA	Gamma-glutamyl phosphate reductase
X84257	16S rDNA	16S ribosomal RNA		X85965	aroP；dapE	Aromatic amino acid permease；？	Wehrmann et al.“Functional analysis of sequences adjacent to dapE of C. glutamicum proline reveals the presence of aroP，which encodes the aromatic amino acid transporter，”J.Bacteriol.，177(20)：5991-5993(1995)
X86157	argB；argC；argD； argF；argJ	Acetylglutamate kinase；N-acetyl-gamma- glutamyl-phosphate reductase； acetylornithine aminotransferase；omithine carbamoyltransferase；glutamate N- acetyltransfeferase	Sakanyan，V.et al.“Genes and enzymes of the acetyl cycleof arginine biosynthesis in Corynebacterium glutamicum：enzyme evolution in the early steps of the arginine pathway，”Microbiolgy，142：99-108(1996)	X85965	aroP；dapE	Aromatic amino acid permease；？
X86157	argB；argC；argD； argF；argJ			X89084	pta；ackA	Phosphate acetyltransferase；acetate kinase	Reinschheid，D.J.et al.“Cloning，sequence analysis，expression and inactivation of the Corynebacterium glutamicum pta-ack operon encoding phosphotransacetylase and acetate kinase，”Microbiology，145：503-513(1999)
X89850	attB	Attachment site	Le Marrec，C.et al.“Genetic characterization of site-specific integration functions of phi AAU2 infecting”Arthrobacter aureus C70，”J.Bacteriol.， 178(7)：1996-2004(1996)	X89084	pta；ackA	Phosphate acetyltransferase；acetate kinase
X89850	attB	Attachment site		X90356		Promoter fragment F1	Patek，M.et al.“Promoters from Corynebacterium glutanicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)
X90357		Promoter fragment F2	Patek，M.et al.“Promoters from Corynebacterium glutanicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)	X90356		Promoter fragment F1
X90357		Promoter fragment F2		X90358		Promoter fragment F10	Patek，M.et al.“Promoters from Corynebacterium glutamicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)
X90359		Promoter fragment F13	Patek，M.et al.“Promoters from Corynebacterium glutamicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)	X90358		Promoter fragment F10
X90359		Promoter fragment F13

Table 2 (continued)
Table 2 (continued)				X90360		Promoter fragment F22	Patek，M.et al.“Promoters from Corymebacterium glutamicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)
X90361		Promoter fragment F34	Patek，M.et al.“Promoters from Corynebacterium glutamicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)	X90360		Promoter fragment F22
X90361		Promoter fragment F34		X90362		Promoter fragment F37	Patek，M.et al.“Promotets from C.glutamicum：cloning，molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)
X90363		Promoter fragment F45	Patek，M.et al.“Promoters from Corynebacterium glutamicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)	X90362		Promoter fragment F37
X90363		Promoter fragment F45		X90364		Promoter fragment F64	Patek，M.et al.“Promoters from Corynebacterium glutamicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)
X90365		Promoter fragment F75	Patek，M.et al.“Promoters from Corynebacterium glutamicum：cloming， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)	X90364		Promoter fragment F64
X90365		Promoter fragment F75		X90366		Promoter fragment PF101	Patek，M.et al.“Promoters from Corynebacterium glutamicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)
X90367		Promoter fragment PF104	Patek，M.et al.“Promoters from Corynebacterium glutamicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)	X90366		Promoter fragment PF101
X90367		Promoter fragment PF104		X90368		Promoter fragment PF109	Patek，M.et al.“Promoters from Corynebacterium glutamicum：cloning， molecular analysis and search for a consensus motif，”Microbiology， 142：1297-1309(1996)
X93513	amt	Ammonium transport system	Siewe，R.M.et al.“Functional and genetic characterization of the(methyl) ammonium uptake carrier of Corynebacterium glutamicum，”J.Biol.Chem.， 271(10)：5398-5403(1996)	X90368		Promoter fragment PF109
X93513	amt	Ammonium transport system		X93514	betP	Glycine betaine transport system	Peter，H.et al.“Isolation，characterization，and expression of the Corynebacterium glutamicum betP gene，encoding the transport system for the compatible solute glycine betaine，J.Bacteriol.，178(17)：5229-5234(1996)
X95649	orf4		Patek，M.et al.“Identification and transcriptional analysis of the dapB-ORF2- dapA-ORF4 operon of Corynebacterium glutamicum，encoding two enaymes involved in L-lysine synthesis，”Biotechnol.Lett.，19：1113-1117(1997)	X93514	betP	Glycine betaine transport system
X95649	orf4			X96471	lysE；lysG	Lysine exporter protein；Lysine export regulator protein	Vrljic，M.et al.“A new type of transporter with a new type of cellular function：L-lysine export from Corynebacterium glutamicum，”Mol. Microbiol，22(5)：815-826(1996)

Table 2 (continued)
Table 2 (continued)				X96580	panB；panC；xylB	3-methy-2-oxobutanoate hydroxymethyltransferase；pantoate-beta- alanine ligase；xylulokinase	Sahm，H.et al.“D-pantothenate synthesis in Corynebacte rium glutamicum and use of panBC and genes encoding L-valine synthesis for D-pantothenate overproduction，”Appl.Environ.Microbiol.，65(5)：1973-1979(1999)
X96962		Insertion sequence IS 1207 and transposase		X96580	panB；panC；xylB
X96962		Insertion sequence IS 1207 and transposase		X99289		Elongation factor P	Ramos，A.et al.“Cloning，sequencing and expression of gene encoding elongation factor P in the amino-acid producer Brevibacterium lactofermentum (Corynebacrerium glutamicum ATCC 13869)，”Gene，198：217-222(1997)
Y00140	thrB	Homoserine kinase	Mateos，L.M.et al.“Nucleotide sequence of the homoserine kinase(thrB)gene of the Brevibacterium lactofermentum，”Nucleic Acids Res.，15(9)：3922(1987)	X99289		Elongation factor P
Y00140	thrB	Homoserine kinase		Y00151	ddh	Meso-diaminopimelate D-dehydrogenase (EC 1.4.1.16)	Ishino，S.et al.“Nucleotide sequence of the meso-diaminopimelate D- dehydrogenase gene from Corynebacterium glutamicum，”Nucleic Acids Res.， 15(9)：3917(1987)
Y00476	thrA	Homoserine dehydrogenase	Mateos，L.M.et al.“Nucleotide sequence of the homoserine dehydrogenase (thrA)gene of the Brevibacterium lactofermentum，”Nucleic Acids Res.， 15(24)：10598(1987)	Y00151	ddh	Meso-diaminopimelate D-dehydrogenase (EC 1.4.1.16)
Y00476	thrA	Homoserine dehydrogenase		Y00546	hom；thrB	Homoserine dehydrogenase；homoserine kinase	Peoples，O.P.et al.“Nucleotide sequence and fine structural analysis of the Corynebacterium glutamicum hom-thrB operon，”Mol.Microbiol.，2(1)：63-72 (1988)
Y08964	murC；ftsQ/divD；ftsZ	UPD-N-acetylmuramate-alanine ligase； division initiation protein or cell division protein；cell division protein	Honrubia，M.P.et al.“Identification，characterization，and chromosomal organization of the ftsZ gene from Brevibacterium lactofermentum，”Mol.Gen. Genet.，259(1)：97-104(1998)	Y00546	hom；thrB	Homoserine dehydrogenase；homoserine kinase
Y08964	murC；ftsQ/divD；ftsZ			Y09163	putP	High affinity proline transport system	Peter，H.et al.“Isolation of the putP gene of Corynebacterium glutamicumproline and characterization of a low-affinity uptake system for compatible solutes，”Arch.Microbiol.，168(2)：143-151(1997)
Y09548	pyc	Pyruvate carboxylase	Peters-Wendisch，P.G.et al.“Pyruvate carboxylase from Corynebacterium glutamicum：characterization，expression and inactivation of the pyc gene，” Microbiology，144：915-927(1998)	Y09163	putP	High affinity proline transport system
Y09548	pyc	Pyruvate carboxylase		Y09578	leuB	3-isopropylmalate dehydrogenase	Patek，M.et al.“Analysis of the leuB gene from Corynebacterium glutamicum，”Appl.Microbiol.Biotechnol，50(1)：42-47(1998)
Y12472		Attachmentsite bacteriophage Phi-16	Moreau，S.et al.“Site-specific integration of corynephage Phi-16：The construction of an integration vector，”Microbiol.，145：539-548(1999)	Y09578	leuB	3-isopropylmalate dehydrogenase
Y12472		Attachmentsite bacteriophage Phi-16		Y12537	proP	Proline/ectoine uptake system protein	Peter，H.et al.“Corynebacterium glutamicum is equipped with four secondary carriers for compatible solutes：ldentification，sequencing，and characterization of the prolinee/ectoine uptake system，ProP，and the ectoine/proline/glycine betaine carrier，EctP，”J.Bacteriol.，180(22)：6005-6012(1998)

Table 2 (continued)
Table 2 (continued)				Y13221	glnA	Glutamine synthetase I	Jakoby，M.et al.“Isolation of Corynebacte rium glutamicum glnA gene encoding glutamine synthetase I，”FEMS Microbiol.Lett.，154(1)：81-88(1997)
Y16642	lpd	Dihydrolipoamide dehydrogenase		Y13221	glnA	Glutamine synthetase I
Y16642	lpd	Dihydrolipoamide dehydrogenase		Y18059		Attachment site Corynephage 304L	Moreau，S.et al.“Analysis of the integration functions of &phi；304L：An integrase module among corynephages，”Virology，255(1)：150-159(1999)
Z21501	argS；lysA	Arginyl-tRNA synthetase；diaminopimelate decarboxylase(partial)	Oguiza，J.A.et al“A gene encoding arginyl-tRNA synthetase is located in the upstream region of the lysA gene in Brevibacterium lactofermentum： Regulation of argS-lysA cluster expression by arginine，”J. Bacteriol.，175(22)：7356-7362(1993)	Y18059		Attachment site Corynephage 304L
Z21501	argS；lysA			Z21502	dapA；dapB	Dihydrodipicolinate synthase； dihydrodipicolinate reductase	Pisabarro，A.et al.“A cluster of three genes(dapA，orf2，and dapB)of Brevibacterium lactofermentum encodes dihydrodipicolinate reductase，and a third polypeptide of unknown function，”J.Bacteriol.，175(9)：2743-2749 (1993)
Z29563	thrC	Threonine synthase	Malumbres，M.et al.“Analysis and expression of the thrC gene of the encoded threonine synthase，”Appl.Environ.Microbiol.，60(7)2209-2219(1994)	Z21502	dapA；dapB	Dihydrodipicolinate synthase； dihydrodipicolinate reductase
Z29563	thrC	Threonine synthase		Z46753	16S rDNA	Gene for 16S ribosomal RNA
Z49822	sigA	SigA sigma factor	Oguiza，J.A.et al“Multiple sigma factor genes in Brevibacterium lactofermentum：Characterization of sigA and sigB，”J.Bacteriol.，178(2)：550- 553(1996)	Z46753	16S rDNA	Gene for 16S ribosomal RNA
Z49822	sigA	SigA sigma factor		Z49823	galE；dtxR	Catalytic activity UDP-galacrose 4- epimerase；diphtheria toxin regulatory protein	Oguiza，J.A.et al“The galE gene encoding the UDP-galactose 4-epimerase of Brevibacterium lactofermentum is coupled transcriptionally to the dmdR gene，”Gene，177：103-107(1996)
Z49824	orf1；sigB	？；SigB sigma factor	Oguiza，J.A.et al“Multiple sigma factor genes in Brevibacterium lactofermentum：Characterization of sigA and sigB，”J.Bacteriol.，178(2)：550- 553(1996)	Z49823	galE；dtxR
Z49824	orf1；sigB	？；SigB sigma factor		Z66534		Transposase	Correia，A.et al.“Cloning and characterization of an IS-like element presentin the genome of Brevibacterium lactofermentum ATCC 13869，”Gene， 170(1)：91-94(1996)
The gene sequences in the cited references have been disclosed. However, the present inventors disclosed a sequence obtained was compared to the sequence length. Speculate publicly initiation codon sequence error, therefore only Actual fragment of the coding region.				Z66534		Transposase

Table 3: can be used in the practice of the invention, Corynebacterium and Brevibacterium

Generic name	Species	ATCC	FERM	NRRL	CECT	NCIMB	CBS	NCTC	DSMZ
Generic name	Species	ATCC	FERM	NRRL	CECT	NCIMB	CBS	NCTC	DSMZ	Brevibacterium	ammoniagenes	21054
Brevibacterium	ammoniagenes	19350								Brevibacterium	ammoniagenes	21054
Brevibacterium	ammoniagenes	19350								Brevibacterium	ammoniagenes	19351
Brevibacterium	ammoniagencs	19352								Brevibacterium	ammoniagenes	19351
Brevibacterium	ammoniagencs	19352								Brevibacterium	ammoniagenes	19353
Brevibacterium	ammoniagenes	19354								Brevibacterium	ammoniagenes	19353
Brevibacterium	ammoniagenes	19354								Brevibacterium	ammoniagenes	19355
Brevibacterium	ammoniagenes	19356								Brevibacterium	ammoniagenes	19355
Brevibacterium	ammoniagenes	19356								Brevibacterium	ammoniagenes	21055
Brevibacterium	ammoniagenes	21077								Brevibacterium	ammoniagenes	21055
Brevibacterium	ammoniagenes	21077								Brevibacterium	ammoniagenes	21553
Brevibacterium	ammoniagenes	21580								Brevibacterium	ammoniagenes	21553
Brevibacterium	ammoniagenes	21580								Brevibacterium	ammoniagenes	39101
Brevibacterium	butanicum	21196								Brevibacterium	ammoniagenes	39101
Brevibacterium	butanicum	21196								Brevibacterium	divaricatum	21792	P928
Brevibacterium	flayum	21474								Brevibacterium	divaricatum	21792	P928
Brevibacterium	flayum	21474								Brevibacterium	flavum	21129
Brevibacterium	flavum	21518								Brevibacterium	flavum	21129
Brevibacterium	flavum	21518								Brevibacterium	flavum			B11474
Brevibacterium	flavum			B11472						Brevibacterium	flavum			B11474
Brevibacterium	flavum			B11472						Brevibacterium	flavum	21127
Brevibacterium	flavum	21128								Brevibacterium	flavum	21127
Brevibacterium	flavum	21128								Brevibaeterium	flavum	21427
Brevibacterium	flavum	21475								Brevibaeterium	flavum	21427
Brevibacterium	flavum	21475								Brevibacterium	flavum	21517
Brevibacterium	flavum	21528								Brevibacterium	flavum	21517
Brevibacterium	flavum	21528								Brevibacterium	flavum	21529
Brevibacterium	flavum			B11477						Brevibacterium	flavum	21529
Brevibacterium	flavum			B11477						Brevibacterium	flavum			B11478
Brevibacterium	flavum	21127								Brevibacterium	flavum			B11478
Brevibacterium	flavum	21127								Brevibacterium	flavum			B11474
Brevibacterium	healii	15527								Brevibacterium	flavum			B11474
Brevibacterium	healii	15527								Brevibacterium	ketoglutamicum	21004
Brevibacterium	ketoglutamicum	21089								Brevibacterium	ketoglutamicum	21004
Brevibacterium	ketoglutamicum	21089								Brevibacterium	ketosoreductum	21914
Brevibacterium	lactofermentum				70					Brevibacterium	ketosoreductum	21914
Brevibacterium	lactofermentum				70					Brevibacterium	lactofermentum				74
Brevibacterium	lactofermentum				77					Brevibacterium	lactofermentum				74
Brevibacterium	lactofermentum				77					Brevibacterium	lactofermentum	21798
Brevibacterium	lactofermentum	21799								Brevibacterium	lactofermentum	21798
Brevibacterium	lactofermentum	21799								Brevibacterium	lactofermentum	21800
Brevibacterium	lactofermentum	21801								Brevibacterium	lactofermentum	21800
Brevibacterium	lactofermentum	21801								Brevibacterium	lactofermentum			B11470
Brevibacterium	lactofermentum			B11471						Brevibacterium	lactofermentum			B11470

Generic name	Species	ATCC	FERM	NRRL	CECT	NCIMB	CBS	NCTC	DSMZ
Generic name	Species	ATCC	FERM	NRRL	CECT	NCIMB	CBS	NCTC	DSMZ	Brevibacterium	lactofermentum	21086
Brevibacterium	lactofermentum	21420								Brevibacterium	lactofermentum	21086
Brevibacterium	lactofermentum	21420								Brevibacterium	lactofermentum	21086
Brevibacterium	lactofermentum	31269								Brevibacterium	lactofermentum	21086
Brevibacterium	lactofermentum	31269								Brevibacterium	linens	9174
Brevibacterium	linens	19391								Brevibacterium	linens	9174
Brevibacterium	linens	19391								Brevibacterium	linens	8377
Brevibacterium	paraffinolyticum					11160				Brevibacterium	linens	8377
Brevibacterium	paraffinolyticum					11160				Brevibacterium	spec.			717.73
Brevibacterium	spec.						717.73			Brevibacterium	spec.			717.73
Brevibacterium	spec.						717.73			Brevibacterium	spec.	14604
Brevibacterium	spcc.	21860								Brevibacterium	spec.	14604
Brevibacterium	spcc.	21860								Brevibacterium	spec.	21864
Brevibacterium	spec.	21865								Brevibacterium	spec.	21864
Brevibacterium	spec.	21865								Brevibacterium	spec.	21866
Brevibacterium	spec.	19240								Brevibacterium	spec.	21866
Brevibacterium	spec.	19240								Corynebacterium	acetoacidophilum	21476
Corynebactcrium	acetoacidophilum	13870								Corynebacterium	acetoacidophilum	21476
Corynebactcrium	acetoacidophilum	13870								Corynebacterium	acetoglutamicum		B11473
Corynebacterium	acetoglutamicum			B11475						Corynebacterium	acetoglutamicum		B11473
Corynebacterium	acetoglutamicum			B11475						Corynebacterium	acetoglutamicum	15806
Corynebacterium	acetoglutamicum	21491								Corynebacterium	acetoglutamicum	15806
Corynebacterium	acetoglutamicum	21491								Corynebacterium	acetoglutamicum	31270
Corynebacterium	acetophilum			B3671						Corynebacterium	acetoglutamicum	31270
Corynebacterium	acetophilum			B3671						Corynebacterium	ammoniagenes	6872			2399
Corynebacterium	ammoniagenes	15511								Corynebacterium	ammoniagenes	6872			2399
Corynebacterium	ammoniagenes	15511								Corynebacterium	fujiokense	21496
Corynebacterium	glutamicum	14067								Corynebacterium	fujiokense	21496
Corynebacterium	glutamicum	14067								Corynebacterium	glutamicum	39137
Corynebacterium	glutamicum	21254								Corynebacterium	glutamicum	39137
Corynebacterium	glutamicum	21254								Corynebacterium	glutamicum	21255
Corynebacterium	glutamicum	31830								Corynebacterium	glutamicum	21255
Corynebacterium	glutamicum	31830								Corynebacterium	glutamicum	13032
Corynebacterium	glutamicum	14305								Corynebacterium	glutamicum	13032
Corynebacterium	glutamicum	14305								Corynebacterium	glutamicum	15455
Corynebacterium	glutamicum	13058								Corynebacterium	glutamicum	15455
Corynebacterium	glutamicum	13058								Corynebacterium	glutamicum	13059
Corynebacterium	glutamicum	13060								Corynebacterium	glutamicum	13059
Corynebacterium	glutamicum	13060								Corynebacterium	glutamicum	21492
Corynebacterium	glutamicum	21513								Corynebacterium	glutamicum	21492
Corynebacterium	glutamicum	21513								Corynebacterium	glutamicum	21526
Corynebacterium	glutamicum	21543								Corynebacterium	glutamicum	21526
Corynebacterium	glutamicum	21543								Corynebacterium	glutamicum	13287
Corynebacterium	glutamicum	21851								Corynebacterium	glutamicum	13287
Corynebacterium	glutamicum	21851								Corynebacterium	glutamicum	21253
Corynebacterium	glutamicum	21514								Corynebacterium	glutamicum	21253
Corynebacterium	glutamicum	21514								Corynebacterium	glutamicum	21516
Corynebacterium	glutamicum	21299								Corynebacterium	glutamicum	21516

Generic name	Species	ATCC	FERM	NRRL	CECT	NCIMB	CBS	NCTC	DSMZ
Generic name	Species	ATCC	FERM	NRRL	CECT	NCIMB	CBS	NCTC	DSMZ	Corynebacterium	glutamicum	21300
Corynebacterium	glutamicum	39684								Corynebacterium	glutamicum	21300
Corynebacterium	glutamicum	39684								Corynebacterium	glutamicum	21488
Corynebacterium	glutamicum	21649								Corynebacterium	glutamicum	21488
Corynebacterium	glutamicum	21649								Corynebacterium	glutamicum	21650
Corynebacterium	glutamicum	19223								Corynebacterium	glutamicum	21650
Corynebacterium	glutamicum	19223								Corynebacterium	glutamicum	13869
Corynebacterium	glutamicum	21157								Corynebacterium	glutamicum	13869
Corynebacterium	glutamicum	21157								Corynebacterium	glutamicum	21158
Corynebacterium	glutamicum	21159								Corynebacterium	glutamicum	21158
Corynebacterium	glutamicum	21159								Corynebacterium	glutamicum	21355
Corynebacterium	glutamicum	31808								Corynebacterium	glutamicum	21355
Corynebacterium	glutamicum	31808								Corynebacterium	glutamicum	21674
Corynebacterium	glutamicum	21562								Corynebacterium	glutamicum	21674
Corynebacterium	glutamicum	21562								Corynebacterium	glutamicum	21563
Corynebacterium	glutamicum	21564								Corynebacterium	glutamicum	21563
Corynebacterium	glutamicum	21564								Corynebacterium	glutamicum	21565
Corynebacterium	glutamicum	21566								Corynebacterium	glutamicum	21565
Corynebacterium	glutamicum	21566								Corynebacterium	glutamicum	21567
Corynebacterium	glutamicum	21568								Corynebacterium	glutamicum	21567
Corynebacterium	glutamicum	21568								Corynebacterium	glutamicum	21569
Corynebacterium	glutamicum	21570								Corynebacterium	glutamicum	21569
Corynebacterium	glutamicum	21570								Corynebacterium	glutamicum	21571
Corynebacterium	glutamicum	21572								Corynebacterium	glutamicum	21571
Corynebacterium	glutamicum	21572								Corynebacterium	glutamicum	21573
Corynebacterium	glutamicum	21579								Corynebacterium	glutamicum	21573
Corynebacterium	glutamicum	21579								Corynebacterium	glutamicum	19049
Corynebacterium	glutamicum	19050								Corynebacterium	glutamicum	19049
Corynebacterium	glutamicum	19050								Corynebacterium	glutamicum	19051
Corynebacterium	glutamicum	19052								Corynebacterium	glutamicum	19051
Corynebacterium	glutamicum	19052								Corynebacterium	glutamicum	19053
Corynebacterium	glutamicum	19054								Corynebacterium	glutamicum	19053
Corynebacterium	glutamicum	19054								Corynebacterium	glutamicum	19055
Corynebacterium	glutamicum	19056								Corynebacterium	glutamicum	19055
Corynebacterium	glutamicum	19056								Corynebacterium	glutamicum	19057
Corynebacterium	glutamicum	19058								Corynebacterium	glutamicum	19057
Corynebacterium	glutamicum	19058								Corynebacterium	glutamicum	19059
Corynebacterium	glutamicum	19060								Corynebacterium	glutamicum	19059
Corynebacterium	glutamicum	19060								Corynebacterium	glutamicum	19185
Corynebacterium	glutamicum	13286								Corynebacterium	glutamicum	19185
Corynebacterium	glutamicum	13286								Corynebacterium	glutamicum	21515
Corynebacterium	glutamicum	21527								Corynebacterium	glutamicum	21515
Corynebacterium	glutamicum	21527								Corynebacterium	glutamicum	21544
Corynebacterium	glutamicum	21492								Corynebacterium	glutamicum	21544
Corynebacterium	glutamicum	21492								Corynebacterium	glutamicum		B8183
Corynebacterium	glutamicum			B8182						Corynebacterium	glutamicum		B8183
Corynebacterium	glutamicum			B8182						Corynebacterium	glutamicum		B12416
Corynebacterium	glutamicum			B12417						Corynebacterium	glutamicum		B12416

Generic name	Species	ATCC	FERM	NRRL	CECT	NCIMB	CBS	NCTC	DSMZ
Generic name	Species	ATCC	FERM	NRRL	CECT	NCIMB	CBS	NCTC	DSMZ	Corynebacterium	glutamicum			B12418
Corynebacterium	glutamicum			B11476						Corynebacterium	glutamicum			B12418
Corynebacterium	glutamicum			B11476						Corynebacterium	glutamicum	21608
Corynebacterium	lilium		P973							Corynebacterium	glutamicum	21608
Corynebacterium	lilium		P973							Corynebacterium	nitrilophilus	21419			11594
Corynebacterium	spec.		P4445							Corynebacterium	nitrilophilus	21419			11594
Corynebacterium	spec.		P4445							Corynebacterium	spec.		P4446
Corynebacterium	spec.	31088								Corynebacterium	spec.		P4446
Corynebacterium	spec.	31088								Corynebacterium	spec.	31089
Corynebacterium	spec.	31090								Corynebacterium	spec.	31089
Corynebacterium	spec.	31090								Corynebacterium	spec，	31090
Corynebacterium	spec.	31090								Corynebacterium	spec，	31090
Corynebacterium	spec.	31090								Corynebacterium	spec.	15954				20145
Corynebacterium	spec.	21857								Corynebacterium	spec.	15954				20145
Corynebacterium	spec.	21857								Corynebacterium	spec.	21862
Corynebacterium	spec.	21863								Corynebacterium	spec.	21862

ATCC: American Type Culture Collection, Rockville, MD, USA

FERM: Fermentation Research Institute, Chiba, Japan

NRRL: ARS Agricultural Research Institutions Collection, NRRL, Peoria, IL, USA

CECT: Spanish Type Culture Collection, Valencia, Spain

NCIMB: National Collection of Industrial and Marine Microorganisms Limited, Aberdeen, UK

CBS: Fungal Culture Collection, Baarn, NL

NCTC: National Type Culture Collection, London, UK

DSMZ: Deutsche Microorganismes, Braunschweig, Germany

See also Sugawara, H.et al. (1993) World directory of collections of cultures of microorganisms: Bacteria, fungi and yeasts (4^th edn)，World federation for culture collections world data center on microorganisms，Saimata，Japen.

Table 4: Sequence comparison results

ID#	Length (NT)	Hit Results	Length	Accession number	Genbank Hit name	Source Genbank Hit	Homology (GAP)	Warehousing Day
ID#	Length (NT)	Hit Results	Length	Accession number	Genbank Hit name	Source Genbank Hit	Homology (GAP)	Warehousing Day	rxa00023 rxa00044 rxa00064 rxa00072 rxa00105 rxa00106 rxa00115	3579 1059 1401 798 579 1170	GB_EST33：A1776129 GB_EST33：A1776129 EM_PAT：E11760 GB_PAT：126124 GB_BA2：ECOUW89 GB_PAT：E16763 GB_HTG2：AC007892 GB_HTG2：AC007892 GB_BA1：MTV002 GB_BA1：ECU29581 GB_BA2：AE000366 GB_EST15：AA494237 GB_BA2：AF161327 GB_PAT：AR041189 GB_PR4：AC007110 GB_HTG3：AC008537 GB_HTG3：AC008537	483 483 6911 6911 176195 2517 134257 13257 56414 71128 10405 367 2021 654 148336 170030 170030	A1776129 A1776129 E11760 126124 U00006 E16763 AC007892 AC007892 AL008967 U29581 AE000366 AA494237 AF161327 AR041189 AC007110 AC008537 AC008537	EST257217 tomato resistant，Cornell Lycopersicon esculentum cDNA clone cLER17D3，mRNA sequence. EST257217 tomato resistant，Cornell Lycopersicon esculentum cDNA clone cLER17D3.mRNA sequence. Base sequence of sucrase gene. Sequence 4 from patent US 5556776. E.coli chromosomal region from 89.2 to 92.8 minutes. gDNA encoding aspartate transferase(AAT). Drosophlia melanogaster chromosome 3 clone BACR02O03(D797)RPCI-98 02.0.3map 99B-99B strain y；cn bw sp，*SEQUENCING IN PROGRESS ，113 unordered pieces. Drosophila melanogaster chromosome 3 clone BACR02O03(D797)RPC1-98 02.0.3map 99B-99B strain y；cn bw sp，SEQUENCING IN PROGRESS，113 unordered pieces. Mycobacterium tuberculosis H37Rv complete genome；segment 122/162. Escherichia coli K-12 genome；approximately 63 to 64 minutes. Escherichia coli K-12 MG1655 section 256 of 400 of the complete genome- ng83f04.s1 NCI_CGAP_Pr6 Homo sapiens cDNA clone IMAGE：941407 similar to SW：DYR_LACCA P00381 DIHYDROFOLATE REDUCTASE；， mRNA sequence. Corynebacterium diphtheriae histidine kinase ChrS(chrS)and response regulator ChrA(chrA)genes，complete cds. Sequence 4 from patent US 5811286 Homo sapiens chromosome 17，clone hRPK.472_J_18，complete sequence. Homo sapiens chromosome 19 clone CIT-HSPC_490E21.SEQUENCING IN PROGRESS，93 unordered pieces. Homo sapiens chromosome 19 clone CIT-HSPC_490E21.SEQUENCING IN PROGRESS*，93 unordered pieces.	Lycopersicon esculentum Lycopersicon esculentum Corynebacterium glutamicum Unknown. Escherichia coli Corynebacterium glutamicum Drosophila melanogaster Drosophila melanogaster Mycobacterium tuberculosis Escherichia coli Escherichia coli Homo sapiens Corynebacterium diphtheriae Unknown Homo sapiens Homo sapiens Homo sapiens	40,956 40,956 42,979 42,979 39,097 95,429 31,111 31,111 37,753 35,669 35,669 42,896 40,210 41,176 36,783 40,296 40,296	29-Jun-99 29-Jun-99 08-OCT- 1997(Rel. 52，Created) 07-OCT- 1996 17-DEC- 1993 28-Jul-99 2-Aug-99 2-Aug-99 17-Jun-98 14-Jan-97 12-Nov-98 20-Aug-97 9-Sep-99 29-Sep-99 30-MAR- 1999 2-Sep-99 2-Sep-99

Table 4 (continued)

rxa00116 rxa00131 rxa00132 rxa00145 rxa00146 rxa00147 rxa00156

1284 732 1557 1059 1464 1302 1233

GB_BA2：AF062345 16458 AF062345 GB_PAT：118647 3300 118647 GB_GSS13：AQ44619 751 AQ446197 7 GB_BA1：MTY20B11 36330 Z95121 GB_BA1：SAR7932 15176 AJ007932 GB_BA1：MTY20B11 36330 Z95121 GB_BA1：MTY20B11 36330 Z95121 GB_IN2：TVU40872 1882 U40872 GB_HTG6：AC010706 169265 AC010706 GB_BA1：MTCY2B12 20431 Z81011 GB_BA1：PSEPYRBX 2273 L19649 GB_BA1：LLPYRBDNA 1468 X84262 GB_BA1：MTCY2B12 20431 Z81011 GB_BA1：MTCY154 13935 Z98209 GB_BA1：MSGY154 40221 AD000002 GB_BA1：MTCY2B12 20431 Z81011 GB_BA1：MSGB937C 38914 L78820 S GB_BA1：PAU81259 7285 U81259 GB_BA1：SC9B10 33320 AL009204

Caulobacter crescentus Sst1(sst1)，S-layer protein subunit(rsaA)，ABC transporter(rsaD)，membrane forming unit(rsaE)，putative GDP-mannose-4，6- dehydratase(lpsA)，putative acetyltransferase(lpsB)，putative perosamine synthetase(lpsC)，putative mannosyltransferase(lpsD)，putative mannosyltransferase(lpsE)，outer membrane protein(rsaF).and putative perosamine transferase(lpsE)genes，complete cds. Sequence 6 from patent US 5500353. nbxb0062D16r CUGI Rice BAC Library Oryza sativa genomic clone nbxb0062D16r，genomic survey sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 139/162. Streptomyces argillaceus mithramycin biosynthetic genes. Mycobacterium tuberculosis H37Rv complete genome；segment 139/162. Mycobacterium tuberculosis H37Rv complete genome；segment 139/162. Trichomonas vaginalis S-adenosyl-L-homocysteine hydrolase gene，complete cds. Drosophila melanogaster chromosome X clone BACR36D15(D887)RPC1-98 36.D.15 map 13C-13E strain y；cn bw sp， ^***SEQUENCING IN PROGRESS ^***，74 unordered pieces. Mycobacterium tuberculosis H37Rv complete genome；segment 61/162. Pseudomonas aeruginosa aspartate transcarbamoylase(pyrB)and dihydroorotase-like(pyrX)genes，complete cds’s. L.leichmannii pyB gene. Mycobacterium tuberculosis H37Rv complete genome；segment 61/162. Mycobacterium tuberculosis H37Rv complete genome；segment 121/162. Mycobacterium tuberculosis sequence from clone y154. Mycobacterium tuberculosis H37Rv complete genome；segment 61/162. Mycobacterium leprae cosmid B937 DNA sequence. Pseudomonas aeruginosa dihydrodipicolinate reductase(dapB)gene，partial cds.carbamoylphosphate synthetase small subunit(carA)and carbamoylphosphate synthetase large subunit(carB)genes.complete cds. and FtsJ homolog(ftsJ)gene.partia cds. Streptomyces coelicolor cosmid 9B10.

Caulobacter crescentus Unknown. Oryza sativa Mycobacterium tuberculosis Streptomyces argillaceus Mycobacterium tuberculosis Mycobacterium tuberculosis Trichomonas vaginalis Drosophila melanogaster Mycobacterium tuberculosis Pseudomonas aeruginosa Lactobacillus leichmannii Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacltrium tuberculosis Mycobacterium tuberculosis Mycobacterium leprae Pseudomonas aeruginosa Streptomyces coelicotor

36,235 36,821 38,124 43,571 41,116 39,726 36,788 61,914 51,325 63,365 56,080 47,514 60,714 39,229 36,618 61,527 59.538 55,396 52,666

19-OCT- 1999 07-OCT- 1996 8-Apr-99 17-Jun-98 15-Jun-99 17-Jun-98 17-Jun-98 31-OCT- 1996 22-Nov-99 18-Jun-98 26-Jul-93 29-Apr-97 18-Jun-98 17-Jun-98 03-DEC- 1996 18-Jun-98 15-Jun-96 23-DEC- 1996 10-Feb-99

Table 4 (continued)

rxa00166 783 rxa00198 672 rxa00216 1113 rxa00219 1065 rxa00223 1212 rxa00229 803 rxa00241 1626

GB_BA2：AF002133 GB_BA1：D85417 GB_HTG3：AC008167 GB_HTG3：AC008167 GB_HTG4：AC010118 GB_BA1：AB024708 GB_BA1：AB024708 GB_EST24：A1232702 GB_HTG2：HSDJ850E 9 GB_HTG2：HSDJ850E 9 GB_PR2：CNS01DSA GB_HTG2：AC005079 _0 GB_HTG2：AC005079 _1 GB_HTG2：AC005079 _1 GB_BA1：PPEA3NIF GB_BA2：AF128444 GB_HTG4：AC010111 GB_BA2：AF124518 GB_PR3：AC004593 GB_HTG2：AC006907 GB_BA1：CGLYSI

15437 7984 174223 174223 80605 8734 8734 528 117353 117353 159400 110000 110000 110000 19771 2477 138938 1758 150221 188972 4232

AF002133 D85417 AC008167 AC008167 AC010118 AB024708 AB024708 AI232702 AL121758 AL121758 AL121766 AC005079 AC005079 AC005079 X99694 AF128444 AC010111 AF124518 AC004593 AC006907 X60312

Mycobacterium avium strain GIR10 transcriptional regulator(mav81)gene， partial cds，aconitase(acn)，invasin 1(inv1)，invasin 2(inv2)，transcriptional regulator(moxR)，ketoacyl-reductase(fabG)，enoyl-reductase(inhA)and ferrochelatase(mav272)genes，complete cds. Propionibacterium freudenreichii hemY，hemH，hemB，hemX，hemR and heml，genes，complete cds. Homo sapiens clone NH0172013，***SEQUENClNG IN PROGRESS***，7 unordered pieces. Homo sapiens clone NH0172013，***SEQUENCING IN PROGRESS***，7 unordered pieces. Drosophila melanogaster chromosome 3L/62B1 clone RPCI98-10D15，*** SEQUENCING IN PROGRESS***，51 unordered pieces. Corynebacterium glutamicum gltB and gltD genes for glutamine 2- oxocglutarate aminotransferase large and small subunits，complete cds. Corynebacterium glutamicum gltB and gttD genes for glutamine 2- oxoglutarate aminotransferase large and small subunits，complete cds. EST229390 NormaLized rat kidney.Bento Soares Rattus sp.cDNA clone RKICF35 3′end，mRNA sequence. Homo sapiens chromosome 20 clone RP5-850E9，***SEQUENCING IN PROGRESS***，in unordered pieces. Homo sapiens chromosome 20 clone RP5-850E9，***SEQUENCING IN PROGRESS***，in unordered pieces. Human chromosome 14 DNA sequence***IN PROGRESS***BAC R-412H8 of RPCI-11 library from chromosome 14 of Homo sapiens(Human)，complete sequence. Homo sapiens clone RG252P22，***SEQUENCING IN PROGRESS***，3 unordered pieces. Homo sapiens clone RG252P22，***SEQUENCING IN PROGRESS***，3 unordered pieces. Homo sapiens clone RG252P22，***SEQUENCING IN PROGRESS***，3 unordered pieces Plasmid pEA3 nitrogen fixation genes. Rhodobacter capsulatus molybdenum cofactor biosynthetic gene cluster. partial sequence. Drosophila melanogaster chromosome 3L/70C1 clone RPC198-9B18.*** SEQUENCING IN PROGRESS***，64 unordered pieces. Conynebacterium glutamicum 3-dehydroquinase(aroD)and shikimate dehydrogenase(aroE)genes，complete cds. Homo sapiens PAC clone DJ0964C11 from 7p14-p15.complete sequence Caenorhabditis elegans clone Y76B12，***SEQUENCING IN PROGRESS*** 25 unordered pieces. C.glutamicum lysl gene for L-lysine permease.

Mycobacterium avium Propionibacterium freudenreichii Homo sapiens Homo sapiens Drosophila melanogaster Corynebacterium glutamicum Corynebacterium glutamicum Rattus sp. Homo sapiens Homo sapiens Homo sapiens Homo sapiens Homo sapiens Homo sapiens Enterobacter agglomerans Rhodobacter capsulatus Drosophila melanogaster Corynebacterium glutamicum Homo sapiens Caenorhabditis elegans Corynebacterium qlutamicum

54,191 46,667 37,451 37,451 38,627 92,113 93,702 34,221 37,965 37,965 38,796 38,227 38,227 38,227 48,826 40,135 39,527 98,237 36,616 37,095 100,000

26-MAR- 1998 6-Feb-99 21-Aug-99 21-Aug-99 16-OCT- 1999 13-MAR- 1999 13-MAR- 1999 31-Jan-99 03-DEC- 1999 03-DEC- 1999 11-Nov-99 22-Nov-98 22-Nov-98 22-Nov-98 2-Aug-g6 22-MAR- 1999 16-OCT- 1999 18-MAY- 1999 18-Apr-98 26-Feb-99 30-Jan-92

Table 4 (continued)

rxa00262 1197 rxa00266 531 rxa00278 1155 rxa00295 1125 rxa00323 1461 rxa00324 3258 rxa00330 1566 rxa00335 1554

GB_HTG1：PFMAL13P 1 GB_HTG1：PFMAL13P 1 GB_IN2：EHU899655 GB_IN2：EHU89655 GB_RO：AF016190 EM_PAT：E09719 GB_PAT：E02133 GB_IN1：CELK05F6 GB_BA1：CGU43535 GB_RO：RNU30789 GB_BA2：CGU31281 GB_BA1：BRLBIOBA GB_PATE：03937 GB_BA1：MTCY427 GB_BA1：MSGB32CS GB_BA1：MTCY427 GB_BA1：MSGB32CS GB_BA1：MTCY427 GB_OM：BOVELA GB_BA1：CGTHRC GB_PAT：109078 GB_BA1：BLTHRESY N GB_BA1：CGGLNA

192581 192581 3219 3219 2939 3505 3494 36912 2531 3510 1614 1647 1005 38110 364D4 38110 36404 38110 3242 3120 3146 1892 3686

AL049180 AL049180 U89655 U89655 AF016190 E09719 E02133 AF040653 U43535 U30789 U31281 D14084 E03937 Z70692 L78818 Z70692 L78818 Z70692 J02717 X56037 109078 Z29563 Y13221

Plasmodium talciparum chromosome 13 strain 3D7，***SEQUENCING IN PROGRESS***，in unordered pieces Plasmodium falciparum chromosome 13 strain 3D7，***SEQUENCING IN PROGRESS***，in unordered pieces. Entamoeba histolytica unconventional myosin IB mRNA，complete cds. Entamoeba histolytica unconventional myosin IB mRNA，complete cds. Mus musculus connexin-36(Cx36)gene，complete cds. DNA encoding precursor protein of alkaline cellulase. gDNA encoding alkaline cellulase. Caenorhabditis elegans cosmid K05F6. Corynebacterium glutamicum multidrug resistance protein(cmr)gene， complete cds. Rattus norvegicus clone N27 mRNA. Corynebacterium glutamicum biotin synthase(bioB)gene，complete cds. Brevibacterium flavum gene for biotin synthetase，complete cds. DNA sequence encoding Brevibacterium flavum biotin-synthase. Mycobacterium tuberculosis H37Rv complete genome；segment 99/162. Mycobacterium leprae cosmid B32 DNA sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 99/162. Mycobacterium leprae cosmid B32 DNA sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 99/162. Bovine elastin a mRNA，complete cds. Corynebacterium glutamicum thrC gene for threonine synthase(EC 4.2 99.2). Sequence 4 from Patent WO 8809819. Brevibacterium lactofermentum；ATCC 13869：：DNA(genomic)：. Corynebacterium glutamicum glnA gene.

Plasmodium falciparum Plasmodium falciparum Entamoeba histolytica Entamoeba histolytica Mus musculus Bacillus sp. Bacillus sp. Caenorhabditis elegans Corynebacterium glutamicum Rattus norvegicus Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium tuberculoss Bos taurus Corynebacterium glutamicum Unknown. Corynebacterium glutamicum Corynebacterium glutamicum

34,947 34,947 36,496 37,544 41,856 34,741 34,741 36,943 36,658 38,190 99,111 98,489 98,207 35,615 60,917 44,606 52,516 38,079 39,351 99,808 99,617 99,170 100,000

11-Aug-99 11-Aug-99 23-MAY- 1997 23-MAY- 1997 9-Feb-99 08-OCT- 1997(Rel 52.Created 29-Sep-97 6-Jan-98 9-Apr-97 20-Aug-96 21-Nov-96 3-Feb-99 29-Sep-97 24-Jun-99 15-Jun-96 24-Jun-99 15-Jun-96 24-Jun-99 27-Apr-93 17-Jun-97 02-DEC- 1994 20-Sep-95 28-Aug-97

Table 4 (continued)

rxa00347 891 rxa00351 1578 rxa00365 727 rxa00366 480 rxa00367 4653 rxa00371 1917

GB_BA2：AF005635 GB_BA1：MSGB27CS GB_EST27：A1455217 GB_6A2：SSU30252 GB_EST21：AA911262 GB_BA1：MLU15187 GB_IN2：AC004373 GB_IN2：AF145653 GB_BA1：AB024708 GB_BA1：MTCY1A6 GB_BA1：SC3A3 GB_BA1：AB024708 GB_BA1：MTCY1A6 GB_BA1：SC3A3 GB_BA1：AB024708 GB_BA1：MTCY1A6 GB_BA1：SC3A3 GB_V1：SBVORFS GB_EST37：A1967505 GB_IN1：CELK09H9

1690 38793 624 2891 581 36138 72722 3197 8734 37751 15901 8734 37751 15901 8734 37751 15901 7568 380 37881

AF005635 L78817 A1455217 U30252 AA911262 U15187 AC004373 AF145653 AB024708 Z83864 AL109849 AB024708 Z83864 AL109849 AB024708 Z83864 AL109849 M89923 A1967505 AF043700

Corynebacterium glutamicum glutamme synthetase(glnA)gene，complete cds. Mycobacterium leprae cosmid B27 DNA sequence. LD21828.3prime LD Drosophila melanogaster embryo pOT2 Drosophila melanogaster cDNA clone LD21828 3prime.mRNA sequence. Synechococcus PCC7942 nucleoside diphosphate kinase and ORF2 protein genes.complete cds，ORF1protein gene，partial cds，and neutral site I for vector use. oe75a02.s1 NCI_CGAP_Lu5 Homo sapiens cDNA clone IMAGE：1417418 3′ similar to gb：A1B757 UROKINASE PLASMINOGEN ACTIVATOR SURFACE RECEPTOR，GPI-ANCHORED(HUMAN)；，mRNA sequence. Mycobacterium leprae cosmid L296. Drosophila melanogaster DNA sequence(P1 DS05273(D80))，complete sequence. Drosophila melanogaster clone GH08860 BcDNA.GH08860 (BcDNA.GH08860)mRNA，complete cds. Corynebacterium glutamicum gltB and gltD genes for glutamine 2- oxoglutarate aminotransferase large and small subunits，complete cds. Mycobacterium tuberculosis H37Rv complete genome；segment 159/162. Streptomyces coelicolor cosmid 3A3. Corynebacterium glutamicum gltB and gltD genes for glutamine 2- oxoglutarate aminotransferase large and small subunits，complete cds. Mycobacterium tuberculosis H37Rv complete genome；segment 159/162. Streptomyces coelicolor cosmid 3A3. Corynebacterium glutamicum gltB and gltD genes for glutamine 2. oxoglutarate aminotransferase large and small subunits，complete cds. Mycobacterium tuberculosis H37Rv complete geome；segment 159/162. Streptomyces coelicolor cosmid 3A3. Sugarcane bacilliform virus ORF 1，2，and 3 DNA，complete cds. Ljirnpest03-215-c10 Ljirnp Lambda HybriZap two-hybrid library Lotus japonicus cDNA done LP215-03-c10 5’similar to 60S ribosomal protein L39， mRNA sequence. Caenorhabditis elegans cosmid K09H9.

Corynebacterium glutamicum Mycobacterium leprae Drosophila melanogaster Synechococcus PCC7942 Homo sapiens Mycobacterium leprae Drosophila melanogaster Drosophila melanogaster Corynebacterium glutamicum Mycobaclerium tuberculosis Streptomyces coelicolor A3(2) Corynebacterium glutamicum Mycobacterium tuberculosis Streptomyces coelicolor A3(2) Corynebacterium glutamicum Mycobacterium tuberculosis Streplomyces coelicolor A3(2) Sugarcane bacilliform virus Lotus japonicus Caenorhabditis elegans

98,906 66,345 34,510 37,084 37,500 52,972 46,341 49,471 96,556 39,496 37,946 99,374 41,333 37,554 99,312 36,971 37,905 35,843 42,593 34,295

14-Jun-99 15-Jun-96 09-MAR- 1999 29-OCT- 1999 21-Apr-98 09-MAR- 1995 17-Jul-98 14-Jun-99 13-MAR- 1999 17-Jun-98 16-Aug-99 13-MAR- 1999 17-Jun-98 16-Aug-99 13-MAR- 1999 17-Jun-98 16-Aug-99 12-Jun-93 24-Aug-99 22-Jan-98

Table 4 (continued)

rxa00377 rxa00382 rxa00383 rxa00391 rxa00393 rxa00402 rxa00403

1245 1425 1467 843 1017 623 1254

GB_BA1：CCU13664 GB_PL1：ANSDGENE GB_GSS4：AQ730303 GB_BA1：PAHEML GB_BA1：MTY25D10 GB_BA1：MSGY224 GB_BA1：MLCB1222 GB_HTG2：AC006269 GB_HTG2：AC007638 GB_EST38：AW01705 3 GB_PAT：AR065852 GB_VI：AF148805 GB_BA1：MTY25D10 GB_BA1：MSGY224 GB_BA1：MLB1306 GB_BA2：AF052652 GB_BA2：AF109162 GB_BA2：AF092918 GB_BA2：AF052652 GB_BA1：MTV016 GB_EST23：Al111288

678 299 483 4444 40838 40051 34714 167171 178053 613 32207 28559 40838 40051 7762 2096 4514 20758 2096 53662 750

U13664 Y08866 AQ730303 X82072 Z95558 AD000004 AL049491 AC006269 AC007638 AW017053 AR065852 AF148805 Z95558 AD000004 Y13803 AF052652 AF109162 AF092918 AF052652 AL021841 A1111288

Caulobacter crescentus uroporphynogen decarboxylase homolog(hemE) gene，partial cds. A.nidulans sD gene. HS_5505_B1_C04_T7A RPC1-11 Human Male BAC Library Homo sapiens genomic clone Plate＝1081 Col＝7 Row＝F，genomic survey sequence. P.aeruginosa hemL gene. Mycobacterium tuberculosis H37Rv complete genome；segment 28/162. Mycobacterium tuberculosis sequence from clone y224. Mycobacterium leprae cosmid B1222. Homo sapiens chromosome 17 clone hRPK.515_E_23map 17，*** SEQUENCING IN PROGRESS***，2 ordered pieces. Homo sapiens chromosome 17 clone hRPK.515_O_17map 17，*** SEQUENCING IN PROGRESS***，8 unordered pieces EST272398 Schistosoma mansoni male.Phil LoVerde/Joe Merrick Schistosoma mansoni cDNA clone SMMAS14 5′end，mRNA sequence. Sequence 20 from patent US 5849564. Kaposi′s sarcoma-associated herpesvirus ORF 68 gene，parial cds；and ORF 69，kaposin，v-FLIP，v-cyclin，latent nuclear antigen，ORF K14，v-GPCR， putative phosphoribosylformylglycinamidine synthase，and LAMP (LAMP)genes，complete cds. Mycobacterium tuberculosis H37Rv complete genome；segment 28/162. Mycobacterium tuberculosis sequence from clone y224. Mycobacterium leprae cosmid B1306 DNA. Corynebacterium glutamicum homoserine O-acetyltransferase(metA)gene， complete cds. Corynebacterium diphtheriae heme uptake locus，complete sequence. Pseudomonas alcaligenes outer membrane Xcp-secretion system gene cluster. Corynebacterium glutamicum homoserine O-acetyltransferase(metA)gene. complete cds. Mycobacterium tuberculosis H37Rv complete genome；segment 143/162. SWOvAMCAQ02A05SK Onchocerca volvulus adult male cDNA(SAW98MLW OvAM)Onchocerca volvulus cDNA clone SWOvAMCAQ02A05 5′，mRNA sequence.

Caulobacter crescentus Emericella nidulans Homo sapiens Pseudomonas aeruginosa Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium leprae Homo sapiens Homo sapiens Schistosoma mansoni Unknown. Kaposi’s sarcoma- associated herpesvirus Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium leprae Corynebacterium glutamicum Corynebacterium diphtheriae Pseudomonas alcaligenes Corynebacterium glutamicum Mycobacterium tuberculosis Onchocerca volvulus

36,832 39,603 36,728 54,175 61,143 61,143 43,981 35,444 34,821 40,472 38,586 38,509 36,308 39,282 39,228 99,672 40,830 50,161 99,920 52,898 37,565

24-MAR- 1995 17-OCT- 1996 15-Jul-99 18-DEC- 1995 17-Jun-98 03-DEC- 1996 27-Aug-99 10-Jun-99 22-MAY- 1999 10-Sep-99 29-Sep-99 2-Aug-99 17-Jun-98 03-DEC- 1996 24-Jun-97 19-MAR- 1998 8-Jun-99 06-DEC- 1998 19-MAR- 1998 23-Jun-99 31-Aug-98

Table 4 (continued)

rxa00405 rxa00420 rxa00435 rxa00437 rxa00439 rxa00440 rxa00441 rxa00446

613 1587 1296 579 591 582 1287 987

GB_BA1：MTV016 GB_PR4：AC005145 GB_BA1：MTV016 GB_BA1：MTY13D12 GB_BA1：MSGY126 GB_BA1：MSGB971C S GB_BA1：AFACBBTZ GB_HTG4：AC009541 GB_HTG4：AC009541 GB_PR4：AC005951 GB_BA1：SC2A11 G8_PR4：AC005951 GB_BA1：MTV016 GB_PL2：AF167358 GB_HTG3：AC009120 GB_BA2SKZ86111 GB_BA1：SC2E1 GB_BA1：SC2E1 GB_PR2：HS173D1 GB_HTG2：HSDJ719K 3 GB_HTG2：HSDJ719K 3 GB_BA1：SCD78 GB_HTG4：AC009367 GB_HTG4：AC009367

53662 143678 53662 37085 37164 37566 2760 169583 169583 155450 22789 155450 53662 1022 269445 7860 38962 38962 11733B 267114 267114 36224 226055 226055

AL021841 AC005145 AL021841 Z80343 AD000012 L78821 M68904 AC009541 AC009541 AC005951 AL031184 AC005951 AL021841 AF167358 AC009120 Z86111 AL023797 AL023797 AL031984 AL109931 AL109931 AL034355 AC009367 AC009367

Mycobacterium tuberculosis H37Rv complete genome；segment 143/162. Homo sapiens Xp22-166-169 GSHB-523A23(Genome Systems Human BAC library)complete sequernce. Mycobacterium tuberculosis H37Rv complete genome；segment 143/162. Mycobacterium tuberculosis H37Rv complete genome；segment 156/162. Mycobacterium tuberculosis sequence from clone y126. Mycobacterium leprae cosmid B971 DNA sequence. Alcaligenes eutrophus chromsomal transketolase.(ctbTc)and phosphoglycolate phosphatase(cbbZc)genes，complete cds. Homo sapiens chromosome 7，***SEQUENCING IN PROGRESS***，25 unordered pieces. Homo sapiens chromosome 7，***SEQUENCING IN PROGRESS***，25 unordered pieces. Homo sapiens chromosome 17.clone hRPK.372_K_20，complete sequence. Streptomyces coelicolor cosmid 2A11. Homo sapiens chromosome 17.clone hRPK.372_K_20，complete sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 143/162. Rumex acetosa expansin(EXP3)gene，partial cds. Homo sapiens chromosome 16 clone RPC1-11_484E3，***SEQUENCING IN PROGRESS***，34 unordered pieces. Streptomyces lividans rpsP，trmD，rplS，sipW，sipX，sipY，sipZ，mutT genes and 4 open reading frames. Streptomyces coelicolor cosmid 2E1. Streptomyces coelicolor cosmid 2E1. Human DNA sequence from clone 173D1 on chromosome 1p36.21- 36.33.Contains ESTs，STSs and GSSs，complete sequence. Homo sapiens chromosome X clone RP4-719K3 map q21.1-21.31，*** SEQUENCING IN PROGRESS***，in unordered pieces. Homp sapiens chromosome X clone RP4-719K3 map q21.1-21.31，*** SEQUENCING IN PROGRESS***，in unordered pieces. Streptomyces coelicolor cosmid D78. Drosophila melano9aster chromosome 3L/76A2 clone RPC198-48B15，*** SEQUENCING IN PROGRESS***，44 unordered pieces. Drosophila metanogaster chromosome 3L/76A2 clone RPC198-48B15，*** SFOUENGING IN PROGRESS***，44 unordered pieces.

Mycobacterium tuberculosis Homo sapiens Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium leprae Ralstonia eutropha Homo sapiens Homo sapiens Homo sapiens Streptomyces coelicolor Homo sapiens Mycobacterium tuberculosis Rumex acetosa Homo sapiens Strepromyces lividans Streptomyces coelico1or Streptomyces coelicolor Homo sapiens Homo sapiens Homo sapiens Streptomyces coelicolor Drosophila melanogaster Drosophila melanogaster

57,259 34,179 40,169 62,031 61,902 39,651 38,677 36,335 36,335 31,738 43,262 37,647 37,088 46,538 43,276 43,080 42,931 36,702 38,027 34,521 34,521 56,410 34,959 34,959

23-Jun-99 08-DEC- 1998 23-Jun-99 17-Jun-98 10-DEC- 1996 15-Jun-96 27-Jul-94 12-OCT- 1999 12-OCT- 1999 18-Nov-98 5-Aug-98 18-Nov-98 23-Jun-99 17-Aug-99 3-Aug-99 27-OCT- 1999 4-Jun-98 4-Jun-98 23-Nov-99 03-DEC- 1999 03-DEC- 1999 26-Nov-98 16-OCT- 1999 16-OCT- 1999

Table 4 (continued)

rxa00448 rxa00450 rxa00461 rxa00465 rxa00487 rxa00488 rxa00489 rxa00533

1143 424 975 1692 1641 1245 1155

GB_PR3：AC003670 GB_HTG2：AF029367 GB_HTG2：AF029367 GB_HTG2：AC007824 GB_HTG2：AC007824 GB_EST35：A1818057 GB_BA1：MLCB1779 GB_IN1：DMC86E4 GB_GSS15：AQ64032 5 GB_BA1：BAGUAA GB_BA2：U00015 GB_BA1：MTCY78 GB_BA1：MTCY78 GB_BA2：U00015 GB_BA1：SCAJ10601 GB_BA2：U00015 GB_HTG2：HS225E12 GB_HTG2：HS225E12 GB_BA1：CGLYS

88945 148676 148676 133361 133361 412 43254 29352 467 3866 42325 33818 33818 42325 4692 42325 126464 125464 2803

AC003670 AF029367 AF029367 AC007824 AC007824 A1818057 Z98271 AL021086 AQ640325 Y10499 U00015 Z77165 Z77165 U00015 AJ010601 U00015 AL031772 AL031772 X57226

Homo sapiens 12q13.1 PAC RPC11-130F5(Roswell Park Cancer Institute Human PAC library)complete sequence. Homo sapiens chromosome 12 clone RPC1-1 130F5 map 12q13.1，*** SEQUENCING IN PROGRESS***，156 unordered pieces. Homo sapiens chromosome 12 clone RPC1-1 130F5 map 12q13.1，*** SEQUENCING IN PROGRESS***，156 unordered pieces. Drosophila melanogaster chromosome 3 clone BACR02L16(D715)RPC1-98 02.L.16 map 89E-90A strainy；cn bw sp，***SEQUENCING IN PROGRESS ***，91 unordered pieces. Drosophila melanogaster chromosome 3 clone BACR02L16(D715)RPC1-98 02.L.16 map 89E-90A strainy；cn bw sp，***SEQUENCING IN PROGRESS ***，91 unordered pieces. wk14a08.x1 NCI_CGAP_Lym12 Homo sapiens cDNA clone IMAGE：2412278 3′similar to gb：Y00764 UBIQUINOL-CYTOCHROME C REDUCTASE 11 KD PROTEIN(HUMAN)：.mRNA sequeence. Mycobacterium leprae cosmid B1779. Drosophila melanogaster cosmid clone 86E4. 927P1-2H3.TP 927P1 Trypanosoma brucei genomic clone 927P1-2H3. genomic survey sequence. B.ammoniagenes guaA gene. Mycobacterium leprae cosmid B1620. Mycobacterium tuberculosis H37Rv complete genome；segment 145/162. Mycobacterium tuberculosis H37Rv complete genome；segment 145/162. Mycobacterium leprae cosmid B1620. Streptomyces coelicolor A3(2)DNA for whiD and whiK loci. Mycobacterium leprae cosmid B1620. Homo sapiens chromosome 6 clone RP1-225E12 map q24，*** SEQUENCING IN PROGRESS***，in unordered pieces. Homo sapiens chromosome 6 clone RP1-225E12 map q24，*** SEQUENCING IN PROGRESS***，in unordered pieces C.glutamicum lysC-alpha，lysC-beta and asd genes for aspartokinase-alpha and-beta subunits，and aspartate beta semialdehyde dehydrogenase， respectively(EC 2.7.2.4；EC 1.2.1.11).

Homo sapiens Homo sapiens Homo sapiens Drosophila melanogaster Drosophila melanogaster Homo sapiens Mycobacterium leprae Drosophila melanogaster Trypanosoma brucei Corynebacterium ammoniagenes Mycobacterium leprae Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium leprae Streptomyces coelicolor Mycobacterium leprae Homo sapiens Homo sapiens Corynebacterium glutamicum

35,682 31,373 31,373 40,000 40,000 35,714 39,308 37,487 38,116 74,259 37,248 39,725 39,451 39,178 60,835 38,041 36,756 36,756 99,913

9-Jun-98 18-OCT- 1997 18-OCT- 1997 2-Aug-99 2-Aug-99 24-Aug-99 8-Aug-97 27-Apr-99 8-Jul-99 8-Jan-98 01-MAR- 1994 17-Jun-98 17-Jun-98 01-MAR- 1994 17-Sep-98 01-MAR- 1994 03-DEC- 1999 03-DEC- 1999 17-Feb-97

Table 4 (continued)

rxa00534 1386 rxa00536 1494 rxa00537 2409 rxa00541 792 rxa00558 1470 rxa00579 1983 rxa00580 1425

GB_BA1：CGCYSCA5 D GB_PAT：A07546 GB_BA1：CGLYS GB_BA1：CORASKD GB_PAT：E14514 GB_BA1：CGLEUA GB_BA1：MTV025 GB_BA1：MTU88526 GB_BA2：SCD25 GB_BA1：MTCY7H7A GB_BA1：MTU34956 GB_PAT：192052 GB_BA1：MLCB5 GB_BA1：MTCY369 GB_BA1：BAPURF GB_BA1：MLU15182 GB_BA1：MTCY7H7A GB_PAT：AR016483 EM_PAT：E11273 GB_PAT：E12594 GB_PAT：E12594

1591 2112 2803 2957 1643 3492 121125 2412 41622 10451 2462 2115 38109 36850 1885 40123 10451 2104 2104 2104 2104

X82928 A07546 X57226 L16848 E14514 X70959 AL022121 U88526 AL118514 Z95618 U34956 192052 Z95151 Z80226 X91252 U15182 Z95618 AR016483 E11273 E12594 E12594

C.glutamicum aspartate-semialdehyde dehydrogenasegene. Recombinant DNA fragment(Pstl-Xhol). C.glutamicum lysC-alpha，lysC-beta and asd genes for aspartokinase-alpha and-beta subunits，and aspartate beta semialdehyde dehydrogenase， respectively(EC 2.7.2.4；EC 1.2.1.11). Corynebacterium flavum aspartokinase(ask)，and aspartate-semialdehyde dehydrogenase(asd)genes，complete cds. DNA encoding Brevibacterium aspartokinase. C.glutamicum gene leuA for isopropylmalate synthase. Mycobacterium tuberculosis H37Rv complete genome；segment 155/162. Mycobacterium tuberculosis putative alpha-isopropyl malate synthase(leuA) gene，complete cds. Streptomyces coelicolor cosmid D25. Mycobacterium tuberculosis H37Rv complete genome；segment 39/162. Mycobacterium tuberculosis phosphoribosylformylglycinamidine synthase (purL)gene，complete cds. Sequence 19 from patent US 5726299. Mycobacterium leprae cosmid B5. Mycobacterium tuberculosis H37Rv complete genome；segment 36/162. B.ammoniagenes purF gene. Mycobacterium leprae cosmid B2266. Mycobacterium tuberculosis H37Rv complete genome；segment 39/162. Sequence 1 from patent US 5776740. DNA encoding serine hydroxymethyl transferase. DNA encoding serine hydroxymethyltransferase from Brevibacterium flavum. DNA encoding serine hydroxymethyltransferase from Brevibacterium flavum.

Corynebactenum glutamicum synthetic construct Corynebacterium glutamicum Corynebacterium flavescens Corynebacterium glutamicum Corynebacterium glutamicum Mycobacterium tuberculosis Mycobacterium tuberculosis Streptomyces coelicolor A3(2) Mycobacterium tuberculosis Mycobacterium tuberculosis Unknown. Mycobacterium leprae Mycobacterium tuberculosis Corynebacterium ammoniagenes Mycobacterium leprae Mycobacterium tuberculosis Unknown. Corynebacterium glutamicum Corynebacterium glutamicum Corynebaclerium glutamicum

99,221 99,391 99,856 98,701 98,773 100,000 68,003 68,185 63,187 62,401 62,205 98,359 62,468 60,814 66,095 64,315 64,863 98,810 98,810 98,810 99,368

17-Feb-97 30-Jul-93 17-Feb-97 11-Jun-93 28-Jul-99 10-Feb-99 24-Jun-99 26-Feb-97 21-Sep-99 17-Jun-98 28-Jan-97 01-DEC- 1998 24-Jun-97 17-Jun-98 5-Jun-97 09-MAR- 1995 17-Jun-98 05-DEC- 1998 08-OCT- 1997(Rel. 52，Created) 24-Jun-98 24-Jun-98

rxa00581 rxa00584 rxa00618 rxa00619 rxa00620 rxa00624 rxa00626

1092 1248 1230 1551 1014 810 1386

GB_PAT：AR016483 EM_PAT：E11273 GB_PAT：E12594 EM_PAT：E11273 GB_PAT：AR016483 GB_BA1：CORAHPS GB_BA1：AOPCZA361 GB_BA1：D90714 GB_EST19：AA802737 GB_EST28：A1534381 GB_IN1：DMANILLIN GB_BA1：MTCY369 GB_BA1：MLC85 GB_PAT：A60305 GB_PL2：AF063247 GB_BA1：STMAPP GB_HTG3：AC008763 GB_IN1：CEY41E3 GB_EST13：AA362167 GB_N1：CEY41E3 GB_BA1：MTCY369 GB_BA1：MLCB5 GB_BA1：MLU15187

2104 2104 2104 2104 2104 2570 37941 14358 280 581 4029 36850 38109 1845 1450 2069 214575 150641 372 150641 36850 38109 36138

AR016483 E11273 E12594 E11273 AR016483 L07603 AJ223998 D90714 AA802737 A1534381 X89858 Z80226 Z95151 A60305 AF063247 M91546 AC008763 Z95559 AA362167 Z95559 Z80226 Z95151 U15187

Sequence 1 from patent US 5776740. DNA encoding serine hydroxymethyl transferase. DNA encoding serine hydroxymethyltransferase from Brevibacterium flavum. DNA encoding serine hydroxymethyltransferase. Sequence 1 from patent US 5776740. Corynebacterium gtutamicum 3-deoxy-D-arabinoheptulosonate-7-phosphate synthase gene，complete cds. Amycolatopsis orientalis cosmid PCZA361. Escherichia coli genomic DNA.(16.8-17.1min). GM06236.5prime GM Drosophila melanogaster ovary BlueScript Drosophila melanogaster cDNA clone GM06236 5prime，mRNA sequence. SD07186.5prime SD Drosophila melanogaster Schneider L2 cell culture pOT2 Drosophila melanogaster cDNA clone SD07186 5prime similar to X89858：Ani FBgn0011558 PID：g927407 SPTREMBL：Q24240，mRNA sequence. D.melanogaster mRNA for anillin protein. Mycobacterium tuberculosis H37Rv complete genome；segment 36/162. Mycobacte rium leprae cosmid B5. Sequence 5 from Patent WO9708323. Pneumocystis carinii f.sp.ratti enolase mRNA，complete cds. Streptomyces lividans aminopeptidase P(PepP)gene.complete cds. Homo sapiens chromosome 19 clone CITB-E1_3214H19，***SEQUENCING IN PROGRESS***，21 unordered pieces. Caenorhabditis elegans cosmid Y41E3，complete sequence. EST71561 Macrophage I Homo sapiens cDNA 5′end，mRNA sequence. Caenorhabditis elegans cosmid Y41E3，complete sequence Mycobacterium tuberculosis H37Rv complete genome；segment 36/162. Mycobacterium leprae cosmid B5. Mycobacterium leprae cosmid L296.

Unknown. Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Unknown. Corynebacterium glutamicum Amycolatopsis orientalis Escherichia coli Drosophila melanogaster Drosophila melanogaster Drosophila melanogaster Mycobacterium tuberculosis Mycobaclerium leprae unidentified Pneumocystis carinii f.sp. ratti Streptomyces lividans Homo sapiens Caenorhabditis elegans Homo sapiens Caenorhabditis elegans Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium leprae

99,368 99,368 37,071 37,071 37,071 98,236 54,553 53,312 39,928 41,136 34,398 62,776 61,831 61,785 41,060 37,126 40,020 36,986 38,378 37,694 57,971 58,806 38,007

05-DEC- 1998 08-OCT- 1997(Rel. 52，Created 24-Jun-98 08-OCT- 1997(Rel. 52，Created 05-DEC- 1998 26-Apr-93 29-MAR- 1999 7-Feb-99 25-Nov-98 18-MAR- 1999 8-Nov-95 17-Jun-98 24-Jun-97 06-MAR- 1998 5-Jan-99 12-Jun-93 3-Aug-99 2-Sep-99 21-Apr-97 2-Sep-99 17-Jun-98 24-Jun-97 09-MAR- 1995

Table 4 (continued)

rxa00632 rxa00633 rxa00688 rxa00708 rxa00717 rxa00718 rxa00727

795 1392 666 930 1083 831 1035

GB_BA1：BRLBIOAD GB_PAT：E04041 GB_PAT：E04040 GB_BA1：BRLBIOAD GB_PAT：E04040 GB_BA2：EHU38519 GB_BA1：MTV041 GB_BA1：BRLSECY GB_BA2：MBU77912 GB_BA2：AF157493 GB_PAT：100836 GB_PAT：E00311 GB_PAT：178753 GB_PAT：192042 GB_BA1：MTC1125 GB_BA1：MTC1125 GB_BA1：MTC1125 GB_GSS12：AQ42075 5 GB_HTG3：AC008332 GB_HTG3：AC008332

2272 675 1272 2272 1272 1290 28826 1516 7163 25454 1853 1853 1187 1187 37432 37432 37432 671 118545 118545

D14083 E04041 E04040 D14083 E04040 U38519 AL021958 D14162 U77912 AF157493 100836 E00311 178753 192042 Z98268 Z98268 Z98268 AQ420755 AC008332 AC008332

Brevibacterium flavum genes for 7，8-diaminopelargonic acid aminotransterase and dethiobiotin synthetase，complete cds. DNA sequence coding for desthiobiotinsynthetase. DNA sequence coding for diamino pelargonic acid aminotransferase. Brevibacterium flavum genes for 7，8-diaminopelargonic acid aminotransferase and dethiobiotin synthetase，complere cds. DNA sequence coding for diamino pelargonic acid aminotransferase. Erwinia herbicola adenosylmethionine-8-amino-7-oxononanoate transaminase (bioA)gene，complete cds. Mycobacterium tuberculosis H37Rv complete genome；segment 35/162. Brevibacterium flavum gene for SecY protein(complete cds)and gene or adenylate kinase(partial cds). Mycobacterium bovis MBE50a gene，partial cds；and MBE50b，MBE50c， preprotein translocase SecY subunit(secY)，adenylate kinase(adk). methionine aminopeptidase(map)，RNA polymerase ECF sigma factor (sigE50)，MBE50d，and MBE50e genes，complete cds. Zymomonas mobilis ZM4 fosmid clone 42D7，complete sequence. Sequence 1 from Patent US 4758514. DNA coding of 2，5-diketogluconic acid reductase. Sequence 9 from patent US 5693781. Sequence 9 from patent US 5726299. Mvcobacterium tuberculosis H37Rv complete genome；segment 76/162. Mycobacterium tuberculosis H37Rv complete genome；segment 76/162. Mvcobacterium tuberculosis H37Rv complete genome；segment 76/162. RPC1-11-168G18.TJ RPC1-11 Homo sapiens genomic cloneRPC1-11- 168G18，genomic survey sequence. Drosophila melanogaster chromosome 2 clone BACR48D10(D867)RPC1-98 48.D.10 map 34A-34A strain y；cn bw sp，***SEQUENCING IN PROGRESS ***.78 unordered pieces. Drosophila melanogaster chromosome 2 clone BACR48D10(D867)RPC1-98 48 D 10 map 34A-34A strain y；cn bw sp，***SEQUENCING IN PROGRESS***，78 unordered pieces.

Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Erwinia herbicola Mycobacterium tuberculosis Corynebacterium glutamicum Mycobacterium bovis Zymomonas mobilis Unknown. unidentified Unknown. Unknown Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium tuberculosis Homo sapiens Drosophila melanogaster Drosophila melanogaster

97,358 98,074 93,814 95,690 95,755 55,564 60,030 99,563 60,030 39,116 47,419 47,419 37,814 37,814 50,647 55,228 40,300 35,750 40,634 40,634

3-Feb-99 29-Sep-97 29-Sep-97 3-Feb-99 29-Sep-97 4-Nov-96 17-Jun-98 3-Feb-99 27-Jan-99 5-Jul-99 21-MAY- 1993 29-Sep-97 3-Apr-98 01-DEC- 1998 17-Jun-98 17-Jun-98 17-Jun-98 23-MAR- 1999 6-Aug-99 6-Aug-99

Table 4 (continued)

rxa00766 rxa00770 rxa00779 rxa00780 rxa00838 rxa00863 rxa00864

966 1293 1056 669 1023 867 873

GB_HTG3：AC008332 GB_HTG2：AG006789 GB_HTG2：AC006789 GB_BA1：D90810 GB_BA1：MTv043 GB_BA1：MLU15182 GB_BA2：SCD25 GB_HTG1：CER08A5 GB_HTG1：CER08A5 GB_PL2：AF078693 GB_BA1：MTCY98 GB_BA1：AVINIFREG GB_BA2：AF001780 GB_EST1：Z30506 GB_PL2：AC006258 GB_EST37：A1998439 GB_BA1：BLDAPAB GB_PAT：E16749 GB_PAT：E14520 GB_BA1：BLDAPAB GB_BA1：CGDAPB

118545 83823 83823 20476 68848 40123 41622 51920 51920 1492 31225 7099 6701 329 110469 455 3572 2001 2001 3572 1902

AC008332 AC006789 AC006789 D90810 AL022004 U15182 AL118514 Z82281 Z82281 AF078693 Z83860 M60090 AF001780 Z30506 AC006258 A1998439 Z21502 E16749 E14520 Z21502 X67737

Drosop hila melanogaster chromosome 2 clone BACR48D10(D867)RPC1-98 48.D 10 map 34A-34A strain y；cn bw sp，***SEQUENCING IN PROGRESS***，78 unordered pieces. Caenorhabditis elegans clone Y49F6，***SEQUENCING IN PROGRESS***， 2 unordered pieces. Caenorhabditis elegans clone Y49F6，***SEQUENCING IN PROGRESS***， 2 unordered pieoes. E.coli genomic DNA，Kohara clone #319(37.4-37.8min.). Mycobacterium tuberculosis H37Rv complete genome；segment 40/162. Mycobacterium leprae cosmid B2266. Streptomyces coelicolor cosmid D25. Caenorhabditis elegans chromosome V clone R08A5，***SEQUENCING IN PROGRESS***，in unordered pieces. Caenorhabditis elegans chromosome V clone R08A5，***SEQUENCING IN PROGRESS***，in unordered pieces. Chlamydomonas reinhardtii putative O-acetylserine(thiol)lyase precursor (Crcys-1A)mRNA，nuclear gene encoding organellar protein.complete cds. Mycobacterium tuberculosis H37Rv complete genome；segment 103/162. Azotobacter chroococcum nifU，nifS，nifV，nifP，nifW，nifZ and nifM genes， complete cds. Cyanothece PCC 8801 NifP(nifP)，nitrogenase(nifB)，FdxN(fdxN)，NifS(nifS and NifU(nifU)genes，complete cds，and NifH(nifH)gene，partial cds. ATTS2430 AC16H Arabidopsis thaliana cDNA clone TA1306 3′，mRNA sequence. Arabidopsis thaliana BAC F18G18 from chromosome V near 60.5 cM， complete sequence. 701545695 A.thaliana，Columbia Col-0，rosette-2 Arabidopsis thaliana cDNA clone 701545695，mRNA sequence. B.lactofermentum dapA and dapB genes for dihydrodipicolinate synthase and dihydrodipicolinate reductase. gDNA encoding dihydrodipicolinate synthase(DDPS). DNA encoding Brevibacterium dihydrodipicolinic acid synthase. B.lactofemnentum dapAand dapB genes for dihydrodipicolinate synthase and dihydrodipicolinate reductase. C.glutamicum dapB gene for dihydrodipicolinate reductase.

Drosopmila melanogaster Caenorhabditis elegans Caenorhabditis elegans Escherichia coli Mycobacterium tuberculosis Mycobacterium leprae Streptomyces coelicolor A3(2) Caenorhabditis elegans Caenorhabditis elegans Chlamydomonas reinhardti Mycobacterium tuberculosis Azotobacter chroococcum Cyanothece PCC8801 Arabidopsis thaliana Arabidopsis thaliana Arabidopsis thaliana Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum

33,888 36,737 36,737 36,526 66,193 61,443 59,938 64,896 64,896 57,970 54,410 51,729 36,309 44,308 35,571 36,044 99,539 99,539 99,539 99,885 100,000

6-Aug-99 25-Feb-99 25-Feb-99 29-MAY- 1997 24-Jun-99 09-MAR- 1995 21-Sep-99 14-OCT- 1998 14-OCT- 1998 3-Nov-99 17-Jun-98 26-Apr-93 08-MAR- 1999 11-MAR- 1994 2B-DEC- 1998 8-Sep-99 16-Aug-93 28-Jul-99 28-Jul-99 16-Aug-93 1-Apr-93

Table 4 (continued)

rxa00865 rxa00867 rxa00873 rxa00884 rxa00891 rxa00952 rxa00954 rxa00955

1026 650 779 1263 1102 963 644 1545

GB_PAT：E14520 GB_BA1：BLDAPAB GB_PAT：E16752 GB_PAT：AR038113 GB_BA1：MTV002 GB_BA1：MLCB22 GB_BA1：SAU19858 GB_BA1：SCO001206 GB_BA1：SCO001205 GB_BA1：D78198 GB_BA1：MTGY253 GB_BA1：MSGY222 GB_GSS15：AQ65460 0 GB_BA1：MTC14188 GB_BA1：SCO001206 GB_BA1：SCO001205 EM_PAT：E10963 GB_BA1：BLTRP GB_PAT：E01688 GB_PAT：E01375 GB_PAT：E01688 GB_BA1：BLTRP GB_PAT：E01375

2001 3572 1411 1411 56414 40281 2838 9184 9589 2304 41230 41156 468 11700 9184 9589 3118 7725 7725 7726 7725 7725 7726

E14520 Z21502 E16752 AR038113 AL008967 Z98741 U19858 AJ001206 AJ001205 D78198 Z81368 AD000010 AQ654600 Z96071 AJ001206 AJ001205 E10963 X04960 E01688 E01375 E01688 X04960 E01375

DNA encoding Brevibacterium dihydrodipicolinic acid synthase. B.lactofermentum dapA and dapB genes for dihydrodipicolinate synthase and dihydrodipicolinate reductase. gDNA encoding dihydrodipicolinate reductase(DDPR). Sequence 18 from patent US 5804414. Mycobacterium tuberculosis H37Rv complete genome；segment 122/162. Mycobacterium leprae cosmid B22. Streptomyces antibioticus guanosine pentaphosphate synthetase(gpsl)gene. complete cds. Streptomyces coelicolor A3(2)，glycogen metabolism cluster II. Streptomyces coelicolor A3(2)glycogen metabolism clusterl. Pimelobacter sp.DNA for trehalose synthase，complete cds. Mycobacterium tuberculosis H37Rv complete genome；segment 106/162. Mycobacterium tuberculosis sequence from clone y222. Sheared DNA-1014.TF Sheared DNA Trypanosoma brucei genomic clone Sheared DNA-1014.genomc survey sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 7/162. Streptomyces coelicolor A3(2)，glycogen metabolism cluster II. Streptomyces coelicolor A3(2)glycogen metabolism clusterl. gDNA encoding tryptophan synthase. Brevibacterium lactofermentum tryptophan operon. Genomic DNA oftrp operon of prepibacterium latophelmentamn. DNA sequence of tryptophan operon. Genomic DNA of trp operon of prepibacterium latophelmenlamn. Brevibacterium lactofermentum tryptophan operon. DNA sequence of tryptophan operon.

Corynebacterium glutamicum Corynebaderium glutamicum Corynebacterium glutamicun Unknown. Mycobacterium tuberculosis Mycobacterium leprae Streptomyces antibioticus Streptomyces coelicolor Streptomyces coelicolor Pimelobacter sp. Mycobacterium tuberculosis Mycobacterium tuberculosis Trypanosoma brucei Mycobacterium tuberculosis Streptomyoes coelicolor Streptomyces coelicolor Corynebacterium glutamicum Corynebacterium glutamicum unidentified Corynebacterium glutamicum unidentified Corynebacterium glutamicum Corynebacterium glutamicum

100,000 100,000 99,805 99,805 39,179 39,482 69,706 63,415 61,617 60,594 37,785 38,006 33,974 63,297 61,965 61,727 99,688 98,847 98,428 98,758 98,758 98,758 98,372

28-Jul-99 16-Aug-93 28-Jul-99 29-Sep-99 17-Jun-98 22-Aug-97 25-OCT- 1996 29-MAR- 1999 29-MAR- 1999 5-Feb-99 17-Jun-98 03-DEC- 1996 22-Jun-99 18-Jun-98 29-MAR- 1999 29-MAR- 1999 08-OCT- 1997(Rel. 52，Created) 10-Feb-99 29-Sep-97 29-Sep-97 29-Sep-97 10-Feb-99 29-Sep-97

Table 4 (continued)

rxa00956 rxa00957 rxa00958 rxa00970 rxa00972 rxa00981 rxa00989

1237 1677 747 1050 1458 753 1644

GB_BA1：BLTRP GB_PAT：E01688 EM_PAT：E10963 GB_BA1：BLTRP GB_PAT：E01375 GB_BA1：BLTRP GB_PAT：ED1375 GB_PAT：E01688 GB_BA1：BLTRP GB_PAT：E01375 GB_PAT：E01688 GB_BA1：CGHOMTHR GB_PAT：109077 GB_PAT：E01358 GB_PAT：E16755 GB_PAT：AR038110 GB_PAT：E14508 GB_OV：GGA245664 GB_PL2：AC007887 GB_GSS1：CNS00RN W GB_BA1：MTV008 GB_BA1：SCVALSFP GB_BA1：MTV008

7725 7725 3118 7725 7726 7725 7726 7725 7725 7726 7725 3685 3685 2615 3579 3579 3579 512 159434 542 63033 3619 63033

X04960 E01688 E10963 X04960 E01375 X04960 E01375 E01688 X04960 E01375 E01688 Y00546 109077 E01358 E16755 AR038110 E14508 AJ245664 AC007887 AL087338 AL021246 Y13070 AL021246

Brevibacterium lactotermentum tryptophan operon. Genomic DNA of trp operon of prepibaclerium latophelmentamn. gDNA encoding tryptophan synthase. Brevibacterium lactofermentum tryptophan operon. DNA sequence of tryptophan operon. Brevibacterium lactofermentum tryptophan operon. DNA sequence of tryptophan operon. Genomic DNA of trp operon of prepibacterium latophelmentamn. Brevibacterium lactofermentum tryptophan operon. DNA sequence of tryptophan operon. Genomic DNA of trp operon of prepibacterium latophelmentamn. Corynebacterium glutamicum hom-thrB genes for homoserine dehydrogenase and homoserine kinase. Sequence 1 from Patent WO 8809819. DNA encoding for homoserine dehydrogenase(HDH)and homoserine kinase(HK). gDNA encoding diaminopimelate decarboxylase(DDC)and arginyl-tRNA synthase. Sequence 15 from patent US 5804414. DNA encoding Brevibacterium diaminopimelic acid decarboxylase and arginyl- tRNA synthase. Gallus gallus partial mRNA for ATP-citrate lyase(ACL gene). Genomic sequence for Arabidopsis thaliana BAC F 1504 from chromosome I， complete sequence. Arabidopsis thaliana genome survey sequence T7 end of BAC F14D7 of IGF library from strain Columbia of Arabidopsis thaliana，genomic survey sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 108/162. S.coelicolor valS，fpgs，ndk genes. Mycobacterium luberculosis H37Rv complete genome：segment 108/162.

Corynebacterium glutamicum unidentifid Corynebacterium glutamicum Corynebacterium glulamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum unidentified Corynebacterium glutamicum Corynebacterium glutamicum unidentified Corynebacterium glutamicum Unknown. Corynebacterium glutamicum Corynebacterium glutamicum Unknown. Corynebacterium glutamicum Gallus gallus Arabidopsis thaliana Arabidopsis thaliana Mycobacterium tuberculosis Streptomyces coelicolor Mycobacterium tuberculosis

98,372 98,242 98,949 99,107 98,945 99,165 98,927 98,867 98,792 98,792 98,658 99,905 99,810 97,524 99,931 99,931 99,931 37,538 37,600 41,264 40,773 58,119 38,167

10-Feb-99 29-Sep-97 08-OCT- 1997(Rel. 52，Created) 10-Feb-99 29-Sep-97 10-Feb-99 29-Sep-97 29-Sep-97 10-Feb-99 29-Sep-97 29-Sep-97 12-Sep-93 02-DEC- 1994 29-Sep-97 28-Jul-99 29-Sep-99 28-Jul-99 28-Sep-99 04-OCT- 1999 28-Jun-99 17-Jun-98 03-MAR- 1998 17-Jun-98

Table 4 (continued)

rxa00997 rxa01019 rxa01026 rxa01D27 rxa01073 rxa01079 rxa01080 rxa01087

705 1110 1782 1131 954 2226 567 999

GB_BA2：CGU31225 1817 U31225 GB_HTG1：CEY39C12 282838 AL009026 GB_IN1：CEB0001 39416 Z6g634 GB_HTG2：AC005052 144734 AC005052 GB_HTG2：AC005052 144734 AC005052 GB_GSS9：AQ171808 512 AQ171808 GB_BA1：SC1C2 42210 AL031124 GB_BA1：ATLEUCD 2982 X84647 GB_BA1：MTV012 70287 AL021287 GB_BA1：MLC8637 44882 Z99263 GB_BA1：MTCY349 43523 Z83018 GB_BA1：SPUNGMUT 1172 Z21702 X GB_BA1：BACOUTB 1004 M15811 GB_PR4：AC007938 167237 AC007938 GB_PL2：ATAC006282 92577 AC006282 GB_BA2：AF112535 4363 AF112535 GB_BA1：CANRDFGE 6054 Y09572 N GB_BA1：MTV012 70287 AL021287 GB_BA2：AF112535 4363 AF112535 GB_BA1：CANRDFGE 6054 Y09572 N GB_BA1：STNRD 4894 X73226 GB_IN2：AF063412 1093 AF063412 GB_PR3：HS24M15 134539 Z94055 GB_IN2：ARU85702 1240 U85702

Corynebacterium glutamicum L-proline；NADP+5-oxidoreductase(proC)gene， complete cds. Caenorhabditis elegans chromosome IV clone Y39C12，***SEQUENCING IN PROGRESS***，in unordered pieces. Caenorhabditis elegans cosmid B0001，complete sequence. Homo sapiens clone RG038K21，***SEQUENCING IN PROGRESS***，3 unordered pieces. Homo sapiens clone RG038K21，***SEQUENCING IN PROGRESS***，3 unordered pieces. HS_3179_A1_G03_T7 CIT Approved Human Genomic Sperm Library D Homo sapiens genomic clone Plate＝3179 Col＝5 Row＝M，genomic survey sequence. Streptomyces coelicolor cosmid 1C2. A.teichomyceticus leuC and leuD genes. Mycobacterium tuberculosis H37Rv complete genome；segment 132/162. Mycobacterium leprae cosmid8637. Mycobacterium tuberculosis H37Rv complete genome；segment 131/162. S.pneumoniae ung gene and mutX genes encoding uracil-DNA glycosylase and 8-oxodGTP nucleoside triphosphatase. Bacillus subtilis outB gene encoding a sporulation protein，complete cds. Homo sapiens ctone UWGC：djs201 from 7q31，complete sequenoe. Arabidopsis thaliana chromosome II BAC F13K3 genomic sequence. complete sequence. Corycebacterium glutamicum putative glutaredoxin NrdH(nrdH)，NrdI(nrdI)， and ribonucleotide reductase alpha-chain(nrdE)genes，complete cds. Corynebacterium ammoniagenes nrdH，nrdI，nrdE，nrdF genes. Mycobacterium tuberculosis H37Rv complete genome；segment 132/162. Corynebacterium glutamicum putative glutaredoxin NrdH(nrdH)，NrdI(nrdI). and ribonucleotide reductase alpha-chain(nrdE)genes，complete cds. Corynebacterium ammoniagenes nrdH，nrdI，nrdE，nrdF genes. S.typhimurium nrdEF operon. Limnadia lenticularis elongation factor 1-alpha mRNA，partial cds. Human DNA sequence from PAC 24M15 on chromosome 1.Contains tenascin-R(restrictin)，EST. Anathix ralla elongation factor-1 alpha(EF-1a)gene，partial cds.

Corynebacterium glutamicum Caenorhabditis elegans Caenorhabditis elegans Homo sapiens Homo sapiens Homo sapiens Streptomyces coelicolor Actinoplanes teichomyceticus Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium tuberculosis Streptococcus pneumoniae Bacillus subtilis Homo sapiens Arabidopsis thaliana Corynebacterium glutamicum Corynebacterium ammoniagenes Mycobacterium tuberculosis Corynebacterium glutamicum Corynebacterium ammoniagenes Salmonella typhimurium Limnadia lenticularis Homo sapiens Anathix ralla

40,841 36,416 36,416 39,172 39,172 34,661 68,275 65,935 40,454 38,636 51,989 38,088 53,723 34,322 36,181 99,820 75,966 38,296 100,000 65,511 52,477 43,750 37,475 37,319

2-Aug-96 26-OCT- 1999 2-Sep-99 12-Jun-98 12-Jun-9B 17-OCT- 1998 15-Jan-99 04-OCT- 1995 23-Jun-99 17-Sep-97 17-Jun-98 15-Jun-94 26-Apr-93 1-Jul-99 13-MAR- 1999 5-Aug-99 18-Apr-98 23-Jun-99 5-Aug-99 18-Apr-98 03-MAR- 1997 29-MAR- 1999 23-Nov-99 16-Jul-97

Table 4 (continued)

rxa01095 rxa01097 rxa01098 rxa01100 rxa01101 rxa01104 rxa01105 rxa01106

857 477 897 861 756 729 1221 1449

GB_BA1：MTCY01B2 35938 Z95554 GB_HTG5：AC011632 175917 AC011632 GB_HTG5：AC011632 175917 AC011632 GB_BA2：AF030405 774 AF030405 GB_BA2：AF030405 774 AF030405 GB_BA2：AF030405 774 AF030405 GB_BA1：MSGY223 42061 AD000019 GB_BA1：MLCB1610 40055 AL049913 GB_BA2：AF051846 738 AF051846 GB_BA2：AF060558 636 AF060558 GB_HTG1：HSDJ140A 221755 AL109917 9 GB_BA2：AF060558 636 AF060558 GB_BA1：SC4G6 36917 AL096884 GB_BA1：STMHISOPA 3981 M31628 GB_BA1：STMHISOPA 3981 M31628 GB_BA1：SC4G6 36917 AL096884 GB_BA1：MTCY336 32437 Z95586 GB_BA1：MTCY336 32437 Z95586 GB_BA1：MSGY223 42061 AD000019 GB_BA1：MLCB1610 40055 AL049913 GB_BA1：MSGY223 42061 AD000019

Mycobacterium tuberculosis H37Rv complete genome；segment 72/162. Homo sapiens clone RP11-3N13，WORKING DRAFT SEQUENCE，9 unordered pieces. Homo sapiens clone RP11-3N13，WORKING DRAFT SEQUENCE，9 unordered pieces. Corynebacterium glutamicum cyclase(hisF)gene，complete cds. Corynebacterium glutamicum cyclase(hisF)gene，complete cds. Corynebacterium glutamicum cyclase(hisF)gene，complete cds. Mycobacterium tuberculosis sequence from clone y223. Mycobacterium leprae cosmid B1610. Corynebacterium glutamicum phosphoribosylformimino-5-amino-1- phosphoribosyl-4-imidazolecarboxamide isomerase(hisA)gene， complete cds. Corynebacterium glutamicum glutamine amidotransferase(hisH)gene， complete cds. Homo sapiens chromosome 1 clone RP1-140A9，***SEQUENCING IN PROGRESS***，in unordered pieces. Corynebacterium glutamicum glutamine amidotransferase(hisH)gene， complete cds. Streptomyces coelicolor cosmid 4G6. S.coelicolor histidine biosynthesis operon encoding hisD，partial cds.，and hisC，hisB，hisH，and hisA genes，complete cds. S.coelicolor histidine biosynthesis operon encoding hisD，partial cds.，and hisC，hisB，hisH，and hisA genes，complete cds. Streptomyces coelicolor cosmid 4G6. Mycobacterium tuberculosis H37Rv complete genome；segment 70/162. Mycobacterium tuberculosis H37Rv complete genome；segment 70/162. Mycobacterium tuberculosis sequence from clone y223. Mycobacterium leprae cosmid B1610. Mycobacterium tuberculosis sequence from clone y223.

Mycobacterium tuberculosis Homo sapiens Homo sapiens Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Mycobacterium tuberculosis Mycobacterium leprae Corynebacterium glutamicum Corynebacterium glutamicum Homo sapiens Corynebacterium glutamicum Streptomyces coelicolor A3(2) Streptomyces coelicolor Streptomyces coelicolor Streptomyces coelicolor A3(2) Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium tuberculosis

43,243 36,471 36,836 100,000 41,206 97,933 40,972 61,366 97,154 95,455 30,523 94,462 38,378 60,053 58,333 39,045 60,364 60,931 36,851 60,902 37,233

17-Jun-98 19-Nov-99 19-Nov-99 13-Nov-97 13-Nov-97 13-Nov-97 10-DEC- 1996 27-Aug-99 12-MAR- 1998 29-Apr-98 23-Nov-99 29-Apr-98 23-Jul-99 26-Apr-93 26-Apr-93 23-Jul-99 24-Jun-99 24-Jun-99 10-DEC- 1996 27-Aug-99 10-DEC- 1996

Table 4 (continued)

rxa01145 rxa01162 rxa01208 rxa01209 rxa01215 rxa01239 rxa01253

1137 1449 846 1528 1098 2556 873

GB_BA1：MSHISCD GB_BA1：MTCY336 GB_BA1：CORAIA GB_BA1：BRLILVCA GB_PAT：E08232 GB_PAT：A60299 GB_PR3：HS24E5 GB_PR3：AC005265 GB_HTG2：AC004965 GB_HTG2：AC004965 GB_PL2：TAU55859 GB_HTG3：AC011469 GB_HTG3：AC011469 GB_PL1：AB010077 GB_BA1：MTCY10G2 GB_IN1：LEIPRPP GB_HTG2：HSJ799D1 6 GB_BA1：MTCY48 GB_PR2：AB029032 GB_GSS9：AQ107201 GB_PL2：F508 GB_PL2：F508 GB_IN1：CELC06G1

2298 32437 4705 1364 1017 2869 35506 43900 323792 323792 2397 113436 113436 77380 38970 1887 130149 35377 6377 355 99923 99923 31205

X65542 Z95586 L09232 D14551 E08232 A60299 Z82185 AC005265 AC004965 AC004965 U55859 AC011469 AC011469 AB010077 Z92539 M76553 AL050344 Z74020 AB029032 AQ107201 AC005990 AC005990 U41014

M.smegmalis genes hisD and hisC for histidinol dehydrogenase and histidinol- phosphate aminotransferase.respectively. Mycobacterium tuberculosis H37Rv complete genome；segment 70/162. Corynebacterium glutamicum acetohydroxy acid synthase(ilvB)and(ilvN) genes，and acetohydroxy acid isomeroreductase(ilvC)gene，complete cds. Brevibacterium flavum ilvC gene for acetohydroxy acid isomeroreductase. complete cds. DNA encoding aceohydroxy-acid isomeroreductase. Secquence 18 from Patent WO9706261. Human DNA sequence from Fosmid 24E5 on chromosome 22q11.2-qter contains parvalbumin，ESTs，STS. Homo sapiens chromosome 19，cosmid F19750，complete sequence. Homo sapiens clone DJ1106H14，***SEQUENCING IN PROGRESS***，42 unordered pieces. Homo sapiens clone DJ1106H14，***SEQUENCING IN PROGRESS***，42 unordered pieces. Triticum aeslivum heat shock protein 80 mRNA，complete cds. Homo sapiens chromosome 19 clone CIT-HSPC_475D23，***SEQUENCING IN PROGRESS***，31 unordered pieces. Homo sapiens chromosome 19 clone CIT-HSPC_475D23，***SEQUENCING IN PROGRESS***，31 unordered pieces. Arabidopsis thaliana genomic DNA，chromosome 5，P1 clone：MYH19， complete sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 47/162. Leishmania donovani phosphoribosylpyrophosphate synthetase gene， complete cds. Homo sapiens chromosome 1 clone RP4-799D16 map p34.3-36.1，*** SEQUENCING IN PROGRESS***，in unordered pieces. Mycobacterium tuberculosis H37Rv complete genome；segment 69/162. Homo sapiens mRNA for KIAA1109 protein，patial cds. HS_3098_A1_C03_T7 CIT Approved Human Genomic Sperm Library D Homo sapiens genomic clone Plate＝3098 Col＝5 Row＝E，genomic survey sequence. Arabidopsis thaliana chromosome 1 BAC F508 sequence，complete sequence. Arabidopsis thailana chromosome 1 BAC F508 sequence，complete sequence. Caenorhabditis elegans cosmid C06G1.

Mycobacterium smegmatis Mycobacterium tuberculosis Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Aspergillus niger Homo sapiens Homo sapiens Homo sapiens Homo sapiens Triticum aestivum Homo sepiens Homo sapiens Arabidopsis thaliana Mycobacterium tuberculosis Leishmania donovani Homo sapiens Mycobacterium tuberculosis Homo sapiens Homo sapiens Arabidopsis thaliana Arabidopsis thaliana Caenorhabditis elegans

60,111 58,420 100,000 99,560 99,803 38,675 36,204 38,363 36,058 36,058 37,269 40,000 40,000 36,803 37,047 50,738 38,135 38,139 39,394 41,408 36,118 35,574 38,560

30-Jun-93 24-Jun-99 23-Feb-95 3-Feb-99 29-Sep-97 06-MAR- 1998 23-Nov-99 6-Jul-98 12-Jun-98 12-Jun-98 1-Feb-99 07-OCT- 1999 07-OCT- 1999 20-Nov-99 17-Jun-98 7-Jun-93 29-Nov-99 17-Jun-98 4-Aug-99 28-Aug-98 23-DEC- 1998 23-DEC- 1998 30-Nov-95

Table 4 (continued)

rxa01321 rxa01352 rxa01360 rxa01361 rxa01381 rxa01393 rxa01394

1044 706 259 629 944 993 822

GB_GSS14：AQ51884 441 AQ518843 3 GB_HTG2：AC007473 194859 AC007473 GB_HTG4：AC011696 115847 AC011696 G3_PL2：ATAC005167 83260 AC005167 GB_PL2：ATAC005825 97380 AC005825 GB_HTG3：AC011150 127222 AC011150 GB_EST32：A1725583 728 A1725583 GB_PR2：HS227P17 82951 Z81007 GB_EST34：AV171099 173 AV171099 GB_RO：AB008915S1 530 AB008915 GB_EST22：A1050532 293 A1050532 GB_RO：AB008895 3062 AB008895 GB_PL1：AB005237 87835 AB005237 GB_GSS5：AQ766840 491 AQ766840 GB_BA1：MTV043 68848 AL022004 GB_BA1：CGLYSEG 2374 X96471 GB_BA1：SC5A7 40337 AL031107 GB_PR3：AC004054 112184 AC004054 GB_BA1：CGLYSEG 2374 X96471 GB_GSS5：AQ769223 500 AQ769223

HS_5106_A1_D10_SP6E RPCI-11 Human Male BAC Library Homo sapiens genonic clone Plate＝682 Col＝19 Row＝G，genomic survey sequence. Drosophila melanogaster chromosome 2 clone BACR38D12(D590)RPC1-98 38.D.12 map 48A-48B strain y；cn bw sp，***SEQUENCING IN PROGRESS ***，60 unordered pieces. Drosophila melanogaster chromosome 2 clone BACR35F01(D1156)RPC1-98 35.F.1 map 48A-48C strain y；cn bw sp，***SEQUENCING IN PROGRESS ***，108 unordered pieces. Arabidopsis thaliana chromosome 11 BAC F12A24 genomic sequence. complete sequence. Arabidopsis thaliana chromosome 11 BAC T24121 genomic sequence. complete sequence. Homo sapiens clone 4_K_17，LOW-PASS SEQUENCE SAMPLING. BNLGHi12371 Six-day Cotton fiber Gossypium hirsutum cDNA 5′similar to (U86081)root hair defective 3[Arabidopsis thaliana].mRNA sequence. Human DNA sequence from PAC 227P17，between markers DXS6791 andDXS8038 on chromosome X contains CpG island，EST. AV171099 Mus musculus head C57BL/6J 14，17 day embryo Mus musculus cDNA clone 3200002M11，mRNA sequence. Mus musculus mGpi1 gene，exon 1. uc83d10.y1 Sugano mouse kidney mkia Mus musculus cDNA clone IMAGE：1432243 5′similar to TR：O35120 O35120 MGPI1P.；，mRNA sequence. Mus musculus mRNA for mGpi1p.complete cds. Arabidopsis thaliana genoic DNA，chromosome 5，P1 clone：MJJ3，complete sequence. HS_026_A2_C09_T7C ClT Approved Human Genomic Sperm Library D Homo sapiens genomic clone Plate＝2026 Col＝18 Row＝E，genomic survey sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 40/162. C.glutamicum lysE and lysG genes Streptomyces coelicolor cosmid 5A7. Homo sapiens chromosome 4 clone B220GB map 4q21，complete sequence. C.glutamicum lysE and lysG genes. HS_3155_B2_G10_T7C CIT Approved Human Genomic Sperm Library D Homo sapiens genomic clone Plate＝3155 Col＝20 Row＝N，genomic survey sequence.

Homo sapiens Drosophila melanogaster Drosophila melanogaster Arabidopsis thaliana Arabidopsis thaliana Homo sapiens Gossypium hirsutum Homo sapiens Mus musculus Mus musculus Mus musculus Mus musculus Arabidopsis thaliana Homo sapiens Mycobacterium tuberculosis Corynebacterium glutamicum Streptcmyces coelicolor Homo sapiens Corynebacterium glutamicum Homo sapiens

41,121 40,634 38,290 34,311 34,311 37,722 38,492 39,738 46,237 45,574 44,097 41,316 36,606 37,916 37,419 34,831 35,138 37,277 100,000 38,400

05-MAY- 1999 2-Aug-99 26-OCT- 1999 15-OCT- 1998 12-Apr-99 01-OCT- 1999 11-Jun-99 23-Nov-99 6-Jul-99 28-Sep-99 9-Jul-98 23-Nov-97 20-Nov-99 28-Jul-99 24-Jun-99 24-Feb-97 27-Jul-98 9-Jul-98 24-Feb-97 28-Jul-99

Table 4 (continued)

rxa01416 rxa01442 rxa01446 rxa01483 rxa01486 rxa01489 rxa01491 rxa01508 rxa01512

630 1347 1413 1395 757 1146 774 1662 723

GB_BA1：CGLYSEG 2374 X96471 GB_BA1：SC3C3 31382 AL031231 GB_BA1：MLCB22 40281 Z98741 GB_BA1：MTV002 56414 AL008967 GB_BA1：D90827 18886 D90827 GB_BA1：D90828 14590 D90828 GB_BA2：AE000279 10855 AE000279 GB_BA1：SCH10 39524 AL049754 GB_BA1：MTY13E10 35019 Z95324 GB_BA1：MLCB4 36310 AL023514 GB_BA1：MTCY98 31225 Z83860 GB_BA1：MSGB1229C 30670 L78812 S GB_BA2：AF027507 5168 AF027507 GB_BA1：MTV002 56414 AL008967 GB_BA1：MLCB22 40281 Z98741 GB_BA1：SC3C3 31382 AL031231 GB_BA1：CORFADS 1547 D37967 GB_BA1：MLCB22 40281 Z98741 GB_BA1：SC1QA7 39739 AL078618 GB_BA1：MTV002 56414 AL008967 GB_EST13：AA356956 255 AA356956 GB_OV：OMDNAPROI 7327 X92380 GB_IN1：CEF28C12 14653 Z93380 GB_IN1：CEF28C12 14653 Z93380 GB_BA1：SCE9 37730 AL04984 GB_BA1：MAU88875 840 U88875

C.glutamicum lysE and lysG genes. Streptomyces coelicolor cosmid 3C3. Mycobacterium leprae cosmid B22. Mycobacterium tuberculosis H37Rv complete genome；segment 122/162. E.coli genomic DNA，Kohara clone #336(41.2-41.6min.). E.coli genomic DNA，Kohara clone #336gap(41.6-41.9min.). Escherichia coli K-12 MG1655 section 169 of 400 of the complete genome. Streptomyces coelicolor cosmid H10. Mycobacterium tuberculosis H37Rv complete genome；segment 18/162. Mycobacterium leprae cosmid B4. Mycobacterium tuberculosis H37Rv complete genome；segment 103/162. Mycobacterium leprae cosmid B1229 DNA sequence. Mycobacterium smegmatis dGTPase(dgt)，and primase(dnaG)genes. complete cds；tRNA-Asn gene，cornplete sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 122/162. Mycobacterium leprae cosmid B22. Streptomyces coelicolor cosmid 3C3. Corynebaclerium ammoniagenes gene for FAD synthetase.complete cds. Mycobacterium leprae cosmid B22. Streptomyces coelicolor cosmid 10A7. Mycobacterium tuberculosis H37Rv complete genome；segment 122/162. EST65614 Jurkat T-cells III Homo sapiens cDNA 5′end，mRNA sequence. O.mossambicus prolactin I gene. Caenorhabditis elegans cosmid F28C12.complete sequence. Caenorhabditis elegans cosmid F28C12.complete sequence. Streptomyces coelicolor cosmid E9. Mycobacterium avium hyposanthine-guanine phosphoribosyl transferase gene，complete cds.

Corynebacterium glutamicum Streptomyces coelicolor Mycobacterium leprae Mycobacterium tuberculosis Escherichia coli Escherichia coli Escherichia coli Streptomyces coelicolor Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium smegmatis Mycobacterium tuberculosis Mycobacterium leprae Streptomyces coelicolor Corynebacterium ammoniagenes Mycobacterium leprae Streptomyces coelicolor Mycobacterium tuberculosis Homo sapiens Tilapia mossambica Caenorhabditis elegans Caenorhabditis elegans Streptomyces coelicolor Mycobacterium avium

33,665 62,726 39,159 37,340 58,517 56,151 56,021 39,037 40,130 37,752 39,057 54,382 52,941 40,941 38,451 61,194 58,021 38,414 36,930 37,062 37,647 38,289 37,984 38,469 39,021 57,521

24-Feb-97 10-Aug-98 22-Aug-97 17-Jun-98 21-MAR- 1997 21-MAR- 1997 12-Nov-98 04-MAY- 1999 17-Jun-98 27-Aug-99 17-Jun-98 15-Jun-96 16-Jan-98 17-Jun-98 22-Aug-97 10-Aug-98 8-Feb-99 22-Aug-97 9-Jun-99 17-Jun-98 21-Apr-97 19-OCT- 1995 23-Nov-98 23-Nov-98 19-MAY- 1999 05-MAR- 1997

Table 4 (continued)

rxa01514 711 rxa01515 975 rxa01516 513 rxa01517 600 rxa01521 921 rxa01528 651 rxa01551 1998 rxa01561 1053 rxa01599 1785

GB_BA1：MTY15C10 33050 295436 GB_BA1：MTCY7H78 24244 Z95557 GB_BA1：MLCB2548 38916 AL023093 GB_PL1：EGGTPCHI 242 Z49757 GB_BA1：ECOUW93 338534 U14003 GB_BA1：ECOUW93 338534 U14003 GB_BA1：MTCY49 39430 Z73966 GB_IN1：DME238847 5419 AJ238847 GB_HTG3：AC009210 103814 AC009210 GB_IN2：AF132179 4842 AF132179 GB_PL2：F6H8 82596 AF178045 GB_PL2：AF038831 647 AF038831 GB_PL2：ATAC005957 108355 AC005957 GB_BA1：ANANIFBH 5936 J05111 GB_PR2：AC002461 197273 AC002461 GB_PR2：AC002461 197273 AC002461 GB_RO：MM437P9 165901 AL049866 GB_PR3：AC005740 186780 AC005740 GB_PR3：AC005740 186780 AC005740 GB_BA1：MTCY22G10 35420 Z84724 GB_BA2：ECOUW89 176195 U00006 GB_BA1：SCQ11 15441 AL096823 GB_IN1：CEY62HgA 47396 AL032630 GB_PR4：HSU51003 3202 U51003 GB_OM：PIGDAO1 395 M18444 GB_BA1：MTC1125 37432 Z98268 GB_BA1：U00021 39193 U00021

Mycobacterium tuberculosis H37Rv complete genome；segment 154/162. Mycobacterium tuberculosis H37Rv complete genome；segment 153/162. Mycobacterium leprae cosmid B2548. E.gracilis mRNA for GTP cyclohydrolese I(core region). Escherichia coli K-12 chromosomal region from 92.8 to 00.1 minutes. Escherichia coli K-12 chromosomal region from 92.8 to 00.1 minutes. Mycobacterium tuberculosis H37Rv complete genome；segment 93/162. Drosophila melanogaster mRNA for drosophila dodeca-satellite protein 1 (DDP-1). Drosophila melanogaster chromosome 2 clone BACR01106(D1054)RPC1-98 01.1.6 map 55D-55D strain y；cn bw sp，***SEQUENCING IN PROGRESS ***，86 unordered pieces. Drosophila melanogaster clone LD21677 unknown mRNA. Arabidopsis thalina BAC F6H8. Sorosporium saponariae internal transcribed spacer 1，5.8S ribosomal RNA gene；and internal transcribed spaoer 2，complete sequence. Arabidopsis thaliana chromosome II BAC T15J14 genomic sequence， complete sequence. Anabaena sp.(clone AnH20.1)nitrogen fixation operon nifB，fdxN，nifS，nifU， and nifH genes.complete cds. Human BAC clone RG204116 from 7q31，complete sequence. Human BAC clone RG204116 from 7q31，complete sequence. Mus musculus chromosome X，clone 437P9. Homo sapiens chromosome 5p，BAC clone 50g21(LBNL H154)，complete sequence. Homo sapiens chromosome 5p，BAC clone 50g21(LBNL H154)，complete sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 21/162. E.coli chromosomal region from 89.2 to 92.8 minutes. Streptomyces coelicolor cosmid Q11. Caenorhabditis elegans cosmid Y62H9A，complete sequence. Homo sapiens DLX-2(DLX-2)gene，complete cds. Pig D-amino acid oxidase(DAO)gene，exon 1. Mycobacterium tuberculosis H37Rv complete genome；segment 76/162. Mycobacterium leprae cosmid L247.

Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium leprae Euglena gracilis Escherichia coli Escherichia coli Mycobacterium tuberculosis Drosophila melarogaster Drosophila melanogaster Drosophila melanogaster Arabidopsis thaliana Sorosporium saponariae Arabidopsis thaliana Anabaena sp. Homo sapiens Homo sapiens Mus musculus Homo sapiens Homo sapiens Mycobacterium tuberculosis Escherichia coli Streptomyces coelicolor Caenorhabditis elegans Homo sapiens Sus scrofa Mycobacterium tuberculosis Mycobacterium leprae

40,086 43,343 38,177 64,876 38,943 37,500 38,010 36,346 37,897 36,149 35,846 40,566 38,095 38,206 36,623 34,719 37,500 37,031 38,035 38,371 38,064 60,775 38,514 37,730 39,340 63,300 36,756

17-Jun-98 18-Jun-98 27-Aug-99 20-OCT- 1995 17-Apr-96 17-apr-96 24-Jun-99 13-Aug-99 20-Aug-99 3-Jun-99 19-Aug-99 13-Apr-99 7-Jan-99 26-Apr-93 20-Aug-97 20-Aug-97 29-Jun-99 01-OCT- 1998 01-OCT- 1998 17-Jun-98 17-DEC- 1993 8-Jul-99 2-Sep-99 07-DEC- 1999 27-Apr-93 17-Jun-98 29-Sep-94

Table 4 (continued)

rxa01617 rxa01657 rxa01660 rxa01678 rxa01679 rxa01690 rxa01692 rxa01698

795 723 675 651 1359 1224 873 1353

GB_BA1：MLCB51351 38936 Z95117 GB_PR2：HSMTM0 217657 AL034384 GB_PR2：HS13D10 153147 AL021407 GB_PR2：HSMTM0 217657 AL034384 GB_BA1：MTCY1A10 25949 Z95387 GB_EST6：D79278 392 D79278 GB_BA2：AF129925 10243 AF129925 GB_BA1：MTV013 11364 AL021309 GB_RO：MMFV1 6480 X97719 GB_PAT：A67508 6480 A67508 GB_VI：TVU95309 600 U95309 GB_VI：TVU95303 600 U95303 GB_VI：TVU95302 600 U95302 GB_EST5：H91843 362 H91843 GB_STS：G26925 362 G26925 GB_PL2：AF139451 1202 AF139451 GB_8A1：SC1C2 42210 AL031124 GB_EST22：A1064232 493 A1064232 GB_IN2：AF117896 1020 AF117896 GB_BA2：AF057123 1034 AF067123 GB_RO：RATNFHPEP 3085 M37227 GB_RO：RSNFH 3085 X13804 GB_BA2：AF124600 4115 AF124600 GB_BA1：MTCY159 33818 Z83863

Mycobacterium leprae cosmid B1351. Human chromosome Xq28，cosmid clones 7H3，14D7，C1230，11E7，F1096， A12197，12GB，A09100；complete sequence bases 1..217657. Homo sapiens DNA sequence from PAC 13D10 on chromosome 6p22.3-23. Contains CpG island. Human chromosome Xq28，cosmid clones 7H3，14D7，C1230，11E7，F1096， A12197，12GB，A09100；complete sequence bases 1..217657. Mycobacterium tuberculosis H37Rv complete genome；segment 117/162. HUM213D06B Human aorta polyA+(TFujiwara)Homo sapiens cDNA clone GEN-213D06 5′，mRNA sequencce. Thiobacillus ferrooxidans carboxysome operon，complete cds. Mycobacterium tuberculosis H37Rv complete genome；segment 134/162. M.musculus retrovirus restriction gene Fv1. Sequence 1 from Patent WO9743410. Tula virus O64 nucleocapsid protein gene，partial cds. Tula virus O52 nucleocapsid protein gene，partial cds. Tula virus O24 nucleocapsid protein gene，partial cds. ys81e01.s1 Soares retina N2b4HR Homo sapiens cDNA clone IMAGE：221208 3′similar to gb：X63749_ma1 GUANINE NUCLEDTIDE- BINDING PRDTEIN G(T).ALPHA-1(HUMAN)；.mRNA sequence. human STS SHGC-30023.sequence tagged site. Gossypium robinsonii CelA2 pseudogene，partial sequence. Streptomyces coelicolor cosmid 1C2. GH04563.5prime GH Drosophila melanogaster head pOT2 Drosophi1a melanogaster cDNA clone GH04563 5prime，mRNA sequence. Drosophila melanogaster neuropeptide F(npf)gene，complete cds. Lactobacillus reuteri cobalamin biosynthesis protein J(cbiJ)gene.partial cds； and uroporphyrin-III C-methyltransferase(sumT)gene，complete cds. Rat heavy neurofilament(NF-H)polypeptide，partial cds. Rat mRNA for heavy neurofilament polypeptide NF-H C-terminus. Corynebacterium glutamicum chorismate synthase(aroC)，shikimate kinase (aroK).and 3-dehydroquinate synthase(aroB)genes，complete cds；and putative cytoplasmic peptidase(pepQ)gene，partial cds. Mycobacterium tuberculosis H37Rv complete genome；segment 111/162.

Mycobacterium leprae Homo sapiens Homo sapiens Homo sapiens Mycobacterium tuberculosis Homo sapiens Thiobacillus ferrooxidans Mycobacterium tuberculosis Mus musculus Mus musculus Tula virus Tula virus Tula virus Homo sapiens Homo sapiens Gossypium robinsonii Streptomyces coelicolor Drosophila melanogaster Drosophila melanogaster Lactobacillus reuteri Rattus norvegicus Rattus sp. Corynebacterium glutamicum Mycobacterium tuberculosis

36,756 40,811 38,768 39,018 40,656 44,262 40,709 40,986 35,364 35,364 41,894 41,712 39,576 39,157 39,157 38,910 60,644 38,037 36,122 48,079 37,093 37,093 100,000 36,323

24-Jun-97 5-Jul-99 23-Nov-99 5-Jul-99 17-Jun-98 9-Feb-96 17-MAY- 1999 17-Jun-98 29-Aug-96 05-MAY- 1999 28-OCT- 1997 28-OCT- 1997 28-OCT- 1997 29-Nov-95 14-Jun-96 1-Jun-99 15-Jan-99 24-Nov-98 2-Jul-99 3-Jun-98 27-Apr-93 14-Jul-95 04-MAY- 1999 17-Jun-98

Table 4 (continued)

rxa01699 rxa01712 rxa01719 rxa01720 rxa01746 rxa01747 rxa01757

693 805 684 1332 876 1167 924

GB_BA1：MSGB937C 38914 L78820 S GB_8A2：AF124600 4115 AF124600 GB_BA2：AF016585 41097 AF016585 GB_EST9：C19712 399 C19712 GB_EST21：AA952466 278 AA952466 GB_EST21：AA952466 278 AA952466 GB_HTG1：HSDJ534K 154416 AL109925 7 GB_HTG1：HSDJ534K 154416 AL109925 7 GB_EST27：A1447108 431 A1447108 GB_PR4：AC006322 179640 AC006322 GB_PL2：TM018A10 106184 AF013294 GB_PR4：AC006322 179640 AC006322 GB_EST3：R46227 443 R46227 GB_EST3：R46227 443 R46227 GB_BA1：MTCY190 34150 Z70283 GB_BA1：MLCB22 40281 Z98741 GB_BA1：SC5F7 40024 AL096872 GB_EST21：AA918454 416 AA918454 GB_EST4：H34042 345 H34042 G8_EST20：AA899038 450 AA899038

Mycobacterium leprae cosmid B937 DNA sequence. Corynebacterium glutamicum chorismate synthase(aroC)，shikimate kinase (aroK)，and 3-dehydroquinate synthase(aroB)genes，complete cds；and putative cytoplasmic peptidase(pepQ)gene，partial cds. Streptomyces caelestis cytochrome P-450 hydroxylase homolog(nidi)gene， partial cds；polyketide synthase modules 1 through 7(hidA)genes，complete cds；and N-methyltransferase homolog gene，partial cds. C19712 Rice panicle at ripening stage Oryza sativa cDNA clone E10821_1A， mRNA sequence. TENS1404 T.cruzi epimastigote normalized cDNA Library Trypanosoma cruzi cDNA clone 1404 5′，mRNA sequence. TENS1404 T.cruzi epimastigote normalized cDNA Library Trypanosoma cruzi cDNA clone 1404 5′，mRNA sequence. Homo sapiens chromosome 1 clone RP4-534K7，***SEQUENCING IN PROGRESS***，in unordered pieces. Homo sapiens chromosome 1 clone RP4-534K7，***SEQUENCING IN PROGRESS***，in unordered pieces. mq91e08.x1 Stratagene mouse heart(#937316)Mus musculus cDNA clone IMAGE：586118 3′，mRNA sequence. Homo sapiens PAC clone DJ1060B11 from 7q11.23-q21.1，complete sequence. Arabidopsis thaliana BAC TM018A10. Homo sapiens PAC clone DJ1060B11 from 7q11.23-q21.1，complete sequence. yg52a03.s1 Soares infant brain 1NIB Homo sapiens cDNA clone IMAGE：36000 3′，mRNA sequence. yg52a03.s1 Soares infant brain 1NIB Homo sapiens cDNA clone IMAGE：36000 3′，mRNA sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 98/162. Mycobacterium leprae cosmid B22. Streptomyces coelicolor cosmid 5F7. om38c02.s1 Soares_NFL_T_GBC_S1 Homo sapiens cDNA clone IMAGE：1543298 3′similar to WP：F28F8.3 CE09757 SMALL NUCLEAR RIBONUCLEOPROTEIN E；.mRNA sequence. EST110563 Rat PC-12 cells，NGF-treated(9 days)Rattus sp.cDNA clone RPNB181 5′end，mRNA sequence. NCP6G8T7 Perithecial Neurospora crassa cDNA clone NP6G8 3′end，mRNA sequence.

Mycobacterium leprae Corynebacterium glutamicum Streptomyces caelestis Oryza sativa Trypanosoma cruzi Trypanosoma cruzi Homo sapiens Homo sapiens Mus musculus Homo sapiens Arabidopsis thaliana Homo sapiens Homo sapiens Homo sapiens Mycobacterium tuberculosis Mycobacterium leprae Streptomyces coelicolor A3(2) Homo sapiens Rattus sp. Neurospora crassa

62,780 100,000 40,260 45,425 40,876 41,367 35,651 35,651 39,671 35,817 35,698 37,243 42,812 42,655 59,294 57,584 61,810 39,655 35,942 40,000

15-Jun-96 04-MAY- 1999 07-DEC- 1997 24-OCT- 1996 29-OCT- 1998 29-OCT- 1998 23-Nov-99 23-Nov-99 09-MAR- 1999 18-MAR- 1999 12-Jul-97 18-MAR- 1999 22-MAY- 1995 22-MAY- 1995 17-Jun-98 22-Aug-97 22-Jul-99 23-Jun-98 2-Apr-98 12-Apr-98

Table 4 (continued)

rxa01807 rxa01821 rxa01835 rxa01850 rxa01878 rxa01892 rxa01894 rxa01920

915 401 654 1470 1002 852 978 1125

GB_BA1：AP000063 185300 AP000063 GB_HTG4：AC010694 115857 AC010694 GB_HTG4：AC010694 115857 AC010694 GB_BA1：CGL007732 4460 AJ007732 GB_RO：RATALGL 7601 M24108 GB_OV：APIGY2 1381 X78272 GB_EST30：A1629479 353 A1629479 GB_STS：G48245 515 G48245 GB_GSS3：B49052 515 B49052 GB_BA2：ECOUw67_ 110000 U18997 0 GB_BA2：AE000392 10345 AE000392 GB_BA2：U32715 13136 U32715 GB_HTG1：CEY64F11 177748 Z99776 GB_HTG1：CEY64F11 177748 Z99776 GB_HTG1：CEY64F11 177748 Z99776 GB_BA1：MTCY274 39991 Z74024 GB_BA1：MLCB250 40603 Z97369 GB_BA1：MSGB1529C 36985 L78824 S GB_BA1：MTCY274 39991 Z74024 GB_IN1：CELF46H5 38886 U41543 GB_HTG3：AC009204 115633 AC009204 GB_BA2：AF112536 1798 AF112536 GB_BA1：CANRDFGE 6054 Y09572 N

Aeropyrum pemix genomic DNA，section 677. Drosophila melanogaster clone RPC198-6H2，***SEQUENCING IN PROGRESS***，75 unordered pieces. Drosophila melanogaster clone RPCI98-6H2，***SEQUENCING IN PROGRESS***，75 unordered pieces. Corynebacterium glutamicum 3′ppc gene，secG gene，amt gene，ocd gene and 5′soxA gene. Rattus norvegicus(clone A2U42)alpha2u globulin gene，exons 1-7. Anas platyrhynchos(Super M)lgY upsilon heavy chain gene，exon 2. 486101D10.x1 486-leaf primordia cDNA library from Hake lab Zea mays cDNA，mRNA sequence. SHGC-62915 Human Homo sapiens STS genomic，sequence tagged site. RPC111-4112.TV RPC1-11 Homo sapiens genomic clone RPC1-11-4112. genomic survey sequence. Escherichia coli K-12 chromosomal region from 67.4 to 76.0 minutes. Escherichia coli K-12 MG1655 section 282 of 400 of the complete genome. Haemophilus influenzae Rd section 30 of 163 of the complete genome. Caenorhabditis elegans chromosome IV clone Y64F11，***SEQUENCING IN PROGRESS***，in unordered pieces. Caenorhabditis elegans chromosome IV clone Y64F11，***SEQUENCING IN PROGRESS***，in unordered pieces. Caenorhabditis elegans chromosome IV clone Y64F11，***SEQUENCING IN PROGRESS***，in unordred pieces. Mycobacterium tuberculosis H37Rv complete genome；segment 126/162. Mycobacterium leprae cosmid B250. Mycobacterium leprae cosmid B1529 DNA sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 126/162. Caenorhabditis elegans cosmid F46H5. Drosophila melanogaster chromosome 2 clone BACR03E19(D1033)RPC1-98 03.E.19 map 36E-37C strain y；cn bw sp，***SEQUENCING IN PROGRESS ***，94 unordered pieces. Corynebacterium glutamicum ribonucleotide reductase beta-chain(nrdF) gene，complele cds. Corynebacterium ammoniagenes nrdH，nrdI，nrdE，nrdF genes.

Aeropyrum pernix Drosophila melanogaster Drosophila melanogaster Corynebacterium glutamicum Rattus norvegicus Anas platyrhynchos Zea mays Homo sapiens Homo sapiens Escherichia coli Escherichia coli Haemophilus influenzae Rd Caenorhabditis elegans Caenorhabditis elegans Caenorhabditis elegans Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium leprae Mycobacterium tuberculosis Caenorhabditis elegans Drosophila melanogaster Corynebacterium glutamicum Corynebacterium ammoniagenes

40,067 35,450 35,450 100,000 38,692 36,962 38,109 37,021 37,021 37,196 38,021 39,860 37,564 37,564 37,576 35,910 64,260 64,260 37,229 38,525 31,579 99,733 70,321

22-Jun-99 16-OCT- 1999 16-OCT- 1999 7-Jan-98 15-DEC- 1994 15-Feb-99 26-Apr-99 26-MAR- 1999 8-Apr-99 U18997 12-Nov-98 29-MAY- 1998 14-OCT- 1998 14-OCT- 1998 14-OCT- 1998 19-Jun-98 27-Aug-99 15-Jun-96 19-Jun-98 29-Nov-96 18-Aug-99 5-Aug-99 18-Apr-98

Table 4 (continued)

rxa01928 rxa01929 rxa01940 rxa02022 rxa02024 rxa02027 rxa02031 rxa02072

960 936 1059 1230 859 1464

GB_BA2：AF050168 1228 AF050168 GB_BA1：CGPAN 2164 X96580 GB_PL1：AP000423 154478 AP000423 GB_PL1：AP000423 154478 AP000423 GB_BA1：CGPAN 2164 X96580 GB_BA1：XCU33548 8429 U33548 GB_BA1：XANHRPB6 1329 M99174 A GB_IN2：CFU43371 1060 U43371 GB_BA2：AE001467 11601 AE001467 GB_RO：AF175967 3492 AF175967 GB_BA1：CGDAPE 1966 X81379 GB_BA1：CGDNAARO 2612 X85965 P GB_BA1：APU47055 6469 U47055 GB_BA1：MTC1364 29540 Z93777 GB_BA1：MSGB1912C 38503 L01536 S GB_BA1：MLU15180 38675 U15180 GB_BA1：CGGDHA 2037 X72855 GB_BA1：CGGDH 2037 X59404 GB_BA1：PAE18494 1628 Y18494

Corynebacterium ammoniagenes ribonucleoside diphosphate neductase small subunit(nrdF)gene，complete cds. C.glulamicum panB，panC & xylB genes. Arabidopsis thaliana chloroplast genomic DNA，complete sequence， strain：Columbia. Arabidopsis thaliana chloroplast genomic DNA，complete sequence， strain：Columbia. C.glutamicum panB，panC &xyIB genes. Xanthomonas campestris hrpB pathogenicity locus proteins HrpB1，HrpB2， HrpB3，HrpB4，HrpB5，HrpB6，HrpB7，HrpB8，HrpA1，and ORF62 genes，complete cds. Xanthomonas campestris hrpB6 gene，complete cds. Crithidia fasciculata inosine-uridine preferring nucleoside hydrolase(IUNH) gene，complete cds. Helicobacter pylori，strain J99 section 28 of 132 of the complete genome. Homo sapiens Leman coiled-coil protein(LCCP)mRNA，complete cds. C.glutamicum dapE gene and orf2. C.glutamicum ORF3 and aroP gene. Anabaena PCC7120 nitrogen fixation proteins(nifE，nifN，nifX，nifW)genes. complete cds，and nitrogenase(nifK)and hesA genes，partial cds. Mycobacterium tuberculosis H37Rv complete genome；segment 52/162. M.leprae genomic dna sequence，cosmid b1912. Mycobacterium leprae cosmid B1756. C.glutamicum GDHA gene. Corynebacterium glutamicum，gdh gen fo rglutamate dehydrogenase. Pseudomonas aeruginosa gdhA gene，strain PAC1.

Corynebacterium ammoniagenes Corynebacterium glutamicum Chloroplast Arabidopsis thaliana Chloroplast Arabidopsis thaliana Corynebacterium glutamicum Xanthomonas campestris pv.vesicatoria Xanthomonas campestris Crithidia fasciculata Helicobacter pylori J99 Mus musculus Corynebacterium glutamicum Corynebacterium glutamicum Anabaena PCC7120 Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium leprae Corynebacterium glutamicum Corynebacterium glutamicum Pseudomonas aeruginosa

72,082 100,000 35,917 33,925 100,000 38,749 39,305 61,417 38,560 40,275 100,000 38,889 36,647 59,415 57,093 57,210 99,317 94,387 62,247

23-Apr-98 11-MAY- 1999 15-Sep-99 15-Sep-99 11-MAY- 1999 19-Sep-96 14-Sep-93 18-Jun-96 20-Jan-99 26-Sep-99 8-Aug-95 30-Nov-97 17-Feb-96 17-Jun-98 14-Jun-96 09-MAR- 1995 24-MAY- 1993 30-Jul-99 6-Feb-99

Table 4 (continued)

rxa02085 rxa02093 rxa02106 rxa02111 rxa02112 rxa02134 rxa02135

2358 927 1179 1407 960 1044 1197

GB_BA1 ：MTCY22G8 22550 Z95585 GB_BA1：MLCB33 42224 Z94723 GB_BA1：ECOUW85 91414 M87049 GB_EST14：AA448146 452 AA448146 GB_EST17：AA641937 444 AA641937 GB_PR3：AC003074 143029 AC003074 GB_BA1：SC1A6 37620 AL023496 GB_PR4：AC005553 179651 AC005553 GB_EST3：R49746 397 R49746 GB_BA1：SC6G10 36734 AL049497 GB_BA1：U00010 41171 U00010 GB_BA1：MTCY336 32437 Z95586 GB_HTG3：AC010579 157658 AC010579 GB_GSS3：B09839 1191 B09839 GB_HTG3：AC010579 157658 AC010579 GB_BA1：SCSECYDN 6154 X83011 A GB_EST32：A1731596 568 A1731596 GB_BA1：SCSECYDN 6154 X83011 A GB_PR3：HS525L6 168111 AL023807 GB_PL2：ATF21P8 85785 AL022347 GB_PL2：U89959 106973 U89959

Mycobacterium tuberculosis H37Rv complete genome；segment 49l162. Mycobacterium leprae cosmid B33. E.coli genomic sequence of the region from 84.5 to 86.5 minutes. zw82hD1.r1 Soares_testis_NHT Homo sapiens cDNA clone IMAGE：782737 5′.mRNA sequence. ns18b10.r1 NCI_CGAP_GCB1 Homo sapiens cDNA clone IMAGE：1183963 5′.mRNA sequence. Human PAC clone DJ0596009 from 7p15，complete sequence. Streptomyces coelicolor cosmid 1A6. Homo sapiens chromosome 17，clone hRPK.112_J_9，complete sequence. yg71g10.r1 Soares infant brain 1NIB Homo sapiens cDNA clone IMAGE：38768 5′similar to gb：V00567 BETA-2-MICROGLOBULIN PRECURSOR(HUMAN)；，mRNA sequence. Streptomyces coelicolor cosmid 6G10. Mycobacterium leprae cosmid B1170. Mycobacterium tubercu1osis H37Rv complete genome；segment 70/162. Drosophila melanogaster chromnosome 3 clone BACR09D08(D1101)RPCI-98 09.D.8 map 96F-96F strain y；cn bw sp，***SEQUENCING IN PROGRESS ***，121 unordered pieces. T12A12-Sp6 TAMU Arabidopsis thaliana genomic clone T12A12，genomic survey sequence. Drosophila melanogaster chromosome 3 clone BACR09DD08(D1101)RPC1-98 09.D.8 map 96F-96F strain y；cn bw sp，***SEQUENCING IN PROGRESS ***，121 unordered pieces. S.coelicolor secY locus DNA. BNLGHi10185 Six-day Cotton fiber Gossypium hirsutum cDNA 5′similar to (AC004005)putative ribosomal protein L7[Arabidopsis thaliana]，mRNA sequence. S.coelicolor secY locus DNA. Human DNA sequence from clone RP3-525L6 on chromosome 6p22.3-23 Contains CA repeat，STSs，GSSs and a CpG Island，complete sequence. Arabidopsis thaliaha DNA chromosome 4，BAC clone F21P8(ESSA project). Arabidopsis thaliana BAC T7123，complete sequence.

Mycobacterium tuberculosis Mycobacterium leprae Escherichia coli Homo sapiens Homo sapiens Homo sapiens Streptomyces coelicolor Homo sapiens Homo sapiens Streptomyces coelicolor Mycobacterium leprae Mycobacterium tuberculosis Drosophila melanogaster Arabidopsis thaliana Drosophila melanogaster Streptomyces coelicolor Gossypium hirsutum Streptomyces coelicolor Homo sapiens Arabidopsis thaliana Arabidopsis thaliana

38,442 56,486 52,127 34,163 35,586 31,917 35,818 34,274 41,162 50,791 37,563 39,504 37,909 37,843 37,909 36,533 33,451 36,756 34,365 34,325 33,874

17-Jun-98 24-Jun-97 29-MAY- 1995 4-Jun-97 27-OCT- 1997 6-Nov-97 13-Jan-99 31-DEC- 1998 18-MAY- 1995 24-MAR- 1999 01-MAR- 1994 24-Jun-99 24-Sep-99 14-MAY- 1997 24-Sep-99 02-MAR- 1998 11-Jun-99 02-MAR- 1998 23-Nov-99 9-Jun-99 26-Jun-98

Table 4 (continued)

rxa02136 rxa02139 rxa02153 rxa02154 rxa02155 rxa02156

645 1962 903 414 1287 1074

GB_PL2：ATAC005819 57752 AC005819 GB_PL2：F15K9 71097 AC005278 GB_PL2：U89959 106973 U89959 GB_BA1：MTCY190 34150 270283 GB_BA1：MSGB1554C 36548 L78814 S GB_BA1：MSGB1551C 36548 L78813 S GB_BA2：AF049897 9196 AF049897 GB_BA1：AF005242 1044 AF005242 GB_BA1：CGARGCJB 4355 X86157 D GB_BA2：AF049897 9196 AF049897 GB_BA1：AF005242 1044 AF005242 GB_BA1：CGARGCJB 4355 X86157 D GB_BA1：CGARGCJB 4355 X86157 D GB_BA2：AF049897 9196 AF049897 GB_BA1：MSGB1133C 42106 L78811 S GB_BA2：AF049897 9196 AF049897

Arabidopsis thaliana chromosome II BAC T3A4 genomic sequence，complete sequence. Arabidopsis thaliana chromosome 1 BAC F15K9 sequence，complete sequence. Arabidopsis thaliana BAC T7123，complete sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 98/162. Mycobacterium leprae cosmid B1 554 DNA sequence. Mycobacterium leprae cosmid B1 551 DNA sequence. Corynebacterium glutamicum N-acetylglutamylphosphate reductase(argC)， omithine acetyltransferase(argJ)，N-acetylglutamate kinase(argB)， acetylomithine transaminase(argD)，omithine carbamoyltransferase (argF)，arginine repressor(argR)，argininosuccinate synthase(argG)，and argininosuccinate lyase(argH)genes，complete cds. Corynebacterium glutamicum N-acetylglutamate-5-semialdehyde dehydrogenase(argC)gene，complete cds. C.glutamicum argC，argJ，argB，argD，and argF genes. Corynebacterium glutamicum N-acetylglutamylphosphate reductase(argC)， ornithine acetyltransferase(argJ)，N-acetylglutamate kinase(argB)， acetyfornithine transaminase(argD)，omithine carbamoyltransferase(argF)， arginine repressor(argR)，argininosuccinate synthase(argG)，and argininosuccinate lyase(argH)genes，comptete cds. Corynebacterium glutamicum N-acetylglutamate-5-semialdehyde dehydrogenase(argG)gene，complete cds. C.glutamicum argC，argJ，argB，argD，and argF genes. C.glutamicum argC，argJ，argB，argD，and argF genes. Corynebacterium glutamicum N-acetylglutamylphosphate reductase(argC)， omithine acetyltransferase(argJ)，N-acetylglutamate kinase(argB)， acetylomithine transaminase(argD)，omithine carbamoyltransferase(argF)， arginine repressor(argR)，argininosuccinate synthase(argG)，and argininosuccinate lyase(argH)genes，complete cds. Mycobacterium leprae cosmid B1133 DNA sequence. Corynebaclerium glutamicum N-acetylglutamylphosphate reductase(argC)， ornithine acetyltransferase(argJ)，N-acetylglutamate kinase(argB)， acetylomithine transaminase(argD)，omithine carbamoyltransferase(argF)， arginine repressor(argR)，argininosuccinate synthase(argG)，and argininosuccinate lyase(argH)genes，complete cds.

Arabidopsis thaliana Arabidopsis thaliana Arabidopsis thaliaha Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium leprae Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Mycobacterium leprae Corynebacterium glutamicum

34,123 31,260 34,281 62,904 36,648 36,648 99,104 99,224 100,000 98,551 98,477 100,000 99,767 99,378 55,504 100,000

3-Nov-98 7-Nov-98 26-Jun-98 17-Jun-98 15-Jun-96 15-Jun-96 1-Jul-98 2-Jul-97 25-Jul-96 1-Jul-98 2-Jul-97 25-Jul-96 25-Jul-96 1-Jul-98 15-Jun-96 1-Jul-98

Table 4 (continued)

rxa02157 rxa02158 rxa02159 rxa02160 rxa02162

1296 1080 636 1326 1554

GB_BA1：CGARGCJB 4355 X86157 D GB_BA2：AE001816 10007 AE001816 GB_BA2：AF049897 9196 AF049897 GB_BA1：CGARGCJB 4355 X86157 D GB_BA1：MTCY06H11 38000 Z85982 GB_BA2：AF049897 9196 AF049897 GB_BA2：AF031518 2045 AF031518 GB_BA1：CGARGCJB 4355 X86157 D GB_BA2：AF049897 9196 AF049897 GB_BA2：AF031518 2045 AF031518 GB_BA2：AF041436 516 AF041436 GB_BA2：AF049897 9196 AF049897 GB_BA2：AF030520 1206 AF030520 GB_BA1：SCARGGH 1909 Z49111 GB_BA2：AF049897 9196 AF049897

C.glutamicum argC，argJ，atgB，argD，and argF genes. Thermotoga maritima section 128 of 136 of the complete genome. Corynebacterium glutamicum N-acetylglutamylphosphate reductase(argC)， ornithine acetyltransferase(argJ)，N-acetylglutamate kinase(argB)， acetylomithine transaminase(argD)，omithine carbamoyltransferase(argF)， arginine repressor(argR)，argininosuccinate synthase(argG)，and argininosuccinate lyase(argH)genes，complete cds. C.glutamicum argC，argJ，argB，argD，and argF genes. Mycobacterium tuberculosis H37Rv complete genome；segmant 73/162. Corynebacterium glutamicum N-acetylglutamylphosphate reductase(argC)， ornithine acetyltransferase(argJ)，N-acetylglutamate kinase(argB)， acetylomithine transaminase(argD)，ornithine carbamoyltransferase(argF)， arginine repressor(argR)，argininosuccinate synthase(argG)，and argininosuccinate tyase(argH)genes，complete cds. Corynebacterium glutamicum omithine carbamolytransferase(argF)gene， complete cds. C.glutamicum argC，argJ，argB，argD，and argF genes. Corynebacterium glutamicum N-acetylglutamylphosphate reductase(argC)， omithine acetyltransferase(argJ)，N-acetylglutamate kinase(argB)， acetylomithine transaminase(argD)，ornithine carbamoyltransferase(argF)， arginine repressor(argR)，argininosuccinate synthase(argG)，and argininosuccinate lyase(argH)genes，complete cds. Corynebacterium glutamicum ornithine carbamolytransferase(argF)gene， complete cds. Corynebacterium glutamicum arginine repressor(argR)gene，complete cds. Corynebacterium glutamicum N-acetylglutamylphosphate reductase(argC)， omithine acetyllransferase(argJ).N-acetylglutamate kinase(argB)， acetylornithine transaminase(argD)，ornithine carbamoyltransferase(argF)， arginine repressor(argR)，argininosuccinate synthase(argG)，and argininosuccinate lyase(argH)genes，complete cds. Corynebacterium glutamicum argininosuccinate synthetase(argG)gene， complete cds. S.clavuligerus argG gene and argH gene(partial). Corynebacterium glutamicum N-acetylglutamylphosphate reductase(argC)， omithine acetyltransferase(argJ)，N-acetylglutamate kinase(argB)， acetylomithine transaminase(argD)，omithine carbamoyltransferase(argF)， arginine repressor(argR)，argininosuccinate synthase(argG)，and argininosuccinate lyase(argH)genes，complete cds.

Corynebacterium glutamicum Thermologa maritima Corynebacterium glutamicum Corynebacterium glutamicum Mycobacterium tuberculosis Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Streptomyces clavuligerus Corynebacterium glutamicum

100,000 50,238 99,612 99,612 57,278 100,000 99,898 100,000 99,843 88,679 100,000 99,774 99,834 65,913 88,524

25-Jul-96 2-Jun-99 1-Jul-98 25-Jul-96 17-Jun-98 1-Jul-98 5-Jan-99 25-Jul-96 1-Jul-98 5-Jan-99 5-Jan-99 1-Jul-98 19-Nov-97 22-Apr-96 1-Jul-98

Table 4 (continued)

rxa02176 1251 rxa02189 861 rxa02193 1701 rxa02194 966 rxa02195 393 rxa02197 551 rxa02198 2599 rxa02208 1025

GB_BA2：AF048764 1437 AF048764 GB_BA1：MTCY06H11 38000 Z85982 GB_BA1：MTCY31 37630 Z73101 GB_BA1：CGGLTG 3013 X66112 GB_PL2：PGU65399 2700 U65399 GB_PR3：AC002468 115888 AC002468 GB_BA1：MSGB1970C 39399 L78815 S GB_PR3：AC002468 115888 AC002468 GB_BA1：BRLASPA 1987 D25316 GB_PAT：E04307 1581 E04307 GB_BA1：ECOUW93 338534 U14003 GB_BA2：AF050166 840 AF050166 GB_BA1：BRLASPA 1987 D25316 GB_PAT：E08649 188 E08649 GB_BA2：AF086704 264 AF086704 GB_BA1：EAY17145 6019 Y17145 GB_STS：G01195 332 G01195 GB_BA1：MTCY261 27322 Z97559 GB_BA1：MLCB2533 40245 AL035310 GB_BA1：U00017 42157 U00017 GB_BA1：U00017 42157 U00017 GB_BA1：MLCB2533 40245 AL035310 GB_BA1：MTCY261 27322 Z97559 GB_BA1：U00017 42157 U00017 GB_BA1：AP000063 185300 AP000063

Coryneoacterium glutamicum argininosuccinate lyase(agH)gene，complete cds. Mycobacterium tuberculosis H37Rv complete genome；segment 73/162. Mycobaclerium tuberculosis H37Rv complete genome；segment 41/162. C.glutamicum glt gene for citrate synthase and ORF. Basidiomycete CECT 20197 phenoloxidase(pox1)gene，complete cds. Human Chromosome 15q26.1 PAC clone pDJ417d7，complete sequence. Mycobacterium leprae cosmid B1970 DNA sequence. Human Chromosome 15q26.1 PAC clone pDJ417d7，complete sequence. Brevibacterium flavum aspA gene for aspartase，complete cds. DNA encoding Brevibacterium flavum aspartase. Escherichia coli K-12 chromosomal region from 92.8 to 00.1 minutes. Corynebacterium glutamicum ATP phosphoribosyltransferase(hisG)gene， complete cds. Brevibacterium flavum aspA gene for aspartase，complete cds. DNA encoding part of aspartase from coryneform bacteria. Corynebacterium glutamicum phosphoribosyl-ATP-pyrophosphohydrolase (hisE)gene，complete cds. Eubacterium acidaminophilum grdR，grdI，grdH genes and partial ldc，grdT genes. fruit fly STS Dm1930 clone DS06959 T7. Mycobacterium tuberculosis H37Rv complete genome；segment 95/162. Mycobacterium leprae cosmid B2533. Mycobacterium leprae cosmid B2126. Mycobacterium leprae cosmid B2126. Mycobacterium leprae cosmid B2533. Mycobacterium tuberculosis H37Rv complete genome；segment 95/162. Mycobacterium leprae cosmid B2126 Aeropyrum pemix genomic DNA，seection 6/7.

Corynebacterium glutamicum Mycobacterium tuberculosis Mycobacterium tuberculosis Corynebacterium glutamicum basidiomycete CECT 20197 Homo sapiens Mycobacterium leprae Homo sapiens Corynebacterium glutamicum Corynebacterium glutamicum Escherichia coli Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Eubacterium acidaminophilum Drosophila melanogastar Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium leprae Mycobacterium lepraa Mycobacterium leprae Mycobacterium tuberculosis Mycobacterium leprae Aeropyrum pernix

87,561 64,732 36,998 39,910 38,474 35,941 40,286 33,689 99,353 99,367 37,651 98,214 93,805 100,000 100,000 39,075 35,542 33,938 65,517 36,770 38,674 65,465 37,577 59,823 39,442

1-Jul-98 17-Jun-98 17-Jun-98 17-Feb-95 19-Jul-97 16-Sep-98 15-Jun-96 16-Sep-98 6-Feb-99 29-Sep-97 17-Apr-96 5-Jan-99 6-Feb-99 29-Sep-97 8-Feb-99 5-Aug-98 28-Feb-95 17-Jun-98 27-Aug-99 01-MAR- 1994 01-MAR- 1994 27-Aug-99 17-Jun-98 01-MAR- 1994 22-Jun-99

Table 4 (continued)

rxa02229 948 rxa02234 3462 rxa02235 727 rxa02237 693 rxa02239 1389 rxa02240 1344 rxa02246 1107

GB_PR4：AC006236 127593 AC006236 GB_BA1：MSGY154 40221 AD000002 GB_BA1：MTCY154 13935 Z98209 G3_BA1：U00019 36033 U00019 GB_BA1：MSGB937C 38914 L78820 S GB_BA1：MTCY2B12 20431 Z81011 GB_BA2：UD1072 4393 U01072 GB_BA1：MSU91572 960 U91572 GB_HTG3：AC009364 192791 AC009364 GB_HTG3：AC009364 192791 AC009364 GB_BA1：MTCY21B4 39150 Z80108 GB_BA2：AF077324 5228 AF077324 GB_EST22：AU017763 586 AU017763 GB_BA1：MTCY2184 39150 Z80108 GB_HTG3：AC010745 193862 AC010745 GB_HTG3：AC010745 193862 AC010745 EM_PAT：E09855 1239 E09855 GB_PAT：A37831 5392 A37831 GB_BA2：AF117274 2303 AF117274 EM_BA1：AB003693 5589 AB003693

Homo sapiens chromosome 17，clone hCIT.162_E_12，complete sequence. Mycobacterium tuberculosis sequence from clone y154. Mycobacterium tuberculosis H37Rv complete genome；segment 121/162. Mycobacterium leprae cosmid B2235. Mycobacterium leprae cosmid B937 DNA sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 61/162. Mycobacterium bovis BCG orotidine-5′-monophosphate decarboxylase(uraA) gene. Mycobacterium smegmatis carbamoyl phosphate synhetase(pyrAB)gene， partial cds and orotidine 5′-monophosphate decarboxylase(pyrF)gene， complete cds. Homo sapiens chromosome 7，***SEQUENCING IN PROGRESS***，57 unordered pieces. Homo sapiens chromosome 7，***SEQUENCING IN PROGRESS***，57 unordered pieoes. Mycobacterium tuberculosis H37Rv complete genome；segment 62/162. Rhodococcus equi strain 103 plasmid RE-VP1 fragment f. AU017763 Mouse two-cell stage embryo cDNA Mus musculus cDNA clone J0744A04 3′.mRNA sequence. Mycobacterium tutberculosis H37Rv complete genome；segment 62/162. Homo sapiens clone NH0549D18，***SEQUENCING IN PROGRESS***，30 unordered pieces. Homo sapiens clone NH0549D18，***SEQUENCING IN PROGRESS***，30 unordered pieces. gDNA encoding S-adenosylmethionine synthelase. Sequence 1 from Patent WO9408014. Streptomyces spectabilis flavoprotein homolog Dfp(dfp)gene，partial cds；and S-adenosylmethionine synthetase(metK)gene，complete cds. Corynebacterium ammoniagenes DNA for rib operon，complete cds.

Homo sapiens Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium leprae Mycobacterium tuberculosis Mycobacterium bovis Mycobacterium smegmatis Homo sapiens Homo sapiens Mycobacterium tuberculosis Rhodococcus equi Mus musculus Mycobacterium tuberculosis Homo sapiens Homo sapiens Corynebacterium glutamicum Streptomyces pristinaespiralis Streptomyces spectabilis Corynebacterium ammoniagenes

37,191 53,541 40,407 40,541 66,027 71,723 67,101 60,870 37,994 37,994 55,844 41,185 38,616 56,282 36,772 36,772 99,515 63,568 65,000 52,909

29-DEC- 1998 03-DEC- 1996 17-Jun-98 01-MAR- 1994 15-Jun-96 18-Jun-98 22-DEC- 1993 22-MAR- 1997 1-Sep-99 1-Sep-99 23-Jun-98 5-Nov-98 19-OCT- 1998 23-Jun-98 21-Sep-99 21-Sep-99 07-OCT- 1997(Rel. 52，Crealed 05-MAR- 1997 31-MAR- 1999 03-OCT- 1997(Rel. 52，Created)

Table 4 (continued)

rxa02247 756 rxa02248 1389 rxa02249 600 rxa02250 643 rxa02262 1269 rxa02263 488 rxa02272 1368 rxa02281 1545

GB_PAT：E07957 5589 ED7957 GB_PAT：132742 5589 132742 GB_PAT：132743 2689 132743 EM_BA1：AB003693 5589 AB003693 GB_PAT：132742 5589 132742 GB_PAT：132742 5589 132742 EM_BA1：AB003693 5589 AB003693 GB_PAT：E07957 5589 E07957 GB_PAT：E07957 5589 E07957 GB_PAT：132742 5589 132742 GB_PAT：132743 2689 132743 GB_PAT：E07957 5589 E07957 GB_PAT：132742 5589 132742 EM_BA1：AB003693 5589 AB003693 GB_BA1：CGL007732 4460 AJ007732 GB_BA1：CGAMTGEN 2028 X93513 E GB_VI：HEHCMVCG 229354 X17403 GB_BA1：CGL007732 4460 AJ007732 GB_BA1：CGL007732 4460 AJ007732 EM_PAT：E09373 1591 E09373 GB_BA1：D38505 1591 D38505 GB_HTG2：AC006595 146070 AC006595 GB_GSS12：AQ41101 551 AQ411010 O

gDNA encoding at least guanosine triphosphate cyclohydrolase and riboflavin synthase. Sequence 1 from patent US 5589355. Sequence 2 from patent US 5589355. Corynebacterium ammoniagenes DNA for rib operon，complete cds. Sequence 1 from patent US 5589355. Sequence 1 from patent US 5589355. Corynebacterium ammoniagenes DNA for rib operon，complete cds. gDNA encoding at least guanosine triphosphate cyclohydrolase and riboflavin synthase gDNA encoding at least guanosine triphosphate cyclohydrolase and riboflavin synthase. Sequence 1 from patent US 5589355. Sequence 2 from patent US 5589355. gDNA encoding at least guanosine triphosphate cyclohydrolase and riboflavin synthase. Sequence 1 from patent US 5589355. Corynebacterium ammoniagenes DNA for rib operon，complete cds. Corynebacterium glutamicum 3′ppc gene，secG gene，amt gene，ocd gene and 5′soxA gene. C.glutamicum amt gene. Human cytomegaloviris strain AD169 complete genome. Corynebacterium glutamicum 3′ppc gene，secG gene，amt gene，ocd gene and 5′soxA gene. Corynebacterium glutamicum 3′ppc gene，secG gene，amt gene，ocd gene and 5′soxA gene. Creatinine deiminase gene. Bacillus sp.gene for creatinine deaminase，conplete cds. Homo sapiens，***SEQUENCING IN PROGRESS***，4 unordered pieces. HS_2257_B1_H02_MR CIT Approved Human Genomic Sperm Library D Homo sapiens genomic clone Plate＝2257 Col＝3 Row＝P，genomic survey sequence.

Corynebacterium ammoniagenes Unknown. Unknown. Corynebacterium ammoniagenes Unknown. Unknown. Corynebacterium ammoniagenes Corynebacterium ammoniagenes Corynebacterium amoniagenes Unknown. Unknown. Corynebacterium ammoniagenes Unknown. Corynebacterium ammoniagenes Corynebacterium gluamicum Corynebacterium glutamicum human herpesvirus 5 Corynebacterium glutamicum Corynebacterium glutamicum Bacillus sp. Bacillus sp. Homo sapiens Homo sapiens

52,909 52,909 57,937 57,937 57,937 61,843 61,843 61,843 64,346 64,346 64,346 56,318 56,318 56,318 100,000 100,000 38,651 100,000 37,526 96,928 96,781 36,264 36,197

29-Sep-97 6-Feb-97 6-Feb-97 03-OCT- 1997(Re1. 52，Created) 6-Feb-n97 6-Feb-97 03-OCT- 1997(Rel. 52，Created) 29-Sep-97 29-Sep-97 6-Feb-97 6-Feb-97 29-Sep-97 6-Feb-97 03-OCT- 1997(Rel 52，Created 7-Jan-99 29-MAY- 1996 10-Feb-99 7-Jan-99 7-Jan-99 08-OCT- 1997(Rel. 52，Created) 7-Aug-98 20-Feb-99 17-MAR- 1999

Table 4 (continued)

rxa02299 531 rxa02311 813 rxa02315 1752 rxa02318 402 rxa02319 1080

GB_EST23：A1128623 363 A128623 GB_PL2：ATAC007019 102335 AC007019 GB_BA2：AF116184 540 AF116184 GB_GSS9：AQ164310 507 AQ164310 GB_VI：MH68TKH 4557 X93468 GB_HTG4：AC006091 176878 AC006091 GB_HTG4：AC006091 176878 AC006091 GB_BA2：RRU65510 16259 U65510 GB_BA1：MSGY224 40051 AD000004 GB_3A1：MTY25D10 40838 Z95558 GB_BA1：MSGY224 40051 AD000004 GB_HTG3：AC011348 111083 AC011348 GB_HTG3：AC011348 111083 AC011348 GB_HTG3：AC011412 89234 AC011412 GB_BA1：MSGY224 40051 AD000004 GB_BA1：MTY25D10 40838 Z95558 GB_EST23：A117213 476 A1117213

qa62c01.s1 Soares_fetal_heart_NbHH19VV Homo sapiens cDNA clone IMAGE：1691328 3′，mRNA sequence. Arabidopsis thaliana chromosome II BAC F7D8 genomic sequence，complete sequence. Corynebacterium glutamicum L-aspartate-alpha-decarboxylase precursor (panD)gene，complete cds. HS_2171_A2_E01_MR CIT Approved Human Genomic Sperm Library D Homo sapiens genomic clone Plate＝2171 Col＝2 Row＝1，genomic survey sequence. Murine herpesvirus type 68 thymidine kinase and glycoprotein H genes. Drosophila melanogaster chromosome 3 clone BACR48G05(D475)RPC1-98 48.G.5 map 91F1-91F13 strain y；cn bw sp，***SEQUENCING IN PROGRESS***，4 unordered pieces. Drosophila melanogaster chromosome 3 clone BACR48G05(D475)RPCI-98 48.G.5 map 91F1-91F13 strain y；cn bw sp，***SEQUENCING IN PROGRESS***，4 unorderd pieces. Rhodospirillum rubrum CO-induced hydrogenase operon(cooM，cooK，cooL， cooX，cooU，cooH)geres，iron sulfur protein(cooF)gene，carbon monoxide dehydrogenase(cooS)gene，carbon monoxide dehydrogenase accessory proteins(cooC，cooT，cooJ)genes.putative transcriptional activator (cooA)gene，nicotinate-nucleotide pyrophosphorylase(nadC)gene，complete cds.L-aspartate oxidase(nadB)gene.and alkyl hydroperoxide reductase(ahpC)gene，partial cds. Mycobacterium tuberculosis sequence from clone y224. Mycobacteriun tuberculosis H37Rv complete genome；segment 28/162. Mycobacterium tuberculosis sequence from clone y224. Homo sapiens chromosome 5 clone CIT-HSPC_303E13，***SEQUENCING IN PROGRESS***，3 ordered pieces. Homo sapiens chromosome 5 clone CIT-HSPC_303E13，***SEQUENCING IN PROGRESS***，3 orderd pieces. Homo sapiens chromosome 5 clone CIT978SKB_81K21，***SEQUENCING IN PROGRESS***，3 ordred pieces. Mycobacterium tuberculosis sequence from clone y224. Mycobacterium tuberculosis H37Rv complete genome；segment 28/162. ub83h02.r1 Soares 2NbMT Mus musculus cDNA clone IMAGE：1395123 5′，mRNA sequence.

Homo sapiens Arabidopsis thaliana Corynebacterium glutamicum Homo sapiens murine herpesvirus 68 Drosophila melanogaster Drosophila melanogaster Rhodospirillum rubrum Mycobacterium tuberculosis Mycobacterium tuberculosis Mycobacterium tuberculosis Homo sapiens Homo sapiens Homo sapiens Mycobaclerium tuberculosis Mycobacterium tuberculosis Mus musculus

37,017 33,988 100,000 37,278 40,288 36,454 36,454 37,828 49,418 49,360 38,150 35,821 35,821 36,181 37,792 37,92 35,084

05-OCT- 1998 16-MAR- 1999 02-MAY- 1999 16-OCT- 1998 3-Sep-96 27-OCT- 1999 27-OCT- 1999 9-Apr-97 03-DEC- 1996 17-Jun-98 03-DEC- 1996 06-OCT- 1999 06-OCT- 1999 06-OCT- 1999 03-DEC- 1996 17-Jun-98 2-Sep-98

Table 4 (continued)

rxa02345 rxa02350 rxa02373 rxa02375 rxa02380 rxa02382 rxa02400 rxa02432

1320 618 1038 1350 777 1419 693 1098

GB_BA1：BAPURKE 2582 X91189 GB_BA1：MTCY71 42729 Z92771 GB_BA1：MTCY71 42729 Z92771 GB_BA1：BAPURKE 2582 X91189 GB_PL1：SC130KBXV 129528 X94335 GB_PL1：SCXVORFS 50984 X90518 GB_PAT：E00311 1853 E00311 GB_PAT：106030 1853 106030 GB_PAT：100836 1853 100836 GB_BA2：CGU31230 3005 U31230 GB_HTG3：AC009946 169072 AC009946 GB_HTG3：AC009946 169072 AC009946 GB_BA1：MTCY253 41230 Z81368 GB_HTG4：AC010658 120754 AC010658 GB_HTG4：AC010658 120754 AC010658 GB_BA1：CGPROAGE 1783 X82929 N GB_BA1：MTCY428 26914 Z81451 GB_BA2：CGU31230 3005 U31230 GB_BA1：CGACEA 2427 X75504 GB_PAT：186191 2135 186191 GB_PAT：113693 2135 113693 GB_GSS15：AQ60684 574 AQ606842 2

B.ammoniagenes purK and purE genes. Mycobacterium tuberculosis H37Rv complete genome；segment 141/162. Mycobacterium tuberculosis H37Rv complete genome；segment 141/162. B.ammoniagenes purK and purE genes. S.cerevisiae 130kb DNA fragment from chromosome XV. S.cerevisiae DNA of 51 Kb from chromosome XV right arm. DNA coding of 2.5-diketogluconic acid reductase. Sequence 4 from Patent EP 0305608. Sequence 1 from Patent US 4758514. Corynebacterium glutamicum Obg protein homolog gene，partial cds，gamma glutamyl kinase(proB)gene，complete cds，and(unkdh)gene，complete cds. Homo sapiens clone NH0012C17，***SEQUENCING IN PROGRESS***，1 unordered pieces. Homo sapiens clone NH0012C17，***SEQUENCING IN PROGRESS***，1 unordered pieces. Mycobacterium tuberculosis H37Rv complete genome；segment 106/162. Drosophila melanogaster chromosome 3L/75C1 clone RPC198-3820，*** SEQUENCING IN PROGRESS***，78 unordered pieces. Drosophila melanogaster chromosome 3L/75C1 clone RPC198-3B20，*** SEQUENCING IN PROGRESS***，78 unordered pieces. C.glutamicum proA gene. Mycobacterium tuberculosis H37Rv complete genome；segment107/162. Corynebacterium glutamicum Obg protein homolog gene，partial cds，gamma glutamyl kinase(proB)gene，complete cds，and(unkdh)gene，complete cds. C.glutamicum aceA gene and thiX genes(partial). Sequence 3 from patent US 5700661. Sequence 3 from patent US 5439822. HS_5404_B2_E07_T7A RPC1-11 Human Male BAC library Homo sapiens genomic clone Plate＝980 Col＝14 Row＝J.genomic survey sequence.

Corynebacterium ammoniagenes Mycobacterium tuberculosis Mycobacterium tuberculosis Corynebacterium ammoniagenes Saccharomyces cerevisiae Saccharomyces cerevisiae unidentified Unknown. Unknown. Corynebacterium glutamicum Homo sapiens Homo sapiens Mycobacterium tuberculosis Drosophila melanogaster Drosophila melanogaster Corynebacterium glutamicum Mycobacterium tuberculosis Corynebacterium glutamicum Corynebacterium glutamicum Unknown. Unknown. Homo sapiens

61,731 39,624 39,847 64,286 36,617 36,617 56,123 56,220 56,220 99,332 36,115 36,115 38,088 35,817 35,817 98,802 38,054 98,529 100,000 100,000 100,000 39,716

14-Jan-97 10-Feb-99 10-Feb-99 14-Jan-97 15-Jul-97 1-Nov-95 29-Sep-97 02-DEC- 1994 21-MAY- 1993 2-Aug-96 8-Sep-99 8-Sep-99 17-Jun-98 16-OCT- 1999 16-OCT- 1999 23-Jan-97 17-Jun-98 2-Aug-96 9-Sep-94 10-Jun-98 26-Sep-95 10-Jun-99

Table 4 (continued)

rxa02458 1413 rxa02469 1554 rxa02497 1050 rxa02499 933 rxa02501 1188 rxa02503 522 rxa02504 681 rxa02516 1386

GB_EST1：T05804 406 T05804 GB_PL1：AB006699 77363 AB006699 GB_3A2：AF114233 1852 AF114233 GB_EST37：AW01306 578 AW013061 1 GB_GSS15：AQ65002 728 AQ650027 7 GB_BA1：MTCY359 36021 Z83859 GB_BA1：MLCB1788 39228 AL008609 GB_BA1：SCAJ10601 4692 AJ010601 GB_BA2：CGU31224 422 U31224 GB_BA1：MTCY20G9 37218 Z77162 GB_BA1：SCE7 16911 AL049819 GB_BA2：CGU31225 1817 U31225 GB_BA1：NG17PILA 1920 X13965 GB_HTG2：AC007984 129715 AC007984 GB_BA1：MTCY20G9 37218 Z77162 GB_BA1：U00018 42991 U00018 GB_VI：HE1CG 152261 X14112 GB_PR3：AC005328 35414 AC005328 GB_PR3：AC005545 43514 AC005545 GB_PR3：AC005328 35414 AC005328 GB_BA1：MTCY20G9 37218 Z77162 GB-PR3：AC005328 35414 AC005328 GB_PR3：AC005545 43514 AC005545 GB_BA1：MLCL536 36224 Z99125 GB_BA1：U00013 35881 U00013

EST03693 Fetal brain，Stratagene(cat#936206)Homo sapiens cDNA clonle HFBDG63 similar to EST containing Alu repeat.mRNA sequence. Arabidopsis thaliana genomic DNA，chromosome 5，P1 clone：MDJ22， complete sequence. Corynebacterium glutamicum 5-enolpyruvylshikimate 3-phosphate synthase aroA)gene，complete cds. ODT-0033 Winter flounder ovary Pleuronectes americanus cDNA clone ODT- 0033 5′similar to FRUCTOSE-BISPHOSPHATE ALDOLASE B(LVER). mRNA sequence. Sheared DNA-5L2，TF Sheared DNA Trypanosoma brucei genomic clone Sheared DNA-5L2，genomic survey sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 84/162. Mycobacterium leprae cosmid B1788. Streptomyces coelicolor A3(2)DNA for whiD and whiK loci. Corynebacterium glutamicum(ppx)gene，partial cds. Mycobacterium tuberculosis H37Rv complete genome；segment 25/162. Streptomyces coelicolor cosmid E7. Corynebacterium glutamicum L-proline：NADP+5-oxidoreductase(proC)gene complete cds. Neisseria gonorrhoeae pilA gene. Drosophila melanogaster chromosome 3 clone BACR05C10(D781)RPC1-98 05.C.10map 97D-97E strain y；cn bw sp，***SEQUENCING IN PROGRESS ***，87 unordered pieces. Mycobacterium tuberculosis H37Rv complete genome；segment 25/162. Mycobacterium leprae cosmid B2168. Herpes simplex virus(HSV)type 1 complete genome. Homo sapiens chromosome 19.cosmid R26660.complete sequence. Homo sapiens chromosome 19.cosmid R26634.complete sequence. Homo sapiens chromosome 19.cosmid R26660.complete sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 25/162. Homo sapiens chromosome 19.cosmid R26660.complete sequence. Homo sapiens chromosome 19.cosmid R26634.complete sequence. Mycobacterium leprae cosmid L536. Mycobacterium leprae cosmid B1496.

Homo sapiens Arabidopsis thaliana Corynebacterium glutamicum Pleurronectes americanus Trypanosoma brucei Mycobacterium tuberculosis Mycobacterium leprae Streptomyces coelicolor Corynebacterium glutamicum Mycobacterium luberculosis Streptomyces coelicolor Corynebacterium glutamicum Neisseria gonorrhoeae Drosophila melanogaster Mycobacterium tuberculosis Mycobacterium leprae human herpesvirus 1 Homo sapiens Homo sapiens Homo sapiens Mycobacterium tuberculosis Homo sapiens Homo sapiens Mycobacterium leprae Mycobacterium leprae

37,915 35,526 100,000 39,175 39,281 39,634 59,343 48,899 96,445 59,429 39,510 97,749 43,249 33,406 39,357 51,768 39,378 39,922 39,922 34,911 54,940 41,265 41,265 37,723 37,723

30-Jun-93 20-Nov-99 7-Feb-99 10-Sep-99 22-Jun-99 17-Jun-98 27-Aug-99 17-Sep-98 2-Aug-96 17-Jun-98 10-MAY- 1999 2-Aug-96 30-Sep-93 2-Aug-99 17-Jun-98 01-MAR- 1994 17-Apr-97 28-Jjl-98 3-Sep-98 28Jul-98 17Jun-98 28-Jul-98 3-Sep-98 04-DEC- 1998 01-MAR- 1994

Table 4 (continued)

rxa02517 570 rxa02532 1170 rxa02536 879 rxa02550 1434 rxa02559 1026 rxa02622 1683 rxa02623 714

GB_BA1：MTV007 32806 AL021184 GB_BA1：MLCL536 36224 Z99125 GB_BA1：U00013 35881 U00013 GB_BA1：SCC22 22115 AL096839 GB_OV：AF137219 831 AF137219 GB_EST30：A1645057 301 A1645057 GB_EST20：AA822595 429 AA822595 GB_HTG2：AF130866 118874 AF130866 GB_HTG2：AF130866 118874 AF130866 GB_PL1：ATT12J5 84499 AL035522 GB_BA1：MTCY279 9150 Z97991 GB_BA1：MSGB1970C 39399 L78815 S GB_BA2：SC2H4 25970 AL031514 GB_BA1：MTV004 69350 AL009198 GB_PAT：128684 5100 128684 GB_BA1：MTU27357 5100 U27357 GB_BA2：AE001780 11997 AE001780 GB_OV：AF064564 49254 AF064564 GB_OV：AF064564 49254 AF064564 GB_GSS5：AQ818728 444 AQ818728 GB_HTG5：AC011083 198586 AC011083 GB_GSS6：AQ826948 544 AQ826948

Mycobacterium tuberculosis H37Rv complete genome；segment 647162. Mycobacterium leprae cosmid L536. Mycobacterium leprae cosmid B1496. Streptomyces coelicolor cosmid C22. Amia calva mixed lineage leukemia-like protein(MII)gene，partial cds. vs52a10.y1 Stratagene mouse Tcell 937311 Mus musculus cDNA clone IMAGE：11498825′.mRNA sequence. vs52a10.r1 Stratagene mouse Tcell 937311 Mus musculus cDNA clone IMAGE：1149882 5′，mRNA sequence. Homo sapiens chromosome 8 clone PAC 172N13 map 8q24，*** SEQUENCING IN PROGRESS***，in unordere pipieces. Homo sapiens chromosome 8 clone PAC 172N13 map 8q24，*** SEQUENCING IN FROGRESS***，in unordered pieces. Arabidopsis thaliana DNA chromosome 4，BAC clone T12J5(ESSAII project) Mycobacterium tuberculosis H37Rv complete genome；segment 17/162. Mycobacterium leprae cosmid B1970 DNA sequence. Streptomyces coelicolor cosmid 2H4. Mycobacterium tuberculosis H37Rv complete genome；segment 144/162. Sequence 1 from patent US 5573915. Mycobacterium tuberculosis cyclopropane mycolic acid synthase(cma1) gene，complete cds. Thermotoga maritima section 92 of 136 of the complete genome. Fugu rubripes neurofibromatosis type 1(NF1)，A-kinase anchor protein (AKAP84).BAVV protein(BAVV)，and WSB1 protein(VVSB1)genes，complete cds. Fugu rubripes neurofibromatosis type 1(NF1).A-kinase anchor protein (AKAP84).BAVV protein(BAVV)，and VVSB1 protein(VVSB1)genes.complete cds. HS_5268_A1_G09 SP6E RPC1-11 Human Male BAC Library Homo sapiens genomic clone Plate＝844 Col＝17 Row＝M，genomic survey sequence. Homo sapiens chromosome 9 clone RP11-111M7 map 9，WORKING DRAFT SEQUENCE，51 unordered pieces. HS_5014_A2_C12_T7A RPC1-11 Human Male BAC Library Homo sapiens geaomic clone Plate＝590 Col＝24 Row＝E genomic survey sequence.

Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium leprae Streptomyces coelicolor Amia calva Mus musculus Mus musculus Homo sapiens Homo sapiens Arabidopsis thaliana Mycobacterium tuberculosis Mycobacterium leprae Streptomyces coelicolor A3(2) Mycobacterium tuberculosis Unknown. Mycobacterium tuberculosis Thermotoga maritima Fugu rubripes Fugu rubripes Homo sapiens Homo sapiens Homo sapiens

61,335 37,018 37,018 37,071 36,853 41,860 42,353 40,754 40,754 35,063 37,773 39,024 37,906 47,358 39,138 39,138 44,914 39,732 36,703 38,801 35,714 39,146

17-Jun-98 04-DEC- 1998 01-MAR- 1994 12-Jul-99 7-Sep-99 29-Apr-99 17-Feb-98 21-MAR- 1999 21-MAR- 1999 24-Feb-99 17-Jun-98 15-Jun-96 19-OCT- 1999 18-Jun-98 6-Feb-97 26-Sep-95 2-Jun-99 17-Aug-99 17-Aug-99 26-Aug-99 19-Nov-9g 27-Aug-99

Table 4 (continued)

rxa02629 708 rxa02645 1953 rxa02646 1392 rxa02648 1326 rxa02653 rxa02687 1068 rxa02717 1005 rxa02754 1461

GB_VI：BRSMGP 462 M86652 GB_VI：BRSMGP 462 M86652 GB_PAT：A45577 1925 A45577 GB_PAT：A45581 1925 A45581 GB_BA1：CORILVA 1925 L01508 GB_BA1：CORILVA 1925 LD1508 GB_PAT：A45585 1925 A45585 GB_PAT：A45583 1925 A45583 GB_OV：mCTCNC 2049 M83111 GB_EST11：AA265464 345 AA265464 GB_GSS8：AQ006950 480 AQ006950 GB_BA1：CORPHEA 1088 M13774 GBB_PAT：E04483 948 E04483 GB_PAT：E06110 948 E06110 GB_PL1：HVCH4H 59748 Y14573 GB_PR2：HS310H5 29718 Z69705 GB_PR3：AC004754 39188 AC004754 GB_HTG2：AC008223 130212 AC008223

Bovine respiratory syncytial virus membrane glyccprolein mRNA.complete cds. Bovine respiratory syncytial virus membrane glycoprotein mRNA.complete cds. Sequence 1 from Patent W09519442 Sequence 5 from Patent W09519442. Corynebacterium glutamicum threonine dehydratase(ilvA)gene，complete cds. Corynebacterium glutamicum threonine dehydratase(ilvA)gene，complete cds. Sequence 9 from Patent WO9519442. Sequence 7 from Patent WO9519442. lctalurus punctatus cyctic nucleotide-gated channel RNA sequence. mx91c06.r1 Soares mouse NML Mus musculus cDNA clone IMAGE：693706 5′，mRNA sequence. CIT-HSP-2294E14.TR CIT-HSP Homo sapiens genomic clone 2294E14. genomic survey sequence. C.glutamicum pheA gene encoding prephenate dehydratase，complete cds. DNA encoding prephenate dehydratase. DNA encodin9 prephenate dehydratase. Hordeum vulgare DNA for chromosome 4H. Human DNA sequence from cosmid 310H5 from a contig from the tip of the short arm of chromosome 16，spanning 2Mb of 16p13.3.Contains EST and CpG island. Homo sapiens chromosome 16，cosmid clone RT286(LANL)，complete sequence. Drosophila melanogaster chromosome 3 clone BACR16118(D815)RPC1-98 16.1.18 map 95A-95A strain y；cn bw sp，***SEQUENCING IN PROGRESS***，101 unordered pleces.

Bovine respiratory syncytia virus Bovine respiratory syncytia virus Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum Corynebacterium glutamicum lctalunus punctatus Mus musculus Homo sapiens Corynebacterium glutamicum Gorynebacterium glutamicum Corynebacterium glutamicum Hordeum vulgare Homo sapiens Homo sapiens Drosophila melanogaster

37,013 37,013 39,130 39,130 39,130 99,138 99,066 99,066 38,402 38,655 36,074 99,715 98,523 98,523 36,593 36,089 36,089 32,757

28-Apr-93 28-Apr-93 07-MAR- 1997 07-MAR- 1997 26-Apr-93 26-Apr-93 07-MAR- 1997 07-MAR- 1997 24-MAY- 1993 20-MAR- 1997 27-Jun-98 26-Apr-93 29-Sep-97 29-Sep-97 25-MAR- 1999 22-Nov-99 28-MAY- 1998 2-Aug-99

Table 4 (continued)

rxa302758 1422 rxa02771 678 rxa02772 1158 rxa02790 1266 rxa02791 951 rxa02802 1194

GB_HTG2：AC008223 130212 AC008223 GB_BA1：MTCY71 42729 Z92771 GB_HTG5：AC011678 171967 AC011678 GB_HTG5：AC011678 171967 AC011678 GB_BA2：AF064070 23183 AF064070 GB_BA2：AF038651 4077 AF038651 GB_IN1：CELT19B4 37121 U80438 GB_EST36：AV193572 360 AV193572 GB_BA2：AF038651 4077 AF038651 GB_BA1：MTCY227 35946 Z77724 GB_BA1：U00011 40429 U00011 GB_BA1：MTCY159 33818 Z83863 GB_PR4：AC006581 172931 AC006581 GB_PR4：AC006581 172931 AC006581 GB_BA1：MTCY159 33818 Z83863 GB_OV：CHKCEK2 3694 M35195 GB_BA1：MSASDASK 5037 Z17372 GB_EST24：A1223401 169 A1223401

Drosophila melanogaster chromosome 3 clone BACR 16118(D815)RPC1-98 16.1.18 map 95A-95A strain y；cnbw sp，***SEQUENCING IN PROGRESS ***，101 unordered pieces. Mycobacterium tuberculosis H37Rv complete genome；segment 141/162. Homo sapiens clone 14_B_7，***SEQUENCING IN PROGRESS***，20 unordered pieces. Homo sapiens clone 14_B_7，***SEQUENCING IN PROGRESS***，20 unordered pieces. Burkholderia pseudomallei putative dihydroorotase(pyrC)gene，partial cds； putative 1-acyl-sn-glycerol-3-phosphate acyltransferase(plsC)，putative diadenosine tetraphosphatase(apaH)，complete cds；type II O-antigen biosynthesis gene cluster，complete sequence；putative undecaprenyl phosphate N-acetyglucosaminyttransferrase，and putative UDP-glucose 4- epimerase genes，complete cds；and putative galactosyl transferase gene， partial cds. Corynebacterium glutamicum dipeptide-binding protein(dciAE)gene，partial cds：adenine phosphoribosyltransferase(apt)and GTP pyrophosphokinase (rel)genes.complete cds；and unknown gene. Caenorhabditis elegans cosmid T19B4. AV193572 Yuii Kohara unpublished cDNA：Strain N2 hermaphrodite embryo Caenorhabditis elegans cDNA clone yk618h85′，mRNA sequence. Corynebacterium glutamicum dipeptide-binding protein(dciAE)gene，partial cds；adenine phosphoribosyltransferase(apt)and GTP pyrophosphokinase (rel)genes，complete cds；and unknown gene. Mycobacterium tuberculosis H37Rv complete genome；segment 114/162. Mycobacterium leprae cosmid B1177. Mycobacterium tuberculosis H37Rv complete genome；segment 111/162. Homo sapiens 12p21 BAC RPCI11-259018(Roswell Park Cancer Institute Human BAC Library)complete sequence. Homo sapiens 12p21 BAC RPCI11-259018(Roswell Park Cancer Institute Human BAC Library)complete sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 111/162. Chicken tyrosine kinase(cek2)mRNA，complete cds. M.smegmatis asd，ask-alpha.and ask-beta genes. qg48g01.x1 Soares_testis_NHT Homo sapiens cDNA clone IMAGE：1838448 3′similar to WP：C25D7.8 CE08394；.mRNA sequence.

Drosophila melanogaster Mycobacte rium tuberculosis Homo sapiens Homo sapiens Burkholderia pseudomallei Corynebacterium glutamicum Caenorhabditis elegans Caenorhabditis elegans Corynebacterium glutamicum Mycobacterium tuberculosis Mycobacterium leprae Mycobacterium tuberculosis Homo sapiens Homo sapiens Mycobacterium tuberculosis Gallus gallus Mycobacterium smegmatis Homo sapiens

32,757 37,838 35,331 33,807 36,929 99,852 43,836 48,588 99,914 38,339 38,996 37,640 37,906 35,280 39,765 38,937 38,495 40,828

2-Aug-99 10-Feb-99 5-Nov-99 5-Nov-99 20-Jan-99 14-Sep-98 04-DEC- 1996 22-Jul-99 14-Sep-98 17-Jun-98 01-MAR- 1994 17-Jun-98 3-Jun-99 3-Jun-99 17-Jun-98 28-Apr-93 9-Aug-94 27-OCT- 1998

Table 4 (continued)

rxa02814 494 rxa02843 6D8 rxs03205 963 rxs03223 1237

GB_EST24：A1223401 169 A.1223401 GB_BA1：MTCY7D11 22070 Z95120 GB_BA1：MTCY7D11 22070 Z95120 GB_PR1：HSAJ2962 778 AJ002962 GB_BA1：CGAJ4934 1160 AJ004934 GB_BA1：MTC1364 29540 Z93777 GB_BA1：MLU15180 38675 U15180 GB_BA1：BLSIGBGN 2906 Z49824 GB_EST21：AA980237 377 AA980237 GB_EST23：A1158316 371 A1158316 G8_IN1：LMFL2743 38368 AL031910 GB_PR3：HSDJ61B2 119666 AL096710 GB_PR3：HSDJ6182 119666 AL096710

qg48g01.x1 Soares_testis_NHT Homo sapiens cDNA clone IMAGE：1838448 3′similar to WP：C25D7.8 CE08394：.mRNA sequence. Mycobacterium tuberculosis H37Rv complete genome；segment 138/162. Mycobacterium tuberculosis H37Rv complete genome；segment 138/162. Homo sapiens mRNA for hB-FABP. Corynebacterium glutamicum dapD gene，complete CDS. Mycobacterium tuberculosis H37Rv complete genome；segment 52/162. Mycobacterium leprae cosmid B1756. B.lactofermentum orf1 gene and sigB gene. ua32a12.r1 Soares_mammary_gland_NbMMG Mus musculus cDNA clone IMAGE：13484145′similar to TR：Q61025 Q61025 HYPOTHETICAL 15.2 KD PROTEIN.；.mRNA sequence. ud27c05.r1 Soares_thymus_2NbMT Mus musculus cDNA clone IMAGE：14471125′，mRNA sequence. Leishmania major Friedtin chromosome 4 cosmid L2743. Human DNA sequence from clone RP1-6182 on chromosome 6p 11.2-12.3 Contains isoforms 1 and 3 of BPAG1(bullous pemphigoid antigen 1 (230/240kD)，an exon of a gene similar to murine MACF cytoskeletal protein. STSs and GSSs.complete sequence. Human DNA sequence from clone RP1-6182 on chromosome 6p11.2-12.3 Contains isoforms1 and 3 of BPAG1(bullous pemphi9oid antigen 1 (230/240kD).an exon of agene similar to murine MACF cytoskeletal protein. STSs and GSSs，complete sequence.

Homo sapiens Mycobacterium tuberculosis Mycobacterium tuberculosis Homo sapiens Corynebacterium glutamicum Mycobacterium tuberculosis Mycobacterium leprae Corynebacterium glutamicu Mus musculus Mus musculus Leishmania major Homo sapiens Homo sapiens

40,828 58,418 40,496 39,826 100,000 37,710 39,626 88,854 41,489 38,005 39,869 34,930 34,634

27-OCT- 1998 17-Jun-98 17-Jun-98 8-Jan-98 17-Jun-98 17-Jun-98 09-MAR- 1995 25-Apr-96 27-MAY- 1998 30-Sep-98 15-DEC- 1999 17-DEC- 1999 17-DEC- 1999

Example

Example 1: Corynebacterium glutamicum ATCC 13032 genomic DNA preparation All

Corynebacterium glutamicum (ATCC 13032) was cultured in BHI medium (Difco) in 30 ℃ vigorous shaking overnight. The cells were collected by centrifugation, the supernatant discarded, cells were resuspended in 5ml Buffer I (original culture volume of 5% - all that is to 100ml volume of culture Volume calculation) in. Composition of Buffer I: 140.34g / l sucrose, 2.46g / l MgSO₄×7 H ₂O，10ml/l KH ₂PO ₄Solution (100g / l, KOH adjusted to PH6.7), 50g / l M12 Concentrate (10g / l (NH₄) ₂SO ₄，lg/l NaCl，2g/l MgSO ₄×7H ₂O，0.2g/l CaCl ₂0.5 g / l yeast extract (Difco)), 10ml / l trace elements mixture (200mg / l FeSO₄×H ₂O， 10mg/l ZnSO ₄×7H ₂O，3mg/l MnCl ₂×4H ₂O，30mg/l H ₃BO ₃，20mg/l CoCl ₂×6H ₂O，lmg/l NiCl ₂×6H ₂O，3mg/l Na ₂MoO ₄×2H ₂O), 500mg / l Complexing agent (EDTA or citrate), 100ml / l vitamin mixture (0.2mg / l biotin, 0.2mg / l folic acid, 20mg / l p-amino benzoic acid, 20mg / l riboflavin, 40mg / l panthothenate, 140mg / l nicotinic acid, 40mg / l hydrochloric acid, pyridoxal, 200mg / l myo-inositol). Hang Floating lysozyme was added to a final concentration of 2.5mg/ml. 37 ℃ for about four hours later, the cells Wall is degraded, and the resulting protoplasts were collected by centrifugation. Precipitate was washed with 5ml buffer I once, With 5ml TE buffer (10mM Tris-HCl, lml EDTA, pH8) wash. Precipitate was washed with 4 ml TE buffer, resuspended, and add 0.5ml SDS solution (10%) and 0.5ml NaCl solution (5M). Proteinase K added to a final concentration of 200μg/ml, suspension was incubated at 37 ℃ about 18 Hours. DNA with phenol, phenol - chloroform - isoamyl alcohol, chloroform - isoamyl alcohol extracted according to standard procedures Purification was then added 1/50 volume of 3M sodium acetate and 2 volumes of ethanol at -20 ℃ Incubate for 30 minutes, using the SS34 rotor (Sorvall) from a high-speed centrifuge 12,000 rpm Heart for 30 minutes, precipitated DNA. The DNA was dissolved in a 1ml TE containing 20μg/ml RNaseA Buffer in 1000ml TE buffer, dialyzed at 4 ℃ for at least 3 hours. During this time, Changing the buffer solution 3 times. Each dialyzed DNA solution 0.4ml, 0.4ml 2M LiCl and added 0.8ml of ethanol. At -20 ℃ for 30 min, centrifuged (13,000 rpm, Biofuge Fresco, Heraeus, Hanau, Germany) collect DNA. DNA pellet was thawed in TE buffer. DNA prepared according to the procedure used for all purposes, including the genomic southern hybridization and Library building. ...

O), 500mg / l Complexing agent (EDTA or citrate), 100ml / l vitamin mixture (0.2mg / l biotin, 0.2mg / l folic acid, 20mg / l p-amino benzoic acid, 20mg / l riboflavin, 40mg / l panthothenate, 140mg / l nicotinic acid, 40mg / l hydrochloric acid, pyridoxal, 200mg / l myo-inositol). Hang Floating lysozyme was added to a final concentration of 2.5mg/ml. 37 ℃ for about four hours later, the cells Wall is degraded, and the resulting protoplasts were collected by centrifugation. Precipitate was washed with 5ml buffer I once, With 5ml TE buffer (10mM Tris-HCl, lml EDTA, pH8) wash. Precipitate was washed with 4 ml TE buffer, resuspended, and add 0.5ml SDS solution (10%) and 0.5ml NaCl solution (5M). Proteinase K added to a final concentration of 200μg/ml, suspension was incubated at 37 ℃ about 18 Hours. DNA with phenol, phenol - chloroform - isoamyl alcohol, chloroform - isoamyl alcohol extracted according to standard procedures Purification was then added 1/50 volume of 3M sodium acetate and 2 volumes of ethanol at -20 ℃ Incubate for 30 minutes, using the SS34 rotor (Sorvall) from a high-speed centrifuge 12,000 rpm Heart for 30 minutes, precipitated DNA. The DNA was dissolved in a 1ml TE containing 20μg/ml RNaseA Buffer in 1000ml TE buffer, dialyzed at 4 ℃ for at least 3 hours. During this time, Changing the buffer solution 3 times. Each dialyzed DNA solution 0.4ml, 0.4ml 2M LiCl and added 0.8ml of ethanol. At -20 ℃ for 30 min, centrifuged (13,000 rpm, Biofuge Fresco, Heraeus, Hanau, Germany) collect DNA. DNA pellet was thawed in TE buffer. DNA prepared according to the procedure used for all purposes, including the genomic southern hybridization and Library building. ...

Use as in Example l, prepared as described in DNA, in accordance with the known and well established in Method (see, for example Sambrook, J.et al. (1989) "Molecular Cloning: A Laboratory Manual "Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, or Ausubel, FMet al. (1994) "Current Protocols in Molecular Bilogy ", John wiley & Sons.), Can be constructed cosmid library and plasmid library.

Use as in Example l, prepared as described in DNA, in accordance with the known and well established in Method (see, for example Sambrook, J.et al. (1989) "Molecular Cloning: A Laboratory Manual "Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, or Ausubel, FMet al. (1994) "Current Protocols in Molecular Bilogy ", John wiley & Sons.), Can be constructed cosmid library and plasmid library....

According to standard methods as in Example 2 using the genomic library as described, can be DNA sequencing, especially with the use of ABI377 chain termination sequencing method (see, for example, Fleischman, RDet al. (1995) "Whole-genome Random Sequencing and Assembly of Haemophilus Influenzae Rd., Science ,269:496-512). Use with Has the following nucleotide sequence sequencing primers: 5'-GGAAACAGTATGACCATG-3 'or 5'-GTAAAACGACGGCCAGT-3 '.

Example 4: In vivo mutagenesis

By E. coli or other microorganisms (e.g., bacteria or the like in some Bacillus The yeast Saccharomyces cerevisiae) plasmid (or other vector) DNA passaged for glutamic acid In vivo mutagenesis of Corynebacterium, wherein the microorganism to maintain the integrity of their genetic information capacity has been Damage. Typical mutant gene in the DNA repair system with mutations (e.g., mutHLS, mutD, mutT, etc.; references see Rupp, WD (1996) DNA repair mechanisms, in: Escherichia coli and Salmonella, p.2277-2294, ASM: Washington.). This These strains for skilled people is well known. The use of these strains are described in, for example, Greener, A.and Callahan, M. (1994) Strategies 7:32-34 in. ...

Corynebacterium and Brevibacterium species contain endogenous spontaneously replicated plasmid (such as for example, pHM1519 or pBL1) (review see, for example, Martin, JFet al. (1987) Biotechnology ,5:137-146). Escherichia coli and Corynebacterium glutamicum shuttle vector, can be E. coli using standard carriers easy to build (Sambrook, J.et al. (1989) "Molecular Cloning: A Laboratory Manual" Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press or Ausubel, FMet al. (1994) "Current Protocols in Molecular Bilogy", John Wiley & Sons.), I.e. in which the Corynebacterium glutamicum added replication fork and a suitable starting point mark. The origin of replication, excellent Is selected from Corynebacterium and Brevibacterium species isolated endogenous plasmid obtained. For these species turn The special-purpose markup kanamycin resistance gene (e.g., from Tn5 or Tn903 Those transposon kanamycin resistance gene) or chloramphenicol resistance gene (Winnacker, EL (1987) "From Genes to Clones-Introduction to Gene Technology, VCH, Weinheim). In building a variety of wild-type shuttle vector literature, there are many examples of these wear Shuttle vector in E. coli and Corynebacterium glutamicum copied, can be used for various purposes and , Including gene overexpression (see reference, for example, Yoshihama, M.et al. (1985) J.Bacteriol.162 :591-597, Martin JFet al. (1987) Biotechnology, 5: 137-146 and Eikmanns, BJet a;. (1991) Gene ,102:93-98). ...

Using standard methods of interest can be cloned into the shuttle vector, and can be With the introduction of the hybrid containing Corynebacterium glutamicum strains. Transformation of Corynebacterium glutamicum can be Protoplast transformation (Kastsumata, R.et al. (1984) J.Bacteriol.159306-311), electrical Chuan Kong (Lieb1, E.et al. (1989) FEMS Microbiol.Letters ,53:399-303) to achieve, When using a special carrier is used, can also be combined action (for example, in Sch_fer, A et al. (1990) J.Bacteriol.172 :1663-1666) implementation. Can also be prepared from Corynebacterium glutamicum plasmid DNA (using standard methods known in the art) and transformed into E. coli, and the wear Corynebacterium glutamicum shuttle vector from transformed into E. coli. This conversion step can use a standard square Method, but using Mcr-deficient E. coli strains, such as NM522 (Gough & Murray (1983) J.Mol.Biol.166 :1-19) is advantageous. ...

In addition to use of other plasmid replication can also be achieved through the integration of genomic gene over The amount of expression. Corynebacterium glutamicum or Brevibacterium or Corynebacterium other species genome integration, Known methods can be achieved, for example genomic regions homologous recombination, restriction endonuclease Enzyme mediated integration (REMI) (see, for example, DE Patent 19823834), or through the By using a transposon. Regulatory regions can also be modified (e.g., promoter, repressor, and / or Enhancer), through the use of targeted loci method (for example, homologous recombination) or based on random events Method (for example transposon mutagenesis or REMI) a sequence modified insert or deletion to adjust The activity of genes of interest. A nucleic acid as a transcription terminator sequences can be inserted into the present Description of one or more genes coding region 3 '; Such terminators are well known in the technology, And are described, for example, Winnacker, EL (1987) From Genes to Clones- Introduction to Gene Technology.VCH: Weinheim in. ...

Transforming a host cell by the activity of the mutant proteins observation relies on the fact that the projection Variable with the wild-type protein protein in a similar manner, and similar quantitative expression. Identified mutations base Due to the transcriptional level (for the gene product of translation of mRNA quantitative indicators) to be a useful way Method is a Northern blot (see reference, for example, Ausubel et al. (1988) Current Protocols in Molecular Biology, Wiley: New York), which designed for binding The primers of interest marked with a detectable tag (usually radioactive or chemiluminescent A), so that, when the culture of the organism is extracted total RNA, running gel electrophoresis, transferred To a stable medium and incubated with this probe, combined with the number of probe binding and it indicates that the gene presence and amount of mRNA. This information is mutated gene transcription level of evidence. You can use several Method was prepared from Corynebacterium glutamicum total cell RNA, which is technically known, for example, As described in Bormann, ERet al. (1992) Mol.Microbiol.6 :317-326 in. ...

Example 7: Genetic modification of the growth of Corynebacterium glutamicum - Media and culture conditions

Of genetically modified Corynebacterium glutamicum can be cultured in a synthetic or natural growth medium. For Corynebacterium glutamicum various growth medium are known and are readily available (Lieb, et al. (1989) Appl.Microbiol.Biotechno. ,32:205-210; von der Osten et al. (1998) Biotechnology Letters ,11:11-16; Patent DE 4,120,867; Liebl (1992) "The Genus Corynebacterium, in: Procaryotes, Volume II, Balows, A. et al., eds.Springer-Verlag). These media contain one or more of carbon, nitrogen, Inorganic salts, vitamins and trace elements. Preferred carbon sources are sugars, such as monosaccharides, disaccharides or Polysaccharides. For example, glucose, fructose, mannose, galactose, ribose, sorbose, ribulose, Lactose, maltose, sucrose, raffinose, starch or cellulose, can be used as a good source of carbon. Complex compound can also be provided to the medium sugars, refined sugars such as molasses, or other Byproducts. Improve the mixture of different carbon sources is also beneficial. Other available carbon sources of alcohol And organic acids such as methanol, ethanol, acetic acid or lactic acid. Typically an organic or inorganic nitrogen Nitrogen compounds, or compounds containing these substances. Representative of the nitrogen sources include ammonia or Ammonium salts such as NH ...₄Of genetically modified Corynebacterium glutamicum can be cultured in a synthetic or natural growth medium. For Corynebacterium glutamicum various growth medium are known and are readily available (Lieb, et al. (1989) Appl.Microbiol.Biotechno. ,32:205-210; von der Osten et al. (1998) Biotechnology Letters ,11:11-16; Patent DE 4,120,867; Liebl (1992) "The Genus Corynebacterium, in: Procaryotes, Volume II, Balows, A. et al., eds.Springer-Verlag). These media contain one or more of carbon, nitrogen, Inorganic salts, vitamins and trace elements. Preferred carbon sources are sugars, such as monosaccharides, disaccharides or Polysaccharides. For example, glucose, fructose, mannose, galactose, ribose, sorbose, ribulose, Lactose, maltose, sucrose, raffinose, starch or cellulose, can be used as a good source of carbon. Complex compound can also be provided to the medium sugars, refined sugars such as molasses, or other Byproducts. Improve the mixture of different carbon sources is also beneficial. Other available carbon sources of alcohol And organic acids such as methanol, ethanol, acetic acid or lactic acid. Typically an organic or inorganic nitrogen Nitrogen compounds, or compounds containing these substances. Representative of the nitrogen sources include ammonia or Ammonium salts such as NH ...₄) ₂SO ₄、NH ₄OH, nitrates, urea, amino acids or Complex nitrogen sources, such as corn steep liquor, soybean flour, soy protein, yeast extract, meat Extract or others.

Can be included in the medium inorganic compounds, including hydrochloride, phosphate or a sulfur Salts of calcium, magnesium, sodium, cobalt, molybdenum, potassium, manganese, zinc, copper or iron. Chelating agents may be added to Medium to maintain metal ions in solution. Particularly useful chelating agents include dihydroxybenzene Phenols, such as catechol and protocatechuate, or organic acids, such as citric acid. A typical medium Also contains growth factors, such as vitamins and growth promoters, examples of which include biotin, Riboflavin, thiamine, folic acid, niacin, pantothenate and pyridoxine alcohol. Growth factors and salts frequently from Complex medium components such as yeast extract, molasses, corn steep liquor, and other ingredients. The exact composition of the media compounds depends strongly on the immediate experiment and for each specific Case specific decision. Information about media optimization by the textbook "Applied Microbiol.Physiology, A Practical Approach "(eds.PMRhodes, PF Stanbury, IRL Press (1997) pp.53-73, ISBN 0 19 963577 3) "in can also be from Commercial vendors selective growth medium, such as standard 1 (Merck) or BHI (Grain heart infusion, DIFCO) or others. ...

Culture conditions were determined for each experiment. Temperature should be at 15 ℃ to 45 ℃ range. The temperature can be kept constant, or changes in the experiment. PH of the medium in the range of 5 to 8.5, Preferably at about 7.0, and can be added medium to maintain the buffer. For this Object of a representative buffer is potassium phosphate buffer. Synthetic buffers such as MOPS, HEPES, ACES and others, can also be used instead or in combination. You can also Growth process by adding NaOH or NH₄OH% EF% BC% 8C% E4% BB% A5% E7% BB% B4% E6% 8C% 81% E7% A8% B3% E5% AE% 9A% E7% 9A% 84% E5% 9F% B9% E5% 85% BBpH% E3% 80% 82% E5% A6% 82% E6% 9E% 9C% 0A% 20% 20% 20% 20% 20% 20% 20% 20% 20% 20% 20% 20% E4% BD% BF% E7% 94% A8% E5% 83% 8F% E6% 98% AF% E9% 85% B5% E6% AF% 8D% E6% 8F% 90% E5% 8F% 96% E7% 89% A9% E8% BF% 99% E6% A0% B7% E7% 9A% 84% E5% A4% 8D% E6% 9D% 82% E5% 9F% B9% E5% 85% BB% E5% 9F% BA% E7% BB% 84% E5% 88% 86% EF% BC% 8C% E5% 8F% AF% E4% BB% A5% E5% 87% 8F% E5% B0% 91% E6% B7% BB% E5% 8A% A0% E7% BC% 93% E5% 86% B2% E6% B6% B2% E7% 9A% 84% E5% BF% 85% 0A% 20% 20% 20% 20% 20% 20% 20% 20% 20% 20% 20% 20% E8% A6% 81% E6% 80% A7% EF% BC% 8C% E8% BF% 99% E6% 98% AF% E5% 9B% A0% E4% B8% BA% E8% AE% B8% E5% A4% 9A% E5% A4% 8D% E6% 9D% 82% E5% 8C% 96% E5% 90% 88% E7% 89% A9% E5% 85% B7% E6% 9C% 89% E5% BE% 88% E5% BC% BA% E7% 9A% 84% E7% BC% 93% E5% 86% B2% E8% 83% BD% E5% 8A% 9B% E8% BF% 99% E4% B8% 80% E4% BA% 8B% E5% AE% 9E% E3% 80% 82% E5% A6% 82% E6% 9E% 9C% E4% BD% BF% 0A% 20% 20% 20% 20% 20% 20% 20% 20% 20% 20% 20% 20% E7% 94% A8% E5% 8F% 91% E9% 85% B5% E7% BD% 90% E5% 9F% B9% E5% 85% BB% E5% BE% AE% E7% 94% 9F% E7% 89% A9% EF% BC% 8C% E4% B9% 9F% E5% 8F% AF% E4% BB% A5% E4% BD% BF% E7% 94% A8% E6% B0% A8% E6% B0% 94% E6% 8E% A7% E5% 88% B6pH% E3% 80% 82

Incubation time is usually in the range of hours to days. This time is to allow the selection of Accumulate in the liquid medium the maximum amount of product. However, growth experiments published in a variety of containers , For example microtiter plates, glass tubes, glass flasks or glass or a different size Metal fermenter. In order to screen large numbers of clones should microorganism cultured in a baffle or not A baffle microtiter plates, glass tubes or shake flasks. Preferably use 100ml flask, 10% (volume) of the desired medium. Should be placed on a shaker flask shaking (amplitude 25 mm M), the speed range of 100-300rpm. Can be kept moist air to reduce evaporation losses; Or, mathematically corrected for evaporation losses. ...

Incubation time is usually in the range of hours to days. This time is to allow the selection of Accumulate in the liquid medium the maximum amount of product. However, growth experiments published in a variety of containers , For example microtiter plates, glass tubes, glass flasks or glass or a different size Metal fermenter. In order to screen large numbers of clones should microorganism cultured in a baffle or not A baffle microtiter plates, glass tubes or shake flasks. Preferably use 100ml flask, 10% (volume) of the desired medium. Should be placed on a shaker flask shaking (amplitude 25 mm M), the speed range of 100-300rpm. Can be kept moist air to reduce evaporation losses; Or, mathematically corrected for evaporation losses. ...₆₀₀Value of 0.5-1.5 inoculated medium can pass By introducing the plate from the CM salt Corynebacterium glutamicum cell suspension, or by adding to the Bacterial liquid preculture implementation.

Example 8: In vitro analysis of protein function mutations

Kinetic parameters of the enzyme activity and determination of technically been well established for. Any After a change of a given enzyme activity measurement experiment, must be suitable for the specific activity of wild type enzyme Sex, it is entirely in the ability of skilled in the art within. Summary of comments on the enzyme and on the knot Structure, dynamics, principles, methods, applications and determine the number of instances of explicit details of the activity can be In the example found in the following references: Dixon, M., and Webb, EC, (1979) Enzymes. Longmans: London; Fersht, (1985) Enzyme Structure and Mechanism.Freeman: New York; Walsh, (1979) Enzymatic Reaction Mechanisms.Freeman: San Francisco; Price, NC, Stevens, L. (1982) Fundamentals of Enzymology.Oxford Univ.Press: Oxford; Boyer, P.D., ed. (1983) The Enzymes, 3 ...^rd ed.Academic Press：New York； Bisswanger，H.，(1994)Enzymkinetik，2 ^nd ed.VCH：Weinheim(ISBN 3527300325)； Bergmeyer，H.U.，Bergmeyer，J.，Graβ1，M.，eds.(1983-1986)Methods of Enzymatic Analysis，3 ^rd ed.，vol.I-XII，Verlag Chemie：Weinheim；and Ullmann’s Encyclopedia of Industrial Chemistry(1987)vol.A9，“Enzymes”.VCH：Weinheim，p.352- 363。

Kinetic parameters of the enzyme activity and determination of technically been well established for. Any After a change of a given enzyme activity measurement experiment, must be suitable for the specific activity of wild type enzyme Sex, it is entirely in the ability of skilled in the art within. Summary of comments on the enzyme and on the knot Structure, dynamics, principles, methods, applications and determine the number of instances of explicit details of the activity can be In the example found in the following references: Dixon, M., and Webb, EC, (1979) Enzymes. Longmans: London; Fersht, (1985) Enzyme Structure and Mechanism.Freeman: New York; Walsh, (1979) Enzymatic Reaction Mechanisms.Freeman: San Francisco; Price, NC, Stevens, L. (1982) Fundamentals of Enzymology.Oxford Univ.Press: Oxford; Boyer, P.D., ed. (1983) The Enzymes, 3 ...

Membrane transport protein activity can be determined such as those described in Gennis, RB (1989) "Pores, Channels and Transporters", in Biomembrane, Molecular Structure and Function, Springer: Heidelberg, p.85-137; 199-234; and 270-322 in white that These techniques.

Example 9: production of mutant proteins to effect the desired product analysis

Genetically modified Corynebacterium glutamicum for the desired compound (such as amino acids) produced role Can be estimated, that is, through the appropriate conditions (for example, those described above) has been modified growth Microorganisms, and the increase of the desired product (e.g., amino acids) the production of medium and / or Cell components. These analytical techniques for the skilled person is well known conventional techniques, including spectral Analysis, thin layer chromatography, various staining methods, enzymatic methods and microbiological methods, and such high Performance liquid chromatography (Ullman, Encyclopedia of Industrial Chemistry, vol.A2, p.89-90 and p.443-613, VCH: Weinheim (1985); Fallon, A.et al., (1987) "Applications of HPLC in Biochemistry "in: Laboratory Techniques in Biochemistry and Molecular Biology, vol.17; Rehm et al. (1993) Biotechnology, vol.3, Chapter III: "Product recovery and purification ", page 469-714, VCH: Weinheim; Belter, PAet al. (1988) Bioseparations: downstream processing for biotechnology, John Wiley and Sons; Kennedy, JFand Cabral, JMS (1992) Recovery processes for biological materials, John Wiley and Sons; Shaeiwitz, JAand Henry, JD (1988) Biochemical separations, in: Ulmann's Encyclopedia of Industrial Chemistry, vol.B3, Chapter 11, page 1-27, VCH: Weinheim; and Dechow, FJ (1989) Separation and purification techniques in biotechnology, Noyes Publications) chromatography such analysis. ...

In addition to the determination of the final fermentation products, the desired compound can also be used for the production of metabolic Other components pathway analysis, such as intermediates and by-products, to determine the total raw compound Production efficiency. Analysis methods include nutrient levels in the medium (for example, sugars, hydrocarbons, nitrogen, Phosphate and other ions) determination of composition and growth of the biomass measurement biosynthetic pathway Common analysis of the production of metabolites, as well as the determination of the gas produced by fermentation. These assays The standard method outlined in Applied Microbial Physiology, A Practical Approach, P. M.Rhodes and PFStanbury, eds., IRL Press, p.103-163; and 165-192 (ISBN: 0199635773) and references cited. ...

Corynebacterium glutamicum cells from the culture supernatant or the recovery of the desired product, can be Through a variety of methods known in the art. If the desired product is not secreted by cells, it It can be from the medium by low speed centrifugation cells were collected, lysed by standard techniques, such as Mechanical force or ultrasound. Cell debris was removed by centrifugation, retaining the supernatant containing the soluble protein For further purification the desired compound. If the product is secreted from the cells of Corynebacterium glutamicum, Then use the low-speed centrifugation to remove cells from the culture medium, supernatant retained for further purification.

Corynebacterium glutamicum cells from the culture supernatant or the recovery of the desired product, can be Through a variety of methods known in the art. If the desired product is not secreted by cells, it It can be from the medium by low speed centrifugation cells were collected, lysed by standard techniques, such as Mechanical force or ultrasound. Cell debris was removed by centrifugation, retaining the supernatant containing the soluble protein For further purification the desired compound. If the product is secreted from the cells of Corynebacterium glutamicum, Then use the low-speed centrifugation to remove cells from the culture medium, supernatant retained for further purification....

Purification methods known in the art is very large, the above purification method does not mean just Bureau Limited thereto. These purification methods are described in, for example, Bailey, JE & Ollis, DFBiochmical Engineering Fundamentals, McGraw-Hill: New York (1986) in.

Purification methods known in the art is very large, the above purification method does not mean just Bureau Limited thereto. These purification methods are described in, for example, Bailey, JE & Ollis, DFBiochmical Engineering Fundamentals, McGraw-Hill: New York (1986) in....

Sequence comparison between two sequences and determination of percent homology, are known in the technical Surgery, you can use mathematical algorithms to complete, for example Karlin and Altschul (1990) Proc. Natl.Acad.Sci.USA 87:2264-68 in algorithms, the algorithm in Karlin and Altschul (1993) Proc.Natl.Acad Sci.USA 90:5873-77 has changed. The op- Law is integrated in Altschul, et al. (1990) J Mol.Biol.215 :403-10 in NBLAST and XBLAST program (version 2.0) in. BLAST nucleotide search can NBLAST Program, score = 100, wordlength = 12, can be obtained with the present invention, the nucleic acid molecule MP Homologous nucleotide sequences. Protein BLAST search program can XBLAST, score = 50, wordlength = 3, can be obtained with the present invention, homologous protein amino MP Acid sequence. For comparison purposes, in order to obtain a gap sequence alignment, can be used to describe In Altschul et al., (1997) Nucleic Acids Res.25 (17) :3389-3402 the Gapped BLAST. When using BLAST and Gapped BLAST programs, the person skilled in the art that For a specific sequence to be analyzed how to optimize the program (for example, XBLAST and NBLAST) of Parameters. ...

Percentage between two amino acid sequences homologous to the GCG software package can be used (http://www.gcg.com is provided) in the GAP program, use Blosum 62 matrix Or PAM 250 matrix, gap component 12,10,8,6 or 4, the length of component 2, 3 or In 4. Percentage between two nucleic acid sequence homology can be used in the package GCG GAP Program, using the standard parameters such as the length of the gap component 50 and component 3.

Gene sequences of the invention between the sequences in Genbank comparative analysis techniques can be used Techniques known in the art (see, for example, Bexevanis and Ouellette, eds. (1998) Bioinformatics: A Practical Guide to the Analysis of Genes and Proteins. John Wiley and Sons: New York). Gene sequences of the present invention, three steps of the square Method and compare sequences in Genbank. In the first step of the present invention, each of a sequence of Relative to the nucleotide sequence in Genbank BLASTN analysis (eg, local sequence alignment Analysis), to retain the highest matching 500 for further analysis. Then this 500 matches for FASTA search (for example, a combination of local and worldwide comparative sequence analysis, in which the limited Sequence region sequence comparison). Next, each of the present invention, and FASTA sequences The three highest matching, using the GAP program in the GCG software package (using standard parameters) into the OK Golden World sequence comparison. To get the correct result, the length from the Genbank sequence selected, Technically well known methods using the query sequence length adjustment. The results of the analysis are listed in Table 4. Medium. Although this result, each gene of the present invention with respect to each of Genbank Conducted according to a single GAP (worldwide) analysis of the result is the same, but the relative The large database GAP (worldwide) analysis, the required computing time is greatly reduced. Not Cutoff values obtained over a sequence alignment of the sequence of the present invention, in Table 4 show that the lack of sequence Ratio information. Skilled in the art to understand the deeper, listed in Table 4, the heading "% homology (GPA) "under the GAP percentage sequence homology comparison is based on the European digital format Listed, of which ',' on behalf of the decimal point. For example, the value in the column "40,345" represents "40.345 percent." ...

Sequence of the invention can also be used for DNA microarray (DNA array design, methods, and The application of technology are well known and described, for example, Schena, M.te al. (1995) Science 270: 467-470; Wodicka, L.et al. (1997) Nature Biotechnology 15:1359-1367; DeSaizieu, A.et al. (1998) Nature Biotechnology 16:45-48; and DeRisi, JL et al. (1997) Science 278:680-686) Construction and application.

DNA microarrays using a solid or flexible supports, including nitrocellulose, nylon Dragon, glass, silicon or other materials. Nucleic acid molecules can be connected in an orderly manner on the surface. Suitable marker, the other nucleic acid or nucleic acid mixture can be hybridized with the immobilized nucleic acid molecules, Tag can be used to monitor and measure the area of hybrid molecules determine separate signal strength. This method Allows the simultaneous quantification of the nucleic acid sample, or for all or a mixture of nucleic acids with the selected Right or absolute number. Thus, DNA micro-array allows multiple (up to 6800 or more) Similar analysis of a nucleic acid expression (see, e.g., Schena, M. (1996) BioEssays 18 (5): 427-431). ...

A nucleic acid microarray can also be as in the Wodicka, L.et al. (1997) Nature Biotechnology 15:1359-1367 describes oligonucleotides synthesized in situ building. Through photos Phase plate method, the matrix can be accurately determined area exposed to light. The protecting group is light Unstable, and thus be activated and subjected to nucleotide addition, but disguised and seeing the light district Domain without any modification. Protection and activation of the next light cycle, allowing to determine the location of different Oligonucleotide synthesis. The present invention can be determined by a small region on the microarray by solid-phase oligo Nucleotide synthesis and synthesis.

Nucleotides in the sample or a mixture of the nucleic acid molecule of the present invention can be used with microarray Hybridization. You can mark them according to the standard method of nucleic acid molecules. Simply put, the nucleic acid molecule (case If, mRNA molecules or DNA molecule) can be labeled with the isotope or a fluorescent nucleoside Acid binding is marked, for example, reverse transcriptase or DNA synthesis. Labeled nucleic acid microarray Hybridization is described (for example, in Schena, M.et al. (1995) supra; Wodicka, L.et al. (1997), supra; and DeSaizieu A.et al. (1998), supra IN). Hybrid molecule detection To fit a specific and quantitative binding tag. Radioactive label can be detected, for example, Schena, M.et al. (1995) supra described, but also can detect fluorescence, for example, Shalon et al. (1996) Gemone Research 6:639-645 approach. ...

Example 13: Analysis of cell population dynamics of proteins (proteomics)

Gene of the invention, the composition and method can be used to study protein interactions groups And dynamics, called "proteomics." Protein of interest groups, including, but not Bureau Limited to, all of Corynebacterium glutamicum protein groups (eg, proteins, and other organisms Comparison group), in a special environment or metabolic conditions (e.g., fermentation in temperature or By low temperature or high pH or low pH) that are active protein, or a specific growth or Developmental stages by those proteins are active.

You can use a variety of well-known technical analysis of protein groups, such as gel electrophoresis. Cell protein Quality can be obtained such as cracking or extract can also be separated from each other using a variety of electrophoretic techniques Away. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) separation of proteins, it is A large extent based on their molecular weight. Polyacrylamide gel isoelectric focusing electrophoresis (IEF- PAGE) through such little (which reflects not only the amino acid sequence, and screening of protein translation Translation modification) separation of proteins. A more preferred method of protein analysis, IEF-PAGE And SDS-PAGE with a continuous, known as 2-D-gel electrophoresis (for example, in Hermann et al. (1998) Electrophoresis 19:3217-3221; Fountoulakis et al. (1998) Electrophoresis 19:1193-1202; Langen et al. (1997) Electrophoresis 18: 1184-1192; Antelmann et al. (1997) Electrophoresis18 :1451-1463 described in Above). ...

You can use a variety of well-known technical analysis of protein groups, such as gel electrophoresis. Cell protein Quality can be obtained such as cracking or extract can also be separated from each other using a variety of electrophoretic techniques Away. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) separation of proteins, it is A large extent based on their molecular weight. Polyacrylamide gel isoelectric focusing electrophoresis (IEF- PAGE) through such little (which reflects not only the amino acid sequence, and screening of protein translation Translation modification) separation of proteins. A more preferred method of protein analysis, IEF-PAGE And SDS-PAGE with a continuous, known as 2-D-gel electrophoresis (for example, in Hermann et al. (1998) Electrophoresis 19:3217-3221; Fountoulakis et al. (1998) Electrophoresis 19:1193-1202; Langen et al. (1997) Electrophoresis 18: 1184-1192; Antelmann et al. (1997) Electrophoresis18 :1451-1463 described in Above). ...³⁵S-methionine,³⁵S- Cysteine,¹⁴C-labeled amino acids,¹⁵N-amino acid,¹⁵NO ₃Or¹⁵NH ₄ ⁺Or¹³C-labeled Mind amino acids), can make these cells before the separation of protein labeled protein. Category Like, can also be a fluorescent tag. According to the aforementioned technique can be extracted, isolated and separate these Labeled protein.

Proteins using these techniques show, by measuring a dye or a mark used as a feed Further analysis. The number of specific proteins may be used such as an optical method for quantitative determination, And may be in the same or another on a gel on the gel ratio of the number of other proteins Compared. By, for example optical comparator, spectroscopic analysis, gel image analysis and scanning, or through the By using photographic film or the display of proteins on the gel for comparison. These technologies Technically known.

To determine the characteristics of a specific protein can be determined using direct sequence or other criteria Technologies. For example, you can use the N-and / or C-terminal amino acid sequencing (e.g., Edman degradation), And mass spectrometry (MALDI or ESI particular techniques (see, for example, Langen et al. (1997) Electrophoresis 18:1184-1192)). Protein sequences provided herein may be These techniques for Corynebacterium glutamicum protein identification.

The information obtained by these techniques can be used to compare the protein present, activity, different Biological conditions (for example, other conditions in the different organisms, fermentation time points, medium Conditions, or biological environment) modification between different samples of the various modes. The resulting number of these tests It can be used alone, or combined with other technologies used in various applications, such as comparing special Specified circumstances (such as metabolism) various organisms act to increase the production of fine chemicals Strains production capacity, or to increase the efficiency of fine chemical production.

Equivalent statement

Skilled in the art will recognize, or be able to determine using only routine experimentation, this Described at a particular embodiment of the present invention, there are many equivalents. The following claims are intended to package Containing such equivalents.

Claims

1 Corynebacterium glutamicum isolated nucleic acid molecule which contains selected from SEQ ID NO: 3 - 1155 with an odd sequence number for each listed nucleic acid sequence or complementary sequence, provided that the nucleic acid Molecules do not include any of those listed in Table 1 F-indicated genes.

2 An isolated nucleic acid molecule coding for the molecule is selected from SEQ ID NO :4-1156 per One with an even number of amino acid sequence listed in SEQ ID NO or its complementary sequence, provided that the nucleic acid molecule does not Including any of those listed in Table 1 F-indicated genes.

3 encoding a polypeptide isolated naturally occurring allelic variant of a nucleic acid molecule, the polypeptide package Selected from the group containing SEQ ID NO :4-1156 with an even sequence number for each listed amino acid sequence or Complementary sequences, provided the nucleic acid molecule does not include any of those listed in Table 1 F-indicated genes.

4 An isolated nucleic acid molecule comprising a nucleotide sequence, the sequence of SEQ ID NO: 3 - 1155 either an odd sequence number are listed in the nucleotide sequence of the complete or complementary sequences thereof with at least Identity of 50%, provided that the nucleic acid molecule does not include any of those listed in Table 1 F-indicated base Result.

5 isolated comprising SEQ ID NO :3-1155 in any one of SEQ ID number listed nuclear Or a nucleotide sequence complementary to a sequence of at least 15 consecutive nucleotides of a nucleic acid molecule fragments, the conditions This nucleic acid molecule does not include any of those listed in Table 1 F-indicated genes.

6 An isolated nucleic acid molecule comprises the nucleic acid molecule of any one of claims 1-5 in the nucleic acid Molecule encoding a heterologous polypeptide and nucleotide sequences.

7 contains any one of claims 1-6 in a vector nucleic acid molecule.

8 The vector of claim 7, the vector is an expression vector.

9 expression vector according to claim 8 transfected host cells.

10 The host cell according to claim 9, wherein the cell is a microorganism.

11 The host cell according to claim 10, wherein the cell belongs to the genus Corynebacterium or Brevibacterium Genera.

12 The host cell according to claim 9, wherein the expression of said nucleic acid molecule, resulting in the fine Cells regulate the production of fine chemicals.

13 The host cell of claim 12, wherein said fine chemical is selected from the following physical Quality: organic acids, protein source of amino acids, non-protein source of amino acids, purine and pyrimidine bases Group, nucleosides, nucleotides, lipids, saturated and unsaturated fatty acids, diols, carbohydrates, aromatic Compounds, vitamins, cofactors, polyketides, and enzymes.

14 for producing a polypeptide, comprising culturing in a suitable host according to claim 9 Cells, thereby producing the polypeptide.

15 containing the selected separation SEQ ID NO :4-1156 with an even sequence number for each listed Acid sequence of a polypeptide, provided that the amino acid sequence listed in Table 1, not by any The gene coding for F-marked.

16 containing the selected separation SEQ ID NO :4-1156 with an even sequence number for each listed Acid sequence of the polypeptide allelic variants of naturally occurring polypeptide, the condition that the ammonia Amino acid sequence listed in Table 1, not by any of the gene coding for F-marked.

17 An isolated polypeptide encoded by a nucleic acid molecule, the nucleic acid molecule comprises a nucleotide sequence SEQ ID NO :3-1155 sequence of any one of an odd number of the nucleic acid sequences listed in full at least Identity of 50%, provided that the nucleic acid molecule does not include any of those listed in Table 1 F-marked nuclear Acid molecules.

18 An isolated polypeptide, the polypeptide comprises the amino acid sequence of SEQ ID NO :4-1156 of Either with an even sequence number are listed in full at least 50% amino acid sequence identity, condition The amino acid sequence listed in Table 1, not by any of the gene coding for F-marked.

19 An isolated polypeptide comprising the polypeptide comprises SEQ ID NO :4-1156 either an even The amino acid sequence listed in SEQ ID polypeptide fragments not by the condition that the amino acid sequence in Table 1 Any listed in the gene coding for F-marked, wherein said polypeptide fragment comprising the amino acid sequence to maintain The biological activity of the polypeptide column.

20. To any one of claims 15 to 19 in isolated polypeptide further comprises a heterologous Amino acid sequence.

21 The method of production of fine chemicals, comprising culturing a cell according to claim 9, thereby generating Health fine chemicals.

22 The method of claim 21, wherein said method further comprising the culture from said In the step of recovery of fine chemicals.

23 The method of claim 21, wherein said cell belongs to the genus Corynebacterium or Brevibacterium Genera.

24. The method of claim 21, wherein said cell is selected from the following strains: Corynebacterium glutamicum Bacteria, Corynebacterium Lux, lily Corynebacterium, Corynebacterium acetoacidophilum Corynebacterium, Corynebacterium vinegar Valley, Addicted acetyl Corynebacterium, Corynebacterium ammoniagenes, Corynebacterium fujiokense, Corynebacterium nitrilophilus, Brevibacterium ammoniagenes, Brevibacterium butanicum, differences Brevibacterium, yellow Brevibacterium, the His Brevibacterium ketoglutarate Brevibacterium, Brevibacterium ketosoreductum, Brevibacterium lactofermentum, Brevibacterium, Brevibacterium wax solution and strains listed in Table 3.

25. The method of claim 21, wherein said vector expressing the nucleic acid molecule, resulting in the Cells regulate the production of fine chemicals.

26 The method of claim 21, wherein said fine chemical substance selected from the following materials: Organic acids, non-protein source of amino acids, purine and pyrimidine bases, nucleosides, nucleotides, fatty Quality, saturated and unsaturated fatty acids, diols, carbohydrates, aromatic compounds, vitamins, cofactors Son, polyketide compounds and enzymes.

27 The method of claim 21, wherein said fine chemical is an amino acid.

28. The method of claim 27, wherein said amino acid is selected from the following amino acid: lysine Acid, glutamic acid, glutamine, alanine, aspartic acid, glycine, serine, threonine, Methionine, cysteine, valine, leucine, isoleucine, arginine, proline, Histidine, tyrosine, phenylalanine, tryptophan.

29 The method of production of fine chemicals, comprising culturing such cells, the cellular genes Genomic DNA was inserted into any one of claims 1-6 in which the nucleic acid molecule changed.

30. Diagnosis of Corynebacterium diphtheria in subjects presence or activity, the method comprising detecting the test By the nucleic acid molecule of claims 1-5 or a polypeptide of claims 15 to 19 in the molecule at least one of The presence of substances, to diagnose subjects Corynebacterium diphtheria presence or activity.

31. Least one selected from SEQ ID NO :3-1155 with an odd sequence number for each listed nucleic acid The sequence of the host cell nucleic acid molecule, wherein the nucleic acid molecules are destroyed.

32 selected from the group comprising SEQ ID NO :3-1155 with an odd sequence number for each listed nucleic acid The sequence of the host cell nucleic acid molecule, wherein the nucleic acid molecule contains one or more of SEQ ID NO :3-1155 sequence of any one of an odd number of the nucleic acid sequence listed as compared to a nucleic acid modification.

33 selected from the group comprising SEQ ID NO :3-1155 with an odd sequence number for each listed nucleic acid The sequence of the host cell nucleic acid molecule, wherein the nucleic acid molecule regulating region of the molecule with respect to the wild Wild-type regulatory regions were modified.