CN1500872A - High temperaturebeta- glucosaccharase, coding gene and uses thereof - Google Patents
High temperaturebeta- glucosaccharase, coding gene and uses thereof Download PDFInfo
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- CN1500872A CN1500872A CNA021487766A CN02148776A CN1500872A CN 1500872 A CN1500872 A CN 1500872A CN A021487766 A CNA021487766 A CN A021487766A CN 02148776 A CN02148776 A CN 02148776A CN 1500872 A CN1500872 A CN 1500872A
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Abstract
High temperature beta-glucosidase gene is obtained from Thermoanaerobacter tengcongensis MB4 total DNA to constitute prokaryotic expression plasmid and transferred to colibacillus for expression. Amino acid sequence comparison shows that the beta-glucosidase is one new kind of beta-glucosidase.
Description
Technical field
The invention belongs to enzyme genetically engineered and enzyme engineering field.Specifically, the present invention relates to the beta-glucosidase dna sequence dna of a kind of coding thermophilc anaerobe (Thermoanaerobacter tengcongensis) MB4, also relate to recombinant plasmid that contains this enzyme gene and the recombinant bacterial strain of expressing corresponding enzyme.
Background technology
Beta-glucosidase (EC3.2.1.21) can the multiple β glucoside bond of catalytic hydrolysis, has the substrate broad spectrum.Has significant application value in industry such as medicine, food, seasonings, the energy.As in zymamsis industry, produce focus [Bothast etc., modern Application microbiology (Adv.Appl.Microbiol.) 44:261-286,1997 that alcohol becomes present people's concern from the cellulosic materials agricultural wastes; Pemberton etc., Canadian chemical engineering magazine (Can.J.Chem.Eng.) 58:723-729,1980].Utilize biological enzyme to transform Mierocrystalline cellulose and be glucose, need three kinds of enzyme synergies, comprising: cellulase (EC3.2.1.4), circumscribed cellulase (EC3.2.1.91) and beta-glucosidase (EC3.2.1.21).Cellulase (EC3.2.1.4), circumscribed cellulase (EC3.2.1.91) act synergistically on Mierocrystalline cellulose and produce cellobiose, and cellobiose is hydrolyzed to glucose by beta-glucosidase (EC3.2.1.21) afterwards.Usually cellobiose suppresses cellulase to cellulosic degraded, thereby and beta-glucosidase can be removed its restraining effect to cellulase by the hydrolysis fiber disaccharides, so beta-glucosidase plays an important role in cellulose degradation.
Beta-glucosidase separates from different microorganisms, as Aspergillus[Kwon etc., FEMS microorganism communication (FEMS Microbiol Lett) 97:149-154,1992], Humicola[Peralta etc., FEMS microorganism communication (FEMS Microbiol Lett) 146:291-295,1997], Fusarium[Christakopoulos etc., european journal of biological chemistry (Eur.J.Biochem) 224:379-385,1994], Volvariella[Cai etc., use and environmental microbiology (Appl Environ Microbiol) 65:553-559,1999], Candida[Christopher etc., use and environmental microbiology (Appl EnvironMicrobiol) 61:518-525,1995]. the beta-glucosidase with certain thermostability is mainly from thermophile bacteria, as Sulfolobus solfataricus[Aguilar etc., molecular biology magazine (J Mol Biol), 271:789-802,1997], Pyrococcus furiosus[Kengen etc., european journal of biological chemistry (EurJ Biochem) 213:305-312,1993], Thermotoga maritime[Gabelsberger etc., FEMS microorganism communication (FEMS Microbiol Lett), 109:131-138,1993], Thermosphaeraaggregans[Chi etc., FEMS communication (FEBS Letters), 445:375-383,1999], Clostridium thermocellum[Ait etc., general microbiology magazine (J.Gen.Microbiol.) 128:569-577,1982].
About existing patent of beta-glucosidase gene and bibliographical information.As: Dion etc. have reported the beta-glucosidase gene [sugared conjugate magazine (GlycoconjugateJ), 16:27-37,1999] of Thermus thermophilus; Gonzalez etc. have reported the beta-glucosidase gene [biological chemistry and biophysical research communication (Biochem Biophys Res Commun), 194:1359-1364,1993] of bacillus Bacillus polymyxa; Wright etc. have reported the beta-glucosidase gene [using and environmental microbiology (Appl.Environ Microbil) 58:3455-3465,1992] of Microbispora bispora; Paavilainen etc. have reported the beta-glucosidase gene [using and environmental microbiology (Appl Environ Microbiol) 59:927-932,1993] of Bacillus circulans; Breves etc. have reported the beta-glucosidase gene [using and environmental microbiology (Appl Environ Microbiol) 63:3902-3910,1997] of Thermoanaerobacter brockii; Lieb etc. have reported the beta-glucosidase gene [molecule General Genetics (Mol Gen Genet.) 242:111-115,1994] of Thermotoga mariti; Tonouchi etc. have reported the beta-glucosidase gene (U.S. Pat P6316251, November 31 calendar year 2001) of Acetobacter xylinum.
Microorganisms beta-glucosidase kind is many, and the beta-glucosidase of different sources has many-sided different properties, thereby causes the range of application that differs from one another.Thereby need develop the new features beta-glucosidase continuously to satisfy industrial needs better.Thermostable enzyme has very big superiority on using, can improve speed of response, reduce and pollute, strengthen the tolerance of chemical reagent etc.Therefore developing high temperature beta-glucosidase enzyme has become focus.
Summary of the invention
A kind of new high temperature beta-glucosidase that the present invention proposes, its primary structure is different with known beta-glucosidase, and its characteristic is also different with known beta-glucosidase.
Therefore, the purpose of this invention is to provide a kind of novel beta-glucosidase and coding structure gene thereof, recombinant plasmid and reorganization thalline, this gene expression product beta-glucosidase shows active under hot conditions, can be applicable to cellulose degradation technology and other commercial runs.The invention provides a kind of method simultaneously,, other recipient bacterium is arrived in the enzyme gene clone that the present invention relates to, produce the beta-glucosidase that the present invention relates to by other bacterial strain or at other culture condition promptly by molecular biology method.
The present invention studies have shown that, thermophilc anaerobe (Thermoanaerobacter tengcongensis MB4) (Xue Yanfen etc., System and Surroundings microorganism international magazine IJSEM, 51:1335-1341,2001) produce the high temperature beta-glucosidase under hot conditions, this enzymic hydrolysis cellobiose forms glucose.
The present invention obtains the gene of beta-glucosidase from thermophilc anaerobe (Thermoanaerobacter tengcongensis MB4), and it is the DNA of 1578bp, the protein of being made up of 526 amino acid of encoding.Obtained to contain the recombinant plasmid of this gene by molecular biology method, transformed into escherichia coli makes recombinant bacterial strain express beta-glucosidase.Therefore the present invention provides a kind of possibility simultaneously, promptly by genetic engineering or molecular biology method, other recipient bacterium is arrived in the enzyme gene clone that the present invention relates to, and produces the beta-glucosidase that the present invention relates to by other bacterial strain or at other culture condition.
To achieve the object of the present invention, realize that concrete technological step of the present invention is as follows: from thermophilc anaerobe (Thermoanaerobacter tengcongensis MB4), extract total DNA, utilize the shot-gun technology, obtain needed dna fragmentation, be connected on the pUC18 carrier and transformed into escherichia coli DH5 α, obtain to contain the recombinant plasmid pGLUB and the recombinant escherichia coli strain DH5 α GLUB of high temperature alpha-glucosidase gene.This reorganization bacterium is expressed the high temperature glucosidase activity.Under the condition that above-mentioned beta-glucosidase gene is expressed, cultivate this reorganization bacterium,, prove this reorganization bacterium expression high temperature activity of beta-glucosidase through determination of activity.This protein has the high temperature activity of beta-glucosidase, and operative temperature 60-80 ℃, pH6-7, the hydrolyzable cellobiose produces glucose.Order-checking shows that goal gene contains 1578bp, the protein of being made up of 526 amino acid of encoding.
The beta-glucosidase that the present invention relates to is a novel beta-glucosidase.The primary structure of new beta-glucosidase is different with known beta-glucosidase, compares with the aminoacid sequence of other beta-glucosidase of having reported, and similarity is less than 51%.Should be pointed out that amino acid to the expressed enzyme molecule of beta-glucosidase gene of the present invention carries out one or more amino acid and replaces, inserts or lack resulting functional analogue and also can reach purpose of the present invention.Thereby the present invention also comprises having with the aminoacid sequence shown in the SEQ NO.2 to have 80% homology at least, preferably has at least 90% homology, but has the active functional analogue of beta-glucosidase simultaneously.According to the amino acid sequence similarity of enzyme, present known beta-glucosidase belongs to glycoside hydrolase the 1st and 3 two families.By the beta-glucosidase gene deduced amino acid comparative analysis to the present invention relates to, the beta-glucosidase that the present invention relates to belongs to a member in glycoside hydrolase the 3rd family.
The beta-glucosidase that the present invention relates to can be used aspect chemical industry, weaving, food, medicine industry.
The beta-glucosidase that the present invention relates to, main characteristic is as follows:
(1) derives from high temperature anaerobic bacterium (Thermoanaerobacter tengcongensis MB4) or other bacterium that derives.The bacterium that derives is meant and transforms the recombinant bacterial strain that the dna fragmentation that the present invention relates to is arranged;
(2) its gene has the nucleotide sequence shown in the SEQ NO.1;
(3) it has the aminoacid sequence shown in the SEQ NO.2;
(4) have activity of beta-glucosidase, operative temperature 60-80 ℃, pH6-7, molecular weight 59000 dalton.
Further, the present invention relates to a dna sequence dna according to of the present invention, the beta-glucosidase that this dna sequence encoding the present invention relates to, this dna nucleotide sequence:
(1) forms by the dna sequence dna among the SEQ NO.1.
(2) coding one protein, the aminoacid sequence among its aminoacid sequence such as the SEQ NO.2.
The dna sequence dna that the present invention relates to, except the beta-glucosidase SEQNO.1 that coding the present invention relates to, also should comprise: coding carries out one or more amino acid replacements to the amino acid of the expressed enzyme molecule of beta-glucosidase gene of the present invention, insert or lack resulting functional analogue and also can reach target DNA nucleotide sequence of the present invention, thereby the present invention comprises that also coding and the aminoacid sequence shown in the SEQNO.2 have and have 80% homology at least, preferably have at least 90% homology, but have the dna nucleotide sequence of the functional analogue of activity of beta-glucosidase simultaneously.
Description of drawings
Below in conjunction with accompanying drawing the present invention is described in further detail.
The design of graphics of Fig. 1 .pGLUB plasmid.
Embodiment
Embodiment 1.
The extraction of the total DNA of thermophilc anaerobe (Thermoanaerobacter tengcongensis MB4)
Employing is from the isolating thermophilc anaerobe of Chinese yunnan Tengchong hot spring (Thermoanaerobactertengcongensis MB4), get its fresh wet thallus 20 grams, be suspended from 10 milliliters of 50mMTris damping fluids (pH8.0), add a small amount of N,O-Diacetylmuramidase and 8 milliliters of 0.25mMEDTA (pH8.0), place 20min in 37 ℃ behind the mixing, add 2 milliliters of 10%SDS afterwards, place 5min, use equal-volume phenol respectively for 55 ℃, each extracting of chloroform is once got last supernatant solution, add 2 times of volume ethanol, reclaim DNA, respectively with 70% and dehydrated alcohol wash, precipitate and be dissolved in 0.5 milliliter of TE damping fluid (pH8.0,10mM Tris, 1mMEDTA), add 10mg/ml RNase3 μ l, 37 ℃ are incubated 1 hour, use equal-volume phenol respectively, each extracting of chloroform once, supernatant liquor adds 2 times of volume ethanol, reclaims DNA, respectively with 70% and dehydrated alcohol wash, deionized water dissolving is used in vacuum-drying.The ultraviolet spectrophotometer measurement result of dna solution is A260/A280=1.98, A260/A230=2.18.The clone of beta-glucosidase gene
Get foregoing total dna solution 10 μ l (about 50 μ gDNA), partially digested with restriction enzyme Sau3AI, through agarose gel electrophoresis, electroelution reclaims the 2-10kbDNA fragment.Get 2 μ l (5 μ g) Sau3AI enzymolysis dna fragmentation and 1 μ l (1 μ g) carries out ligation through the plasmid pUC18 DNA of BamHI enzymolysis and dephosphorylation in 20 μ l linked systems, wherein contain 2 μ l (10X connects damping fluid), 1 μ l T4DNA ligase enzyme, 14 μ l water.Linked system behind the transformed competence colibacillus bacillus coli DH 5 alpha, is applied to and contains 50ug/ml Amp (penbritin), on the LB solid medium of 0.01%5-bromo-4-chloro-3-indolyl-beta-glucopyranoside 16 ℃ of reactions 16 hours.Cultivated 16-18 hour for 37 ℃, cultivated 1-5 hour at 60 ℃ then, bacterium colony becomes the blue positive colony that is.Positive colony in the Amp-LB substratum 37 ℃ cultivated 16-18 hour, have heat-resisting activity of beta-glucosidase through active testing.Positive bacterium colony is extracted recombinant plasmid with alkaline process, with various restriction enzyme hydrolysis recombinant plasmids, confirm to have dna fragmentation to insert plasmid according to electrophoresis result, its size is about 3.5kb.The recombinant plasmid that contains this dna fragmentation is called pGLUB, and the recombination bacillus coli that contains this recombinant plasmid pGLUB is called bacillus coli DH 5 alpha GLUB.
But this recombinant plasmid pGLU high frequency transformed into escherichia coli is expressed activity of beta-glucosidase and anti-ammonia benzyl performance.DNA in the recombinant plasmid is inserted fragment digoxin dna marker detection kit mark, carry out Southern blot DNA hybrid experiment with the chromosomal DNA of thermophile bacteria MB4, the dna fragmentation that inserts among the confirmation recombinant plasmid pGLUB is from the chromosomal DNA of thermophile bacteria MB4.
Adopt the Sanger dideoxy method that this dna fragmentation is checked order.Sequencing result shows that the insertion fragment contains the open reading frame (ORF) of a long 1578bp, the protein of being made up of 526 amino acid of encoding.Belong to the beta-glucoside enzyme family, the highest similarity is with the beta-glucosidase gene (51%) of Clostridiumacetobutylicum.
Embodiment 2.
The purifying and the characteristic of reorganization beta-glucosidase
The thalline of reorganization bacterium E.coli DH5 α GLUB is suspended from the 50mM acetate buffer solution (pH6.5), utilizes the ultrasonic disruption cell, and centrifuged supernatant is the crude enzyme liquid of reorganization beta-glucosidase.Clear enzyme solution 70oC heating is 15 minutes on this, and centrifugal removal metaprotein, last clear enzyme solution be through ion-exchange chromatography, and hydroxyapatite adsorpting column chromatography and PAGE prepare step such as electrophoresis and carry out purifying, and the zymin that obtains shows a band on SDS-PAGE.Utilize the standard method of known protein materialization to measure the fundamental characteristics of this reorganization beta-glucosidase.The molecular weight of the recombinase that records with SDS-PAGE is 60000 dalton, and is similar to the molecular weight of calculating in theory (59000 dalton); The recombinase operative temperature is 60-80 ℃, pH6-7.
Embodiment 3
Reorganization beta-glucosidase hydrolysis fiber disaccharides
In the cellobiose solution to 2% (using pH6, the sodium phosphate buffer preparation of 0.2M), add 0.1V enzyme liquid, be warming up to 60 ℃ of insulations 6 hours.High pressure lipuid chromatography (HPLC) records recombinant high temperature beta-glucosidase hydrolysis fiber disaccharides and produces glucose.
SEQUENCE?LISTING
＜110〉Institute of Microorganism, Academia Sinica
＜120〉a kind of high temperature beta-glucosidase, its encoding gene and application thereof
<130>IM021103
<160>2
<170>PatentIn?version?3.1
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<213>Thermoanaerobacter?tengcongensis?MB4
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actctttctt?ataatatcct?tacccgcatc?ttaagggaag?aattcaagtt?tgatgggctt 780
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gcgtcaatgg?ctataaaagc?tggagcagat?atagtacttg?tttctcatac?taaagagcta 900
cagataaaag?cttttgaaaa?tataaaagaa?gctgtagaaa?gaggagatat?ctctgtagaa 960
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attgcaaaag?ccataagttt?aaagagcata?acagttgtta?aagatgagaa?aaaattaatc 1140
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gtagaagaaa?ctatttctgt?aaatccaaac?aatagagaaa?tctcagtaat?tctagaaaag 1320
gcaaaggata?aagaagtagt?tgtcgtaggt?acttacaatg?ctaatttaaa?tgaaggtcaa 1380
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aatccttatg?accttcaaaa?gtttgaaaat?gtgcctactt?atgtatgtgc?ttacgagtac 1500
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<210>2
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<213>Thermoanaerobacter?tengcongensis?MB4
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Met?Lys?Lys?Ile?Glu?Glu?Met?Thr?Leu?Glu?Glu?Lys?Ile?Gly?Gln?Met
1 5 10 15
Leu?Met?Ile?Gly?Phe?Pro?Ser?Ala?Phe?Tyr?Asp?Asp?His?Ile?Arg?Glu
20 25 30
Leu?Val?Thr?Ser?Tyr?Lys?Ile?Gly?Asn?Val?Ile?Leu?Phe?Ser?Arg?Asn
35 40 45
Val?Gln?Asp?Ala?Arg?Gln?Val?Met?Asp?Leu?Cys?Thr?Asp?Ile?Gln?Lys
50 55 60
Glu?Ala?Ile?Glu?Asn?Thr?Gly?Ile?Pro?Ala?Phe?Ile?Ser?Ile?Asp?Gln
65 70 75 80
Glu?Gly?Gly?Met?Val?Thr?Arg?Ile?Tyr?Arg?Gly?Ala?Thr?Tyr?Phe?Pro
85 90 95
Gly?Asn?Met?Ala?Ile?Gly?Ala?Thr?Lys?Asp?Pro?Glu?Asn?Ala?Tyr?Lys
100 105 110
Val?Gly?Glu?Ile?Ala?Gly?Arg?Glu?Leu?Arg?Thr?Ile?Gly?Ile?Asn?Ile
115 120 125
Asn?Phe?Ala?Pro?Val?Met?Asp?Val?Asn?Asn?Asn?Pro?Leu?Asn?Pro?Val
130 135 140
Ile?Gly?Val?Arg?Ser?Tyr?Gly?Glu?Asn?Pro?Glu?Asp?Val?Ala?Asn?Phe
145 150 155 160
Gly?Ile?Asn?Tyr?Ile?Lys?Gly?Leu?Gln?Lys?Glu?Gly?Val?Ile?Ala?Thr
165 170 175
Ala?Lys?His?Phe?Pro?Gly?His?Gly?Asp?Thr?Thr?Val?Asp?Ser?His?Leu
180 185 190
Asp?Leu?Pro?Lys?Val?Asn?His?Gly?Lys?Asp?Arg?Leu?Tyr?Glu?Val?Glu
195 200 205
Leu?Tyr?Pro?Phe?Lys?Lys?Ala?Val?Glu?Asn?Gly?Val?Asp?Ala?Ile?Met
210 215 220
Thr?Ala?His?Ile?Leu?Phe?Pro?Ala?Leu?Glu?Asp?Ser?Lys?Val?Pro?Ala
225 230 235 240
Thr?Leu?Ser?Tyr?Asn?Ile?Leu?Thr?Arg?Ile?Leu?Arg?Glu?Glu?Phe?Lys
245 250 255
Phe?Asp?Gly?Leu?Val?Ile?Thr?Asp?Cys?Met?Glu?Met?Asn?Ala?Ile?Ala
260 265 270
Lys?Tyr?Phe?Gly?Thr?Gln?Lys?Ala?Ala?Ser?Met?Ala?Ile?Lys?Ala?Gly
275 280 285
Ala?Asp?Ile?Val?Leu?Val?Ser?His?Thr?Lys?Glu?Leu?Gln?Ile?Lys?Ala
290 295 300
Phe?Glu?Asn?Ile?Lys?Glu?Ala?Val?Glu?Arg?Gly?Asp?Ile?Ser?Val?Glu
305 310 315 320
Arg?Ile?Asp?Glu?Ser?Val?Arg?Arg?Ile?Leu?Lys?Met?Lys?Glu?Lys?Tyr
325 330 335
Gly?Leu?Phe?Asp?Arg?Pro?Tyr?Pro?Glu?Lys?Asp?Lys?Phe?Asp?Thr?Thr
340 345 350
Val?Gly?Ile?Glu?Lys?His?Arg?Lys?Ile?Ala?Lys?Ala?Ile?Ser?Leu?Lys
355 360 365
Ser?Ile?Thr?Val?Val?Lys?Asp?Glu?Lys?Lys?Leu?Ile?Pro?Leu?Lys?Thr
370 375 380
Lys?Asn?Val?Leu?Ala?Ile?Ser?Pro?Glu?Thr?Val?Pro?Val?Ser?Phe?Val
385 390 395 400
Asp?Asp?Ala?Leu?Met?Asp?Arg?Leu?Ser?Phe?Ala?Arg?Gln?Phe?Thr?Ser
405 410 415
Thr?Phe?Gly?Gly?Val?Glu?Glu?Thr?Ile?Ser?Val?Asn?Pro?Asn?Asn?Arg
420 425 430
Glu?Ile?Ser?Val?Ile?Leu?Glu?Lys?Ala?Lys?Asp?Lys?Glu?Val?Val?Val
435 440 445
Val?Gly?Thr?Tyr?Asn?Ala?Asn?Leu?Asn?Glu?Gly?Gln?Val?Lys?Leu?Val
450 455 460
Asn?Ala?Leu?Leu?Gln?Val?Asn?Lys?Asn?Val?Ile?Val?Val?Ala?Leu?Arg
465 470 475 480
Asn?Pro?Tyr?Asp?Leu?Gln?Lys?Phe?Glu?Asn?Val?Pro?Thr?Tyr?Val?Cys
485 490 495
Ala?Tyr?Glu?Tyr?Thr?Pro?Leu?Ser?Val?Glu?Ser?Val?Ile?Gly?Val?Leu
500 505 510
Thr?Gly?Lys?Tyr?Pro?Ala?Glu?Gly?Lys?Leu?Pro?Val?Thr?Ile
515 520 525
Claims (10)
1. beta-glucosidase or its functional analogue that derives from thermophile bacteria (Thermoanaerobacter tengcongensis) MB4, the aminoacid sequence shown in its aminoacid sequence and the SEQ NO.2 has and has 80% homology at least.
2. enzyme according to claim 1 or its functional analogue, the aminoacid sequence shown in its aminoacid sequence and the SEQNO.2 have and have 90% homology at least.
3. enzyme according to claim 1 or its functional analogue, it has the aminoacid sequence shown in the SEQ NO.2.
4. the gene of any described beta-glucosidase or its functional analogue in the claim 1 to 3 of encoding.
5. gene according to claim 4, it has the nucleotide sequence shown in the SEQ NO.1.
6. a recombinant plasmid that contains the described nucleotide sequence of claim 5 is pGLUB.
7. expression carrier that contains claim 4 or 5 described beta-glucosidases or its functional analogue.
8. reorganization bacterium that contains the described expression vector of claim 7.
9. recombination bacillus coli DH5 α GLUB who contains the described expression vector of claim 7.
10. a method steps and purposes for preparing beta-glucosidase comprises:
(1) according to the described reorganization of claim 8 bacterium;
(2) be a kind of reorganization that contains recombinant plasmid pGLUB according to the described reorganization of claim 8 bacterium
Bacillus coli DH 5 alpha GLUB;
(3) the reorganization bacterium prepares the high temperature beta-glucosidase by microbial fermentation and enzyme engineering;
(4) the high temperature beta-glucosidase that obtains by this method is being used aspect chemical industry, weaving, food, the medicine industry.
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100335640C (en) * | 2005-10-28 | 2007-09-05 | 南开大学 | Thermo philic alkali beta glucosidase and its coding gene |
| CN101082047B (en) * | 2006-05-31 | 2010-05-12 | 中国农业大学 | Beta-glucosidase gene promoter and uses thereof |
| CN102268421A (en) * | 2011-08-03 | 2011-12-07 | 山西恩泽生物技术有限公司 | Cloning, expression and application of beta-glucosaccharase gene |
| CN102321647A (en) * | 2011-09-08 | 2012-01-18 | 杭州师范大学 | Beta-glucosidase, coding gene, vector, engineering bacteria and application thereof |
| CN102358898A (en) * | 2011-10-28 | 2012-02-22 | 武汉新华扬生物股份有限公司 | Medium-temperature beta-glucosidase BglA1, gene of same and application of same |
| CN102899300A (en) * | 2011-07-28 | 2013-01-30 | 中国科学院上海生命科学研究院 | Novel high-temperature beta-glucosidase, its coding gene and application |
| CN101392242B (en) * | 2007-09-18 | 2013-04-10 | 中国科学院微生物研究所 | Alpha-glucosidase, gene thereof, preparation method, vector and host cell |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US8512714B2 (en) * | 2006-05-22 | 2013-08-20 | Biogasol Ipr Aps | Thermoanaerobacter mathranii strain BG1 |
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2002
- 2002-11-19 CN CN 02148776 patent/CN1245507C/en not_active Expired - Fee Related
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100335640C (en) * | 2005-10-28 | 2007-09-05 | 南开大学 | Thermo philic alkali beta glucosidase and its coding gene |
| CN101082047B (en) * | 2006-05-31 | 2010-05-12 | 中国农业大学 | Beta-glucosidase gene promoter and uses thereof |
| CN101392242B (en) * | 2007-09-18 | 2013-04-10 | 中国科学院微生物研究所 | Alpha-glucosidase, gene thereof, preparation method, vector and host cell |
| CN102899300A (en) * | 2011-07-28 | 2013-01-30 | 中国科学院上海生命科学研究院 | Novel high-temperature beta-glucosidase, its coding gene and application |
| CN102899300B (en) * | 2011-07-28 | 2014-12-10 | 中国科学院上海生命科学研究院 | Novel high-temperature beta-glucosidase, its coding gene and application |
| CN102268421A (en) * | 2011-08-03 | 2011-12-07 | 山西恩泽生物技术有限公司 | Cloning, expression and application of beta-glucosaccharase gene |
| CN102321647A (en) * | 2011-09-08 | 2012-01-18 | 杭州师范大学 | Beta-glucosidase, coding gene, vector, engineering bacteria and application thereof |
| CN102321647B (en) * | 2011-09-08 | 2012-11-21 | 杭州师范大学 | Beta-glucosidase, coding gene, vector, engineering bacteria and application thereof |
| CN102358898A (en) * | 2011-10-28 | 2012-02-22 | 武汉新华扬生物股份有限公司 | Medium-temperature beta-glucosidase BglA1, gene of same and application of same |
| CN102358898B (en) * | 2011-10-28 | 2013-01-09 | 武汉新华扬生物股份有限公司 | Medium-temperature beta-glucosidase BglA1, gene of same and application of same |
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| CN1245507C (en) | 2006-03-15 |
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