CN114990147A - Method for constructing recombinant vibrio natriegens for producing geraniol, recombinant vibrio natriegens constructed by method and application of recombinant vibrio natriegens - Google Patents
Method for constructing recombinant vibrio natriegens for producing geraniol, recombinant vibrio natriegens constructed by method and application of recombinant vibrio natriegens Download PDFInfo
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- CN114990147A CN114990147A CN202110226161.2A CN202110226161A CN114990147A CN 114990147 A CN114990147 A CN 114990147A CN 202110226161 A CN202110226161 A CN 202110226161A CN 114990147 A CN114990147 A CN 114990147A
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- GLZPCOQZEFWAFX-UHFFFAOYSA-N Geraniol Chemical compound CC(C)=CCCC(C)=CCO GLZPCOQZEFWAFX-UHFFFAOYSA-N 0.000 title claims abstract description 133
- GLZPCOQZEFWAFX-YFHOEESVSA-N Geraniol Natural products CC(C)=CCC\C(C)=C/CO GLZPCOQZEFWAFX-YFHOEESVSA-N 0.000 title claims abstract description 67
- 239000005792 Geraniol Substances 0.000 title claims abstract description 67
- 229940113087 geraniol Drugs 0.000 title claims abstract description 67
- 241000607365 Vibrio natriegens Species 0.000 title claims abstract description 43
- 238000000034 method Methods 0.000 title claims abstract description 34
- 238000000855 fermentation Methods 0.000 claims abstract description 8
- 230000004151 fermentation Effects 0.000 claims abstract description 8
- 239000013612 plasmid Substances 0.000 claims description 27
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- 238000004519 manufacturing process Methods 0.000 claims description 11
- 239000002243 precursor Substances 0.000 claims description 11
- 239000013598 vector Substances 0.000 claims description 11
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 8
- 241000894006 Bacteria Species 0.000 claims description 7
- 239000001888 Peptone Substances 0.000 claims description 4
- 108010080698 Peptones Proteins 0.000 claims description 4
- 229940041514 candida albicans extract Drugs 0.000 claims description 4
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- 239000013600 plasmid vector Substances 0.000 claims description 3
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- 239000007864 aqueous solution Substances 0.000 claims description 2
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- 241000588724 Escherichia coli Species 0.000 abstract description 9
- 239000001963 growth medium Substances 0.000 abstract description 9
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- BPHPUYQFMNQIOC-NXRLNHOXSA-N isopropyl beta-D-thiogalactopyranoside Chemical compound CC(C)S[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O BPHPUYQFMNQIOC-NXRLNHOXSA-N 0.000 abstract description 4
- KJTLQQUUPVSXIM-ZCFIWIBFSA-N (R)-mevalonic acid Chemical compound OCC[C@](O)(C)CC(O)=O KJTLQQUUPVSXIM-ZCFIWIBFSA-N 0.000 description 4
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- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
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- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 2
- 240000002234 Allium sativum Species 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 2
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- 235000012141 vanillin Nutrition 0.000 description 2
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- FPIPGXGPPPQFEQ-UHFFFAOYSA-N 13-cis retinol Natural products OCC=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-UHFFFAOYSA-N 0.000 description 1
- VVUMWAHNKOLVSN-UHFFFAOYSA-N 2-(4-ethoxyanilino)-n-propylpropanamide Chemical compound CCCNC(=O)C(C)NC1=CC=C(OCC)C=C1 VVUMWAHNKOLVSN-UHFFFAOYSA-N 0.000 description 1
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- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 1
- HIQIXEFWDLTDED-UHFFFAOYSA-N 4-hydroxy-1-piperidin-4-ylpyrrolidin-2-one Chemical compound O=C1CC(O)CN1C1CCNCC1 HIQIXEFWDLTDED-UHFFFAOYSA-N 0.000 description 1
- 206010006458 Bronchitis chronic Diseases 0.000 description 1
- WTEVQBCEXWBHNA-UHFFFAOYSA-N Citral Natural products CC(C)=CCCC(C)=CC=O WTEVQBCEXWBHNA-UHFFFAOYSA-N 0.000 description 1
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- FPIPGXGPPPQFEQ-BOOMUCAASA-N Vitamin A Natural products OC/C=C(/C)\C=C\C=C(\C)/C=C/C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-BOOMUCAASA-N 0.000 description 1
- FPIPGXGPPPQFEQ-OVSJKPMPSA-N all-trans-retinol Chemical compound OC\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-OVSJKPMPSA-N 0.000 description 1
- 230000000507 anthelmentic effect Effects 0.000 description 1
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- 101150006429 atoB gene Proteins 0.000 description 1
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- 239000003054 catalyst Substances 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 208000007451 chronic bronchitis Diseases 0.000 description 1
- 229940043350 citral Drugs 0.000 description 1
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- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
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- 238000005516 engineering process Methods 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 239000000686 essence Substances 0.000 description 1
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- WTEVQBCEXWBHNA-JXMROGBWSA-N geranial Chemical compound CC(C)=CCC\C(C)=C\C=O WTEVQBCEXWBHNA-JXMROGBWSA-N 0.000 description 1
- HIGQPQRQIQDZMP-UHFFFAOYSA-N geranil acetate Natural products CC(C)=CCCC(C)=CCOC(C)=O HIGQPQRQIQDZMP-UHFFFAOYSA-N 0.000 description 1
- HIGQPQRQIQDZMP-DHZHZOJOSA-N geranyl acetate Chemical compound CC(C)=CCC\C(C)=C\COC(C)=O HIGQPQRQIQDZMP-DHZHZOJOSA-N 0.000 description 1
- YCTKONHGXCYDJD-UHFFFAOYSA-N geranylgeranyl formate Natural products CC(=CCCC(=CCCC(=CCCC(=CCOC=O)C)C)C)C YCTKONHGXCYDJD-UHFFFAOYSA-N 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 229930002839 ionone Natural products 0.000 description 1
- 150000002499 ionone derivatives Chemical class 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000002844 melting Methods 0.000 description 1
- 230000008018 melting Effects 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 238000009629 microbiological culture Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 239000012074 organic phase Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 238000011895 specific detection Methods 0.000 description 1
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- 150000003505 terpenes Chemical class 0.000 description 1
- MWOOGOJBHIARFG-UHFFFAOYSA-N vanillin Chemical compound COC1=CC(C=O)=CC=C1O MWOOGOJBHIARFG-UHFFFAOYSA-N 0.000 description 1
- 235000019155 vitamin A Nutrition 0.000 description 1
- 239000011719 vitamin A Substances 0.000 description 1
- 229940045997 vitamin a Drugs 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/74—Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
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Abstract
The invention provides a method for constructing recombinant vibrio natriegens for producing geraniol, the recombinant vibrio natriegens constructed by the method and application of the recombinant vibrio natriegens. The recombinant vibrio natriegens can be subjected to fermentation culture through conventional vibrio natriegens and LBv2 culture media, and geraniol expression with high yield is obtained through IPTG induction in the culture process, wherein the geraniol expression is about 9.97mg/L, the yield is obviously improved compared with the yield (1.22mg/L) of geraniol produced by using escherichia coli as chassis cells, and the recombinant vibrio natriegens has stronger industrial potential of geraniol.
Description
Technical Field
The invention belongs to the field of genetically engineered bacteria, and particularly relates to a method for constructing recombinant vibrio natriegens for producing geraniol, the recombinant vibrio natriegens constructed by the method and application of the recombinant vibrio natriegens.
Background
Geraniol (Geraniol) and ester derivatives thereof are frequently used as raw materials for perfumes. It is colorless, has a sweet rose smell and citrus taste, and is often used for making perfumes and essences. Its liquid state is similar to oil, so its melting point is low, boiling point is high, and it can be dissolved in oil. Geraniol can be used to prepare a variety of drugs. The extract can be used for treating hepatocarcinoma, chronic bronchitis, antibacterial agent, and anthelmintic. Through chemical treatment, geraniol ester (such as geranyl acetate and geranyl formate), citral, vanillin, vanillyl alcohol, vitamin A, ionone and hydroxy vanillin can be prepared.
The geraniol can be extracted from plants such as Murraya paniculata (Murraya paniculata), Allium sativum (Allium sativum), Laurus nobilis (Laurus nobilis), Rosa rugosa, and Geranium japonicum. The method needs to use a large amount of energy and add a chemical catalyst, generates a large amount of emission pollution, and uses a large amount of plants for industrial extraction, thereby having the hidden danger of environmental pollution. Compared with other methods, the traditional biological method for producing the geraniol uses the escherichia coli as the underpan cells, and the biological method uses natural components, does not need plant addition, and has the advantages of greatly reducing the environmental pollution and the production cost and having the defect of low production yield.
Therefore, there is a need to further develop a new geraniol production process.
Disclosure of Invention
Accordingly, it is an object of the present invention to provide a method for constructing recombinant Vibrio natriegens that produce geraniol.
Another object of the present invention is to provide a recombinant Vibrio natriegens constructed by the above method.
It is still another object of the present invention to provide the above recombinant Vibrio natriegens application.
It is still another object of the present invention to provide a method for producing geraniol.
In one aspect, the present invention provides a method for constructing a recombinant vibrio natriegens that produces geraniol, the method comprising: and transforming the vector capable of expressing the geraniol into the initiating bacterium vibrio natriegens to obtain the recombinant bacterium.
In the present invention, the Latin chemical name of Vibrio natriegens is Vibrio natriegens, which may be model strains used for research production, for example, ATCC 14048, CGMCC1.8729, and the like.
In the present invention, the vector capable of expressing geraniol refers to a vector capable of achieving expression of geraniol in a host, and examples thereof include, but are not limited to, plasmid vectors containing a geraniol expression precursor and an expression pathway.
The vector capable of expressing geraniol may consist of one vector, or the vector capable of expressing geraniol may consist of a plurality of vectors to achieve the purpose of expressing geraniol collectively.
In one embodiment, the vector capable of expressing geraniol may consist of a plasmid containing a geraniol expression precursor and a plasmid of a geraniol expression pathway.
The plasmid containing a geraniol expression precursor may refer to a plasmid containing a key gene of mevalonate pathway (MVA pathway). The plasmid for geraniol expression pathway may refer to a plasmid containing a key gene for geraniol synthesis. Mevalonate pathway (MVA pathway) key genes are well known to those skilled in the art and include, for example, the idi, PMD, PMK, MK, HMGR, HMGS and atoB genes. The key genes for geraniol synthesis are well known to those skilled in the art and include, for example, the GPPS and GES synthesis genes.
In one embodiment, the sequence of the plasmid containing the geraniol expression precursor is shown in SEQ ID NO. 1.
In one embodiment, the sequence of the plasmid of the geraniol expression pathway is set forth in SEQ ID NO 2.
In one embodiment, the method for constructing recombinant vibrio natriegens for producing geraniol comprises the step of co-transforming a plasmid containing a geraniol expression precursor shown as SEQ ID NO:1 and a plasmid of a geraniol expression pathway shown as SEQ ID NO:2 into a developing vibrio natriegens (particularly CGMCC1.8729) to obtain a recombinant bacterium.
In the present invention, methods for transforming vectors capable of expressing geraniol into the developing strain Vibrio natriensis are well known to those skilled in the art and include, but are not limited to, chemical transformation methods, electric shock methods, and the like.
In another aspect, the present invention provides a recombinant Vibrio natriegens constructed by the above method.
In a further aspect, the present invention provides the use of the recombinant Vibrio natriegens described above for the production of geraniol.
In still another aspect, the present invention provides a method for producing geraniol, comprising the step of producing geraniol using the above-described recombinant Vibrio natriegens.
In the method for producing geraniol of the present invention, it is preferable to use LBv2 medium as the fermentation medium, and LBv2 medium is a sterile aqueous solution containing the following components: 21.8g/L NaCl, 10g/L peptone, 5g/L yeast extract, 0.31g/L KCl, 4.7g/L MgCl6H 2 O。
The invention develops an engineering bacterium for expressing geraniol, which can be fermented and cultured by a conventional vibrio natriegens and LBv2 culture medium, and the geraniol expression with high yield is obtained by IPTG induction in the culture process, wherein the geraniol expression is about 9.97mg/L, and is obviously improved compared with the geraniol yield (1.22mg/L) generated by using escherichia coli as a chassis cell.
Drawings
FIG. 1A is a plasmid map of a plasmid containing a geraniol expression precursor.
FIG. 1B is a plasmid map of a plasmid for the geraniol expression pathway.
Detailed Description
Hereinafter, the present invention will be described in detail by examples. However, the examples provided herein are for illustrative purposes only and are not intended to limit the present invention.
The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.
Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
The enzymatic reagents used were purchased from ThermoFisher and New England Biolabs (NEB), the kits used for the extraction of plasmids were purchased from Tiangen Biotechnology (Beijing) Ltd, the corresponding procedures were carried out strictly according to the product instructions, and all the media were prepared with deionized water if no special instructions were given. The starting strain is vibrio natriegens with the number of CGMCC1.8729 (purchased from China general microbiological culture Collection center).
The formula of the culture medium is as follows:
1) escherichia coli culture medium
LB medium: 5g/L yeast extract (from OXID, U.K., catalog No. LP0021), 10g/L peptone (from OXID, U.K., catalog No. LP0042),10g/L NaCl, and the balance water. Adjusting pH to 7.0-7.2, and sterilizing with high pressure steam.
LBv2 culture medium: 21.8g/L NaCl, 10g/L peptone, 5g/L yeast extract, 0.31g/LKCl, 4.7g/L MgCl6H 2 O and the balance of water. And (5) high-pressure steam sterilization.
Example 1
The specific operation is as follows:
1. plasmid extraction: plasmids pSC101-TALESP2-GPPS-GES (No.: BBa _ K2753019, http://2018.igem.org/Team: Greatby _ China/Design) containing geraniol expression precursors and plasmid p15A-MVA (Martin, V.J.J., Pitera, D.J., Withers, S.T., Newman, J.D., & Keasling, J.D. (2003). Engineering a methanol pathway in Escherichia coli for production of terpenoids,21 (7)), 796-. The plasmid map of pSC101-TALESP2-GPPS-GES is shown in figure 1A, and the sequence of pSC101-TALESP2-GPPS-GES is shown in SEQ ID NO: 1. The map of the p15A-MVA plasmid is shown in figure 1B, and the sequence of the p15A-MVA plasmid is shown in SEQ ID NO: 2.
By CaCl 2 The two plasmids were co-transformed into Vibrio natriegens Chassis cells (experimental group) and Escherichia coli Chassis cells DH5 alpha (purchased from Tiangen Biochemical technology, Beijing, Ltd., product No. CB10102) (control group), respectively.
2. The Vibrio natriegens and Escherichia coli transformed with the above-obtained two plasmids were cultured and fermented: the fermentation system is 50mL, 3 groups of parallel experiments are set, and are respectively named as vibrio natriuretic-1, vibrio natriuretic-2, vibrio natriuretic-3, escherichia coli-1, escherichia coli-2 and escherichia coli-3. Firstly, 0.5mL of overnight shake bacteria liquid is added into 50mL of LB/LBv2 liquid culture medium with corresponding resistance, the mixture is placed at 37 ℃ and cultured under the condition of 200rpm until the OD600 is about 0.1, then the condition is changed to 30 ℃ and 200rpm, the bacteria is continuously shaken until the OD600 is 0.4-0.6, IPTG with the final concentration of 200 MuM is added for induction, 15% of isopropyl myristate is added at the same time, and fermentation is carried out for about 3 days under the condition of 30 ℃ and 200 rpm. Wherein, the liquid culture medium used by the vibrio natriegens is LBv2, and the liquid culture medium used by the escherichia coli is LB.
And after the fermentation is finished, centrifuging the fermentation liquor, standing for 1h, taking the supernatant, filtering the supernatant through a 0.22-micron organic phase filter membrane, taking the upper oily liquid as a sample, and waiting for GC detection.
The geraniol extracted above was detected using Gas Chromatography (GC) of shimadzu 2010 Pro.
GC analysis for geraniol content: a gas chromatograph model GC-2010Pro from Shimadzu was used.
The chromatograph is configured to: an HP-5 type capillary chromatographic column, a hydrogen flame ionization detector FID and an SPL shunt sample inlet; high-purity nitrogen is used as carrier gas, hydrogen is fuel gas, and air is combustion-supporting gas; an AOC-20S autosampler was used, and acetone was used as a washing solution.
The settings of the GC analysis program were: a chromatographic column: SH-Rtx-5, specification of 30.0m multiplied by 0.25mm, sample injection volume of 1 mu L, split ratio of 20, gasification chamber temperature of 250 ℃, chromatographic column temperature: 100 ℃, the temperature of the FID detector is 250 ℃, and the total time of the temperature rise program is 8.8 min.
The specific detection and calculation results are shown in tables 1 and 2.
TABLE 1
TABLE 2
The recombinant vibrio natriegens can be subjected to fermentation culture through conventional vibrio natriegens and LBv2 culture media, and the high-yield geraniol expression is obtained through IPTG induction in the culture process, wherein the yield is about 9.97mg/L, and is obviously higher than the yield (1.22mg/L) of geraniol produced by using escherichia coli as chassis cells, so that the recombinant vibrio natriegens has stronger industrial potential of geraniol.
SEQUENCE LISTING
<110> Shenzhen Lanjing Biotech Ltd
<120> method for constructing recombinant Vibrio natriegens that produce geraniol and recombinant Vibrio natriegens constructed by the same
And uses thereof
<130> DI20-2217-XC37
<160> 2
<170> PatentIn version 3.5
<210> 1
<211> 2048
<212> DNA
<213> Artificial sequence
<220>
<223> pSC101-TALESP2-GPPS-GES sequence
<400> 1
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aaaaaaagga tctcaagaag atcctttgat cttttctacg gcgcgcccag ctgtctaggg 120
cggcggattt gtcctactca ggagagcgtt caccgacaaa caacagataa aacgaaaggc 180
ccagtctttc gactgagcct ttcgttttat ttgatgcctt taattaaagc ggataacaat 240
ttcacacagg atcgcggccg cttctagagc tcggtaccaa attccagaaa agaggcctcc 300
cgaaaggggg gccttttttc gttttggtcc tactggcgcg cctcagtcag agtattgact 360
taaagtctaa cctataggag atctacagcc atcgagagct gcgagactgt cgccggatgt 420
gtatccgacc tgacgatggc ccaaaagggc cgaaacagtc ctctacaaat aattttgttt 480
aaatcaattc atcgacgtga aaatggtaga tttaagaact ttaggatatt cacagcagca 540
acaggaaaag atcaagccca aagttaggtc gacagtcgcg cagcatcacg aagcgctggt 600
tggtcatggg tttacacatg cccacatcgt agccttatcg cagcaccctg ccgcccttgg 660
cacggtcgcc gtcaagtacc aggacatgat tgcggcgttg ccggaagcca cacatgaggc 720
gatcgtcggt gtggggaaac agtggagcgg agcccgagcg cttgaggccc tgttgacggt 780
cgcgggagag ctgagagggc ctccccttca gctggacacg ggccagttgc tgaagatcgc 840
gaagcgggga ggagtcacgg cggtcgaggc ggtgcacgcg tggcgcaatg cgctcacggg 900
agcacccctc aacctgaccc cggaccaggt agtcgcgatt gcttcacatg acgggggtaa 960
acaagcgctg gaaacggtgc agcgtctgct accggtgtta tgtcaggatc atgggctcac 1020
gccggaacag gtagtggcaa ttgcgagtca tgacggtggc aaacaggccc tggaaaccgt 1080
acagcggctg ctaccggtgc tgtgtcaagc gcatggcctg actccggacc aagttgtagc 1140
cattgcctcg aacgggggcg gcaagcaggc gctggagact gttcaacgtc tgctccccgt 1200
tctgtgtcag gcgcatggcc tgacgcctgc gcaggtcgtg gcgatcgctt caaacatcgg 1260
tgggaagcaa gccctggaga ctgtccaaag actgttgcca gtgttgtgtc aagatcatgg 1320
cttaacccca gatcaggtgg ttgcgattgc atcaaatgga ggtggtaaac aggcgctgga 1380
gactgtgcag cgcctgttgc cggttctgtg ccaagatcat gggctgactc cggaacaggt 1440
tgtggctatc gcaagcaata ttggtggcaa gcaggccctg gaaacagtac agcgcctgct 1500
gcctgtattg tgtcaagccc acggtcttac ccccgatcag gtagtcgcca tagcatcgca 1560
cgacggcggg aagcaggccc ttgagactgt acaacgcctc ctgccggttt tgtgccaggc 1620
gcacggcctg acgccagccc aggtggttgc gatagccagt aataacggcg gtaagcaagc 1680
ccttgaaacc gttcaacgtt tgctgccagt gctgtgccag gatcacggcc tgaccccgga 1740
tcaggtagtc gccattgcta gcaacattgg tggcaaacaa gcactggaga cagttcaacg 1800
cttactgccc gtgctttgcc aggatcatgg actgacccca gagcaagtgg tcgcgattgc 1860
ctcgcatgac ggaggtaaac aggcccttga gactgtccag cgtctgctgc cggtcctttg 1920
ccaggctcat gggctgacgc ccgaccaggt ggtagcaatc gcttcgaacg gcggaggcaa 1980
acaggcgtta gaaaccgttc aacgtctgtt accggtgctg tgccaagctc atggcttaac 2040
cccggccc 2048
<210> 2
<211> 12103
<212> DNA
<213> Artificial sequence
<220>
<223> p15A-MVA sequence
<400> 2
gacgtcggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct 60
ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga 120
ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag 180
ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg 240
ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc 300
ttcggtatcg tcgtatccca ctaccgagat gtccgcacca acgcgcagcc cggactcggt 360
aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac 420
gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc 480
ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag 540
acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc 600
caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact 660
gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc 720
ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg 780
ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat 840
cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg 900
cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc 960
cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac 1020
tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg 1080
ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac 1140
cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc 1200
gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca 1260
gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg 1320
cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca 1380
tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag 1440
caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga 1500
tcgtttaggc accccaggct ttacacttta tgcttccggc tcgtataatg tgtggaattg 1560
tgagcggata acaatttcag aattcaaaag atcttaggag gaatataaaa tgaaaaattg 1620
tgtcatcgtc agtgcggtac gtactgctat cggtagtttt aacggttcac tcgcttccac 1680
cagcgccatc gacctggggg cgacagtaat taaagccgcc attgaacgtg caaaaatcga 1740
ttcacaacac gttgatgaag tgattatggg taacgtgtta caagccgggc tggggcaaaa 1800
tccggcgcgt caggcactgt taaaaagcgg gctggcagaa acggtgtgcg gattcacggt 1860
caataaagta tgtggttcgg gtcttaaaag tgtggcgctt gccgcccagg ccattcaggc 1920
aggtcaggcg cagagcattg tggcgggggg tatggaaaat atgagtttag ccccctactt 1980
actcgatgca aaagcacgct ctggttatcg tcttggagac ggacaggttt atgacgtaat 2040
cctgcgcgat ggcctgatgt gcgccaccca tggttatcat atggggatta ccgccgaaaa 2100
cgtggctaaa gagtacggaa ttacccgtga aatgcaggat gaactggcgc tacattcaca 2160
gcgtaaagcg gcagccgcaa ttgagtccgg tgcttttaca gccgaaatcg tcccggtaaa 2220
tgttgtcact cgaaagaaaa ccttcgtctt cagtcaagac gagttcccga aagcgaactc 2280
aacggctgaa gcgttaggtg cattgcgccc ggccttcgat aaagcaggaa cagtcaccgc 2340
tgggaacgcg tctggtatta acgacggtgc tgccgctctg gtgattatgg aagaatctgc 2400
ggcgctggca gcaggcctta cccccctggc tcgcattaaa agttatgcca gcggtggcgt 2460
gccccccgca ttgatgggta tggggccagt acctgccacg caaaaagcgt tacaactggc 2520
ggggctgcaa ctggcggata ttgatctcat tgaggctaat gaagcatttg ctgcacagtt 2580
ccttgccgtt gggaaaaacc tgggctttga ttctgagaaa gtgaatgtca acggcggggc 2640
catcgcgctc gggcatccta tcggtgccag tggtgctcgt attctggtca cactattaca 2700
tgccatgcag gcacgcgata aaacgctggg gctggcaaca ctgtgcattg gcggcggtca 2760
gggaattgcg atggtgattg aacggttgaa ttgaggatct tgaattaagg aggacagcta 2820
aatgacaata ggtatcgata aaataaactt ttacgttcca aagtactatg tagacatggc 2880
taaattagca gaagcacgcc aagtagaccc aaacaaattt ttaattggaa ttggtcaaac 2940
tgaaatggct gttagtcctg taaaccaaga catcgtttca atgggcgcta acgctgctaa 3000
ggacattata acagacgaag acaaaaagaa aattggtatg gtaattgtgg caactgaatc 3060
agcagttgat gctgctaaag cagccgctgt tcaaattcac aacttattag gtattcaacc 3120
ttttgcacgc tgctttgaaa tgaaagaagc ttgttatgct gcaacaccag caattcaatt 3180
agctaaagat tatttagcaa ctagaccgaa tgaaaaagta ttagttattg ctacagatac 3240
agcacgttat ggattgaact caggcggcga gccaacacaa ggtgctggcg cagttgcgat 3300
ggttattgca cataatccaa gcattttggc attaaatgaa gatgctgttg cttacactga 3360
agacgtttat gatttctggc gtccaactgg acataaatat ccattagttg atggtgcatt 3420
atctaaagat gcttatatcc gctcattcca acaaagctgg aatgaatacg caaaacgtca 3480
aggtaagtcg ctagctgact tcgcatctct atgcttccat gttccattta caaaaatggg 3540
taaaaaggca ttagagtcaa tcattgataa cgctgatgaa acaactcaag agcgtttacg 3600
ttcaggatat gaagatgctg tagattataa ccgttatgtc ggtaatattt atactggatc 3660
attatattta agcctaatat cattacttga aaatcgagat ttacaagctg gtgaaacaat 3720
cggtttattc agttatggct caggttcagt tggtgaattt tatagtgcga cattagttga 3780
aggctacaaa gatcatttag atcaagctgc acataaagca ttattaaata accgtactga 3840
agtatctgtt gatgcatatg aaacattctt caaacgtttt gatgacgttg aatttgacga 3900
agaacaagat gctgttcatg aagatcgtca tattttctac ttatcaaata ttgaaaataa 3960
cgttcgcgaa tatcacagac cagagtaatt aggatctatt caggaaacag accatgtcca 4020
tgcaaagttt agataagaat tttcgacatt tatctcgtaa agaaaagtta caacaattgg 4080
ttgataagca atggttatca gaagaacaat tcgacatttt actgaatcat ccattaatcg 4140
atgaagaagt agccaatagt ttaattgaaa atgtcatcgc gcaaggtgca ttacccgttg 4200
gattattacc gaatatcatt gtggacgata aggcatatgt tgtacctatg atggtggaag 4260
agccttcagt tgtcgctgca gctagttatg gtgcaaagct agtgaatcag actggcggat 4320
ttaaaacggt atcttctgaa cgtattatga taggtcaaat cgtctttgat ggcgttgacg 4380
atactgaaaa attatcagca gacattaaag ctttagaaaa gcaaattcat aaaattgcgg 4440
atgaggcata tccttctatt aaagcgcgtg gtggtggtta ccaacgtata gcgattgata 4500
catttcctga gcaacagtta ctatctttaa aagtatttgt tgatacgaaa gatgctatgg 4560
gcgctaatat gcttaatacg attttagagg ccataactgc atttttaaaa aatgaatttc 4620
cgcaaagcga cattttaatg agtattttat ccaatcatgc aacagcgtcc gttgttaaag 4680
ttcaaggcga aattgatgtt aaagatttag caaggggcga gagaactgga gaagaggttg 4740
ccaaacgaat ggaacgtgct tctgtattgg cccaagtaga tattcatcgt gcagcaacac 4800
ataataaagg tgttatgaat ggcatacatg ctgttgtttt agcaacagga aatgatacgc 4860
gtggtgcaga agcaagtgcg catgcatacg cgagtcgtga cggacagtat cgtggtattg 4920
ctacatggcg ttacgatcaa gatcgtcaac gattgattgg tacaattgaa gtgcctatga 4980
cattggcaat cgttggcggt ggtacaaaag tattaccaat tgctaaagct tcattagagc 5040
tactaaatgt agagtcagca caagaattag gtcatgtagt tgctgccgtt ggtttagcgc 5100
aaaactttgc agcatgtcgc gcgcttgtgt cagaaggtat tcaacaaggt catatgagtt 5160
tacaatataa atcattagct atcgttgtag gggcaaaagg tgatgaaatt gctaaagtag 5220
ctgaagcttt gaaaaaagaa ccccgtgcaa atacacaagc agcggaacat attttacaag 5280
aaattagaca acaataagga tctttttaag gatctccagg catcaaataa aacgaaaggc 5340
tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctctacta 5400
gagtcacact ggctcacctt cgggtgggcc tttctgcgtt tatagcgaat tgatctggtt 5460
tgacagctta tcatcgactg cacggtgcac caatgcttct ggcgtcaggc agccatcgga 5520
agctgtggta tggctgtgca ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca 5580
ctcccgttct ggataatgtt ttttgcgccg acatcataac ggttctggca aatattctga 5640
aatgagctgt tgacaattaa tcatccggct cgtataatgt gtggaattgt gagcggataa 5700
caatttcagg atctaggagg aaataaccat gtctctgcca ttcctgacgt ctgcgccagg 5760
taaggtgatc atcttcggcg agcactctgc ggtgtacaat aagccggccg tcgccgcctc 5820
tgtgtctgcg ttacgcacct acctgctgat cagcgaatct tctgcaccgg acacgatcga 5880
gctggacttt ccggacatca gcttcaacca caagtggagc atcaacgact tcaacgcgat 5940
cacggaggac caggtgaaca gccaaaagct ggccaaagcc cagcaagcaa ccgacggtct 6000
gtctcaggag ctggtgtctc tgctggaccc gctgttagcg cagttaagcg agagcttcca 6060
ttaccacgcc gcgttctgct tcctgtacat gttcgtttgc ctgtgcccgc acgcaaagaa 6120
catcaagttc agcctgaaga gcacgctgcc gattggcgca ggcttaggct ctagcgcatc 6180
tatcagcgtg agcctggcgc tggcgatggc ctatctgggt ggcctgattg gcagcaacga 6240
cctggagaaa ctgagcgaaa acgacaagca catcgtgaac cagtgggcct ttatcggcga 6300
gaagtgcatt catggcaccc cgagcggcat tgacaacgca gttgccacgt atggcaacgc 6360
cctgctgttc gagaaagaca gccacaacgg cacgatcaac acgaacaact tcaagttcct 6420
ggacgacttc ccggcgatcc cgatgattct gacctacacc cgtatcccac gcagcaccaa 6480
ggatttagtc gcccgcgtgc gtgttttagt caccgaaaag ttcccggagg tgatgaagcc 6540
gatcctggac gcgatgggcg agtgcgcgct gcagggtctg gagatcatga ccaagctgag 6600
caagtgcaag ggcaccgacg atgaggcggt ggagaccaac aatgagctgt acgagcagct 6660
gctggagctg atccgtatca atcacggcct gctggtctct atcggtgtgt ctcacccggg 6720
cctggaactg atcaaaaacc tgagcgacga cctgcgcatt ggctctacga aattaacggg 6780
tgcaggtggc ggtggctgct ctttaacgct gctgcgccgt gacattacgc aggagcaaat 6840
cgacagcttc aagaagaagc tgcaggacga cttcagctac gagacgttcg agacggacct 6900
gggcggcacg ggctgttgcc tgctgagcgc caaaaatctg aacaaggacc tgaagatcaa 6960
aagcctggtg ttccagctgt tcgaaaacaa gacgaccacg aagcagcaga tcgacgacct 7020
gttactgccg ggtaacacca atctgccgtg gacgtcttaa ggatctagga gggagatcat 7080
atgagcgaat tacgtgcatt cagcgcgcca ggtaaggcac tgctggccgg tggctacctg 7140
gtgttagaca ccaagtacga ggcgttcgtc gtcggcttat ctgcccgtat gcatgcagtt 7200
gcccacccgt atggtagcct gcagggctct gacaagttcg aagtgcgtgt gaagagcaag 7260
cagttcaagg acggcgagtg gctgtaccac attagcccaa agagcggctt catcccggtt 7320
agcattggtg gcagcaagaa cccatttatc gagaaggtca ttgccaacgt cttcagctac 7380
ttcaagccga atatggacga ttactgcaac cgcaacctgt tcgtcatcga cattttcagc 7440
gacgacgcgt accacagcca agaggactct gttacggagc atcgtggtaa ccgccgcctg 7500
agcttccaca gccatcgcat tgaggaggtg ccgaagacgg gtctgggttc tagcgccggt 7560
ttagttaccg tcttaacgac ggcgttagcg agcttcttcg tgagcgacct ggagaacaac 7620
gtggacaagt accgcgaagt gattcataac ctggcgcagg tggcacattg tcaggcccaa 7680
ggtaagattg gctctggttt tgatgtggca gcggccgcct atggctctat ccgctatcgc 7740
cgctttccgc cggccctgat cagcaatctg ccggacatcg gctctgcgac gtatggtagc 7800
aaactggcgc atctggtgga cgaagaagac tggaacatca ccattaagtc taatcacctg 7860
ccgagcggct taacgttatg gatgggcgat atcaagaacg gcagcgaaac ggttaagctg 7920
gtgcagaaag tgaaaaactg gtacgacagc cacatgccgg aaagcctgaa gatttacacg 7980
gagctggacc acgccaatag ccgtttcatg gatggtctga gcaagctgga ccgcctgcac 8040
gaaacccacg acgactacag cgaccaaatc ttcgagagcc tggagcgcaa tgactgcacc 8100
tgccagaagt acccggagat cacggaggtc cgcgatgccg tggcaacgat tcgccgtagc 8160
ttccgcaaaa ttacgaagga gagcggcgcg gatatcgaac caccggtcca gacgtctctg 8220
ctggacgact gtcaaacctt aaagggcgtg ttaacgtgcc tgattccggg cgcgggtggt 8280
tacgacgcca ttgccgtcat cacgaaacag gacgtcgatc tgcgcgcaca aacggccaac 8340
gacaaacgtt tcagcaaagt ccaatggctg gatgttacgc aggccgactg gggtgttcgc 8400
aaggagaagg acccggaaac gtatctggat aagtgaggat ctaggaggat tatgagatga 8460
ccgtttacac agcatccgtt accgcacccg tcaacatcgc aacccttaag tattggggga 8520
aaagggacac gaagttgaat ctgcccacca attcgtccat atcagtgact ttatcgcaag 8580
atgacctcag aacgttgacc tctgcggcta ctgcacctga gtttgaacgc gacactttgt 8640
ggttaaatgg agaaccacac agcatcgaca atgaaagaac tcaaaattgt ctgcgcgacc 8700
tacgccaatt aagaaaggaa atggaatcga aggacgcctc attgcccaca ttatctcaat 8760
ggaaactcca cattgtctcc gaaaataact ttcctacagc agctggttta gcttcctccg 8820
ctgctggctt tgctgcattg gtctctgcaa ttgctaagtt ataccaatta ccacagtcaa 8880
cttcagaaat atctagaata gcaagaaagg ggtctggttc agcttgtaga tcgttgtttg 8940
gcggatacgt ggcctgggaa atgggaaaag ctgaagatgg tcatgattcc atggcagtac 9000
aaatcgcaga cagctctgac tggcctcaga tgaaagcttg tgtcctagtt gtcagcgata 9060
ttaaaaagga tgtgagttcc actcagggta tgcaattgac cgtggcaacc tccgaactat 9120
ttaaagaaag aattgaacat gtcgtaccaa agagatttga agtcatgcgt aaagccattg 9180
ttgaaaaaga tttcgccacc tttgcaaagg aaacaatgat ggattccaac tctttccatg 9240
ccacatgttt ggactctttc cctccaatat tctacatgaa tgacacttcc aagcgtatca 9300
tcagttggtg ccacaccatt aatcagtttt acggagaaac aatcgttgca tacacgtttg 9360
atgcaggtcc aaatgctgtg ttgtactact tagctgaaaa tgagtcgaaa ctctttgcat 9420
ttatctataa attgtttggc tctgttcctg gatgggacaa gaaatttact actgagcagc 9480
ttgaggcttt caaccatcaa tttgaatcat ctaactttac tgcacgtgaa ttggatcttg 9540
agttgcaaaa ggatgttgcc agagtgattt taactcaagt cggttcaggc ccacaagaaa 9600
caaacgaatc tttgattgac gcaaagactg gtctaccaaa ggaataagga tctaggaggt 9660
aatgataatg caaacggaac acgtcatttt attgaatgca cagggagttc ccacgggtac 9720
gctggaaaag tatgccgcac acacggcaga cacccgctta catctcgcgt tctccagttg 9780
gctgtttaat gccaaaggac aattattagt tacccgccgc gcactgagca aaaaagcatg 9840
gcctggcgtg tggactaact cggtttgtgg gcacccacaa ctgggagaaa gcaacgaaga 9900
cgcagtgatc cgccgttgcc gttatgagct tggcgtggaa attacgcctc ctgaatctat 9960
ctatcctgac tttcgctacc gcgccaccga tccgagtggc attgtggaaa atgaagtgtg 10020
tccggtattt gccgcacgca ccactagtgc gttacagatc aatgatgatg aagtgatgga 10080
ttatcaatgg tgtgatttag cagatgtatt acacggtatt gatgccacgc cgtgggcgtt 10140
cagtccgtgg atggtgatgc aggcgacaaa tcgcgaagcc agaaaacgat tatctgcatt 10200
tacccagctt aaataaggat ctcgcaaaaa accccggatc caaactcgag taaggatctc 10260
caggcatcaa ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg 10320
tttgtcggtg aacgctctct actagagtca cactggctca ccttcgggtg ggcctttctg 10380
cgtttatacc tagggatata ttccgcttcc tcgctcactg actcgctacg ctcggtcgtt 10440
cgactgcggc gagcggaaat ggcttacgaa cggggcggag atttcctgga agatgccagg 10500
aagatactta acagggaagt gagagggccg cggcaaagcc gtttttccat aggctccgcc 10560
cccctgacaa gcatcacgaa atctgacgct caaatcagtg gtggcgaaac ccgacaggac 10620
tataaagata ccaggcgttt ccccctggcg gctccctcgt gcgctctcct gttcctgcct 10680
ttcggtttac cggtgtcatt ccgctgttat ggccgcgttt gtctcattcc acgcctgaca 10740
ctcagttccg ggtaggcagt tcgctccaag ctggactgta tgcacgaacc ccccgttcag 10800
tccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccgga aagacatgca 10860
aaagcaccac tggcagcagc cactggtaat tgatttagag gagttagtct tgaagtcatg 10920
cgccggttaa ggctaaactg aaaggacaag ttttggtgac tgcgctcctc caagccagtt 10980
acctcggttc aaagagttgg tagctcagag aaccttcgaa aaaccgccct gcaaggcggt 11040
tttttcgttt tcagagcaag agattacgcg cagaccaaaa cgatctcaag aagatcatct 11100
tattaatcag ataaaatatt tctagatttc agtgcaattt atctcttcaa atgtagcacc 11160
tgaagtcagc cccatacgat ataagttgtt actagtgctt ggattctcac caataaaaaa 11220
cgcccggcgg caaccgagcg ttctgaacaa atccagatgg agttctgagg tcattactgg 11280
atctatcaac aggagtccaa gcgagctcga tatcaaatta cgccccgccc tgccactcat 11340
cgcagtactg ttgtaattca ttaagcattc tgccgacatg gaagccatca caaacggcat 11400
gatgaacctg aatcgccagc ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca 11460
tggtgaaaac gggggcgaag aagttgtcca tattggccac gtttaaatca aaactggtga 11520
aactcaccca gggattggct gagacgaaaa acatattctc aataaaccct ttagggaaat 11580
aggccaggtt ttcaccgtaa cacgccacat cttgcgaata tatgtgtaga aactgccgga 11640
aatcgtcgtg gtattcactc cagagcgatg aaaacgtttc agtttgctca tggaaaacgg 11700
tgtaacaagg gtgaacacta tcccatatca ccagctcacc gtctttcatt gccatacgaa 11760
attccggatg agcattcatc aggcgggcaa gaatgtgaat aaaggccgga taaaacttgt 11820
gcttattttt ctttacggtc tttaaaaagg ccgtaatatc cagctgaacg gtctggttat 11880
aggtacattg agcaactgac tgaaatgcct caaaatgttc tttacgatgc cattgggata 11940
tatcaacggt ggtatatcca gtgatttttt tctccatttt agcttcctta gctcctgaaa 12000
atctcgataa ctcaaaaaat acgcccggta gtgatcttat ttcattatgg tgaaagttgg 12060
aacctcttac gtgccgatca acgtctcatt ttcgccagat atc 12103
Claims (10)
1. A method for constructing a recombinant vibrio natriegens that produce geraniol, the method comprising: and transforming the vector capable of expressing the geraniol into the initiating bacterium vibrio natriegens to obtain the recombinant bacterium.
2. The method as claimed in claim 1, wherein the vibrio natriegens is CGMCC1.8729 or ATCC 14048.
3. The method according to claim 1, wherein the vector capable of expressing geraniol is a plasmid vector containing a geraniol expression precursor and an expression pathway.
4. The method according to claim 3, wherein the plasmid vector containing geraniol expression precursor and expression pathway consists of a plasmid containing geraniol expression precursor and a plasmid of geraniol expression pathway.
5. The method according to claim 4, wherein the plasmid containing the geraniol expression precursor is represented by SEQ ID NO. 1.
6. The method according to claim 4 or 5, wherein the plasmid of the geraniol expression pathway is represented by SEQ ID NO 2.
7. A recombinant Vibrio natriegens obtainable by the method of any one of claims 1 to 6.
8. Use of the recombinant vibrio natriegens of claim 7 for the production of geraniol.
9. A method for producing geraniol comprising the step of producing geraniol using the recombinant vibrio natriegens of claim 7.
10. The method of claim 9, wherein LBv2 medium is used as the fermentation medium, and LBv2 medium is a sterile aqueous solution comprising: 21.8g/L NaCl, 10g/L peptone, 5g/L yeast extract, 0.31g/L KCl, 4.7g/L MgCl6H 2 O。
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105420135A (en) * | 2016-01-06 | 2016-03-23 | 山东大学 | Recombined saccharomyces cerevisiae bacterial strain of high-yield monoterpene geraniol and application of recombined saccharomyces cerevisiae bacterial strain |
| CN110869500A (en) * | 2017-01-26 | 2020-03-06 | 马努斯生物合成股份有限公司 | Metabolic engineering for microbial production of terpenoid products |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105420135A (en) * | 2016-01-06 | 2016-03-23 | 山东大学 | Recombined saccharomyces cerevisiae bacterial strain of high-yield monoterpene geraniol and application of recombined saccharomyces cerevisiae bacterial strain |
| CN110869500A (en) * | 2017-01-26 | 2020-03-06 | 马努斯生物合成股份有限公司 | Metabolic engineering for microbial production of terpenoid products |
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