CN105462863A - Alkaline-lipase-producing Aspergillus niger mutant strain - Google Patents
Alkaline-lipase-producing Aspergillus niger mutant strain Download PDFInfo
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- CN105462863A CN105462863A CN201510996611.0A CN201510996611A CN105462863A CN 105462863 A CN105462863 A CN 105462863A CN 201510996611 A CN201510996611 A CN 201510996611A CN 105462863 A CN105462863 A CN 105462863A
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- aspergillus niger
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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
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
- C12N9/18—Carboxylic ester hydrolases (3.1.1)
- C12N9/20—Triglyceride splitting, e.g. by means of lipase
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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
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
- C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
- C12N9/2405—Glucanases
- C12N9/2408—Glucanases acting on alpha -1,4-glucosidic bonds
- C12N9/2411—Amylases
- C12N9/2414—Alpha-amylase (3.2.1.1.)
- C12N9/2417—Alpha-amylase (3.2.1.1.) from microbiological source
- C12N9/242—Fungal source
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y301/00—Hydrolases acting on ester bonds (3.1)
- C12Y301/01—Carboxylic ester hydrolases (3.1.1)
- C12Y301/01003—Triacylglycerol lipase (3.1.1.3)
Abstract
The invention aims to provides an alkaline-lipase-producing Aspergillus niger mutant strain which is prepared by the following steps: transforming an alkaline lipase gene into an Aspergillus niger host strain to construct an Aspergillus niger engineering strain capable of efficiently expressing alkaline lipase; and knocking out the amylase gene of the host strain by gene knock-out means, and screening to obtain the mutant strain with obviously enhanced alkaline lipase yield. The collection number of the mutant Aspergillus niger strain is CCTCC NO:M2015761. The alkaline lipase activity of the mutant strain Aspergillus niger Su-12 reaches 15766 u/ml, and the protein expression level is 5.4 g/L, which are respectively enhanced by 56.3% and 18.9% as compared with the strain before mutation, thereby being beneficial to wide application of the enzyme.
Description
Technical field
The invention belongs to gene engineering technology field, be specifically related to a kind of aspergillus niger mutant strain producing alkaline lipase.
Technical background
Lipase is the enzyme that a class has multiple catalytic capability, can the hydrolysis of catalysis triglyceride and some other water-insoluble ester class, alcoholysis, esterification, transesterification and ester class reverse reaction reaction, in addition the activity of some other enzyme is also shown, as Phospholipid hydrolase, lysophospholipase, Sterol esterase, acylpetide hydrolase activity etc.Lipase is with a wide range of applications, and has become the third-largest industrial enzymes on market.Lipase can the reaction such as catalysis solution fat, transesterify, Lipase absobed, is widely used in the industry such as fodder additives, fats and oils processing, food, medicine, daily use chemicals.
Alkaline lipase is the lipase be hydrolyzed in the basic conditions, and the main raw material of production is the agricultural byproduct such as wheat, corn, is transformed by biofermentation technique.It can be hydrolyzed natural fats and oils, produces lipid acid and glycerine, is a kind of enzyme being hydrolyzed special esters class specially on outphasing system profit interface.
Current alkaline lipase is mainly used in detergent industry, and separately or be added on together with Sumizyme MP in washing composition, as a kind of added ingredients of washing composition, it contributes to the removal of fatty oil stain and human sebaceous's dirt.1988, NoVo company of Denmark reported the application of alkaline lipase in washing composition, and is put in suitability for industrialized production.Lion grease company of Japan is also proposed the washing powder of lipase the same year.The joint ventures such as P & G company and Unilever are also proposed the high-grade washing powder of fatty enzyme and proteolytic enzyme.The domestic research to alkaline lipase is started late, very large gap is had with external modern technique in alkaline lipase fermented bacterium, extraction process, enzymic activity and granzyme technology of preparing etc., especially the production cost of product is far above same kind of products at abroad, lack competitiveness, so be badly in need of now exploitation alkaline lipase superior strain, promote the widespread use of alkaline lipase.
Summary of the invention
The object of this invention is to provide aspergillus niger mutant strain and application thereof that alkaline lipase is produced in a strain, be that the alkaline lipase gene transformation deriving from Thermomyces lanuginosus (Thermomyceslanuginosus) is entered in aspergillus niger Host Strains, build the aspergillus niger engineering bacteria of high expression alkaline lipase; Then the amylase gene of Host Strains self is knocked out by gene knockout means, then the mutant strain that screening acquisition alkaline fat production of enzyme significantly improves, for the widespread use of this enzyme is laid a good foundation.
One aspect of the present invention provides a kind of aspergillus niger engineering bacteria, and it carries the recombinant plasmid of expressing alkaline lipase gene.
Described alkaline lipase, its aminoacid sequence is SEQIDNO:1; The sequence of its encoding gene is SEQIDNO:2.
One aspect of the present invention provides plant mutant bacterium aspergillus niger Su-L2 (AspergillusnigerSu-L2), be preserved in the China typical culture collection center of Wuhan, China Wuhan University on December 16th, 2015, deposit number is CCTCCNO:M2015761.
Aspergillus niger Su-L2 of the present invention is producing the application in alkaline lipase.
The aspergillus niger engineering bacteria Su-L1 that the present invention builds, under 30L ferment tank condition, its alkaline fat enzyme activity reaches 10089u/ml, and expressing quantity is 4.54g/L.In order to improve its enzymatic productivity further, the present invention knocks out the amylase gene of Host Strains self by gene knockout means, the mutant strain aspergillus niger Su-L2 that screening obtains, its alkaline fat enzyme activity reaches 15766u/ml, expressing quantity is 5.4g/L, improve 56.3% and 18.9% respectively than before sudden change, be conducive to the widespread use of this enzyme.
Accompanying drawing explanation
Fig. 1 is plasmid pGAU collection of illustrative plates;
Fig. 2 is plasmid pMD18T-pyrG collection of illustrative plates.
Embodiment
The present invention has used routine techniques and the method for genetic engineering and biology field use, such as MOLECULARCLONING:ALABORATORYMANUAL, 3ndEd. (Sambrook, 2001) method and described in CURRENTPROTOCOLSINMOLECULARBIOLOGY (Ausubel, 2003).These general reference provide definition well known by persons skilled in the art and method.But those skilled in the art on the basis of the technical scheme described in the present invention, can adopt the method for other routine of this area, experimental program and reagent, and is not limited to the restriction of the specific embodiment of the invention.
Describe the present invention below in conjunction with embodiment.
The acquisition of embodiment 1 alkaline lipase gene
According to the gene order in public gene database, synthesize alkaline lipase gene LipN (sequence is SEQIDNO:2) by Shanghai JaRa company, the aminoacid sequence of its coding is SEQIDNO:1.
PCR primer and reaction conditions as follows:
Primer 1 (F): ATGCGGTCCTCCCTGGTGCTG
Primer 2 (R): TCACAGACAGGTGCCGATCAG
Reaction conditions is: 94 DEG C of sex change 5min; Then 94 DEG C of sex change 30s, 56 DEG C of renaturation 30s, 72 DEG C extend 45s, after 30 circulations, 72 DEG C of insulation 10min.Agarose electrophoresis result shows, and LipN gene size is 876bp.
Embodiment 2 construction of recombinant vector
Pcr amplification alkaline lipase gene, XbaI site is introduced at primer two ends.Primer sequence is as follows:
Primer 3 (F): GC
tCTAGAaTGCGGTCCTCCCTGGTGCTG
Primer 4 (R): GC
tCTAGAtCACAGACAGGTGCCGATCAG
PCR reaction conditions is: 94 DEG C of sex change 5min; Then 94 DEG C of sex change 30s, 56 DEG C of renaturation 30s, 72 DEG C extend 45s, after 30 circulations, 72 DEG C of insulation 10min.Agarose gel electrophoresis result shows, the fragment of 876bp sized by LipN gene.
The alkaline lipase LipN fragment of above-mentioned acquisition and expression vector pGAU are carried out restriction enzyme XbaI single endonuclease digestion respectively, and enzyme tangent condition is as follows:
37 DEG C of water-bath enzymes cut process 2h, reclaim two object fragments respectively, be dissolved in 20ulddH after electrophoresis
2o.Connect with T4DNA ligase enzyme, linked system is as follows:
22 DEG C connect 1h, transformation of E. coli DH5a competence, and coating LB+AMP is dull and stereotyped, single bacterium colony is grown after 37 DEG C of overnight incubation, bacterium colony PCR verifies that connecting correct transformant extraction plasmid send order-checking, after order-checking is correct, namely obtains the recombinant vectors pGAU-LipN containing alkaline lipase gene LipN.
Embodiment 3 alkaline lipase LipN's is recombinant expressed
Prepared by protoplastis: inoculated aspergillus niger host Su2-1 is dull and stereotyped in PDA+U, cultivates 5-7d for 30 DEG C.Extract the bacterium block of 2cm × 2cm size, in inoculation 100ml liquid PDA+U substratum, cultivate 24h for 30 DEG C and grow mycelium, for transforming.After the mycelium grown is filtered, resuspended with 20ml1.2M Adlerika, add 0.2g N,O-Diacetylmuramidase.30 DEG C, 100rpm cultivates 2-3h.Filtered by cracked mycelia 2 layers of lens wiping paper, the centrifugal 10min of 3000rpm obtains protoplastis.
Transform: protoplastis 1.2M Sorbitol Solution USP cleans 2 times, then uses appropriate Sorbitol Solution USP resuspended, makes protoplast concentration reach 10
8.Add the ready plasmid of 10ul in 200ul protoplastis, add the PEG6000 of 50ul25%, ice bath 20min, then the PEG6000 room temperature adding 2ml25% places 5min, adds 4ml Sorbitol Solution USP and puts upside down mixing.After pouring 50ml conversion upper strata substratum into, pour into 4 and transform in lower floor's flat boards, after the culture medium solidifying of upper strata, be inverted in 30 DEG C of incubators and cultivate 5d.
Transformant screening: after cultivating 5d, the bacterium colony that picking grows, dibbling carries out multiple sieve to conversion lower floor flat board, cultivates 2d for 30 DEG C.The transformant of normal growth is inoculated into respectively fresh PDA dull and stereotyped, cultivates 5-7d for 30 DEG C.Each transformant extracts the bacterium block of 2cm × 2cm size, inoculates respectively in 50ml liquid submerged culture base and ferments, and cultivate 5d for 32 DEG C, every day adds proper ammonia, and control pH is about 4.5.After cultivating 5d, centrifugal thalline obtains supernatant liquor and is crude enzyme liquid, carries out protein electrophoresis detection, and the recombinant bacterial strain filtering out the expression of obvious protein band is positive transformant.
By one of them positive transformant called after aspergillus niger Su-L1 (AspergillusnigerSu-L1).
Embodiment 4:U deficient strain is screened
Above-mentioned aspergillus niger Su-L1 is seeded to PDA flat board, cultivates 7d for 30 DEG C, draw 1ml sterilized water wash-out spore, dilution certain multiple makes spore concentration 1 × 10
5individual/ml.Coating BT flat board (adding the 5-fluororotic acid of final concentration 1.5g/L in PDA, the uridine of 1g/L), cultivates 3-5d, the colony inoculation PDA+U flat board (adding the uridine of final concentration 1g/L in PDA) that picking grows for 30 DEG C, cultivates 3-5d for 30 DEG C.Namely the bacterial strain of U defective type is obtained.
Embodiment 5 knocks out the amylase gene of Host Strains
Design primer pcr amplification from aspergillus niger host Su2-1 genome obtains the upstream and downstream homology arm AU (sequence is SEQIDNO:3) of amylase gene, AD (sequence is SEQIDNO:4).
PCR primer and reaction conditions as follows:
Primer 5 (AUF): GCTGCGGAAGCCGAATGTGAC
Primer 6 (AUR): CCCAAACACAGCCTGACTACA
Primer 7 (ADF): AGGGGATGTAGGTTTGTTGGT
Primer 8 (ADR): TACAGTCTGGCAATTGATCCA
Reaction conditions is: 94 DEG C of sex change 5min; Then 94 DEG C of sex change 30s, 56 DEG C of renaturation 30s, 72 DEG C extend 90s, after 30 circulations, 72 DEG C of insulation 10min.Agarose electrophoresis result shows, and homology arm AU size is 1761bp, and homology arm AD size is 1698bp.
Homology arm AU, AD are connected to the two ends of pyrG mark in knockout carrier pMD18T-pyrG by SphI, KpnI, build knockout carrier p Δ A.
According to method in embodiment 3, by the U deficient strain of knockout carrier p Δ A transform A. niger Su-L1, screening obtains transformant, is verified by primer 9,10.The bacterial strain knocking out amylase gene can not increase object band, and the bacterial strain not knocking out amylase gene amplifies 1032bp fragment.
Primer 9 (AF): AGTATGGTATTGCTGTACTTC
Primer 10 (AR): GTAGAGGTTGCTGATGCTGCC
The Aspergillus niger strain that a strain alkaline fat production of enzyme significantly improves is screened, called after aspergillus niger Su-L2 ((AspergillusnigerSu-L2) from the bacterial strain that amylase gene knocks out.On December 16th, 2015 mutant strain aspergillus niger Su-L2 (AspergillusnigerSu-L2) has been preserved in the China typical culture collection center of Wuhan, China Wuhan University, deposit number is CCTCCNO:M2015761.
Embodiment 6 alkaline lipase enzyme activity determination
(1) definition of alkaline lipase Mei Huo unit
40 DEG C, under pH value is the condition of 7.5, the enzyme amount required for titratable lipid acid that 1min hydrolysis substrate produces 1 μm of ol is an enzyme activity unit U.
(2) substrate solution
Weigh polyvinyl alcohol (PVA: the polymerization degree 1750 ± 50) 40g, add water 800ml, soaks 4-6h, heat in boiling water bath, stirs, until all dissolve, is settled to 1000ml after stirring cooling, and by clean 6-8 layer filtered through gauze, it is for subsequent use to get filtrate.
Measure above-mentioned filtrate 150ml, add sweet oil 50ml, with high-speed homogenization machine process 10min (point 4 process, interval 5min, processes 2-3min) at every turn, obtain oyster white PVA emulsion.Matching while using.
(3) enzyme activity determination
Get two 100ml triangular flasks, substrate solution 4ml and pH7.5 phosphoric acid buffer 5ml is respectively added respectively in blank bottle (A) and sample bottle (B), 95% ethanol 15ml is added again, preheating 5min in 40 DEG C of water-baths in blank bottle (A);
Enzyme liquid 1ml to be measured is respectively added in blank bottle (A) and sample bottle (B), mix timing immediately, after accurate response 15min, use a point sample pipettor in sample bottle (B), add 95% ethanol 15ml termination reaction immediately, take out;
Above-mentioned reaction solution is poured in 50ml beaker, in triangular flask, add 5ml pure water, pour into after shaking up in 50ml beaker, add 2 phenolphthalein indicators;
Use the pH meter correcting parlkaline condition, under the condition stirred, add the sodium hydroxide solution of 0.05mol/L in beaker, being titrated to pH value is 9.92; Be titrated to pH value and be no longer changed to terminal at 20s, record consumes the volume of standard solution of sodium hydroxide.
X---alkaline fat enzyme activity, u/ml;
V1---consume standard solution of sodium hydroxide volume during titration sample, ml;
Standard solution of sodium hydroxide volume is consumed, ml when V2---titration is blank;
The concentration of C---standard solution of sodium hydroxide, mol/L;
50---0.05mol/L sodium hydroxide solution 1ml is equivalent to lipid acid 50 μm of ol;
N---enzyme liquid extension rate;
0.05---Concentration of Sodium Hydroxide Solution Standard reduction factor;
15---time scale factor.
Embodiment 730L ferment tank is verified
30 liters of fermentor tanks carry out the fermentation of above-mentioned aspergillus niger Su-L1 and mutant bacteria aspergillus niger Su-L2 thereof, and the culture medium prescription that fermentation uses is: maltose 2.5g/L, ammonium sulfate 0.9%, Sodium phosphate dibasic 0.15g/L, calcium chloride 0.1g/L, potassium sulfate 0.5g/L, magnesium sulfate 16.4g/L, Trisodium Citrate 0.2g/L, potassium primary phosphate 0.25%, bean powder 0.5%, defoamer 0.05%.Supplemented medium: maltose concentration is that 400g/L, pH are adjusted between 4.0-5.0.
Fermentation manufacturing technique: pH value 4.0, temperature 30 DEG C, stir speed (S.S.) 300-700rpm, ventilation 1.0-1.5 (v/v), dissolved oxygen control more than 20%.
Whole fermenting process is divided into three phases: the first stage is the yeast culture stage, in 7% ratio access seed, cultivates 15-25h for 30 DEG C, with dissolved oxygen rise for mark; Subordinate phase is the hungry stage, and after sugar has consumed at the end, stream does not add any carbon source, and dissolved oxygen rises to more than 60% and shows that this stage terminates, and schedules to last about 30-120min; Flow feeding substratum carried out product enzyme and cultivated in order to produce the enzyme stage phase III, and period keeps dissolved oxygen more than 20% and maintain concentration of reduced sugar in fermented liquid being not less than 1g/L, and fermentation period is between 140-170h.After fermentation ends, fermented liquid, by obtaining crude enzyme liquid after flame filter press process, carries out alkaline lipase Enzyme activity assay to crude enzyme liquid.
Detected result shows: under 30L ferment tank condition, aspergillus niger Su-L1 fermented liquid neutral and alkali lipase activity is 10089u/ml, and expressing quantity is 4.54g/L; And aspergillus niger Su-L2 fermented liquid neutral and alkali lipase activity is up to 15766u/ml, expressing quantity is 5.4g/L, improves 56.3% and 18.9% respectively than before sudden change.
Claims (6)
1. an aspergillus niger engineering bacteria, is characterized in that, described aspergillus niger engineering bacteria carries the recombinant plasmid of expressing alkaline lipase gene.
2. aspergillus niger engineering bacteria as claimed in claim 1, it is characterized in that, described alkaline lipase, its aminoacid sequence is SEQIDNO:1.
3. aspergillus niger engineering bacteria as claimed in claim 2, it is characterized in that, the sequence of the encoding gene of described alkaline lipase is SEQIDNO:2.
4. a mutant strain for aspergillus niger engineering bacteria according to claim 1, is characterized in that, described mutant strain is that the novel starch enzyme gene having knocked out aspergillus niger engineering bacteria according to claim 1 obtains.
5. mutant strain as claimed in claim 4, it is characterized in that, described mutant strain is aspergillus niger (Aspergillusniger) Su-L2, and its deposit number is CCTCCNO:M2015761.
6. mutant strain according to claim 5 is producing the application in alkaline lipase.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106591259A (en) * | 2016-11-25 | 2017-04-26 | 山东隆科特酶制剂有限公司 | Alkaline lipase prepared from aspergillus niger and production method of alkaline lipase |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103555596A (en) * | 2013-10-25 | 2014-02-05 | 青岛蔚蓝生物集团有限公司 | Aspergillus niger strain for expression of lipase |
| CN103865896A (en) * | 2014-03-11 | 2014-06-18 | 上海康地恩生物科技有限公司 | Alkaline lipase mutant |
| CN103865949A (en) * | 2014-03-10 | 2014-06-18 | 湖北工业大学 | Method for breeding high-producing strain of cellulase by gene knockout |
| CN104031899A (en) * | 2013-03-05 | 2014-09-10 | 中国科学院天津工业生物技术研究所 | Lipase and application thereof |
| CN104830890A (en) * | 2015-04-30 | 2015-08-12 | 上海泰坦科技股份有限公司 | High yielding cellulase engineering bacterium and application thereof |
-
2015
- 2015-12-25 CN CN201510996611.0A patent/CN105462863A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104031899A (en) * | 2013-03-05 | 2014-09-10 | 中国科学院天津工业生物技术研究所 | Lipase and application thereof |
| CN103555596A (en) * | 2013-10-25 | 2014-02-05 | 青岛蔚蓝生物集团有限公司 | Aspergillus niger strain for expression of lipase |
| CN103865949A (en) * | 2014-03-10 | 2014-06-18 | 湖北工业大学 | Method for breeding high-producing strain of cellulase by gene knockout |
| CN103865896A (en) * | 2014-03-11 | 2014-06-18 | 上海康地恩生物科技有限公司 | Alkaline lipase mutant |
| CN104830890A (en) * | 2015-04-30 | 2015-08-12 | 上海泰坦科技股份有限公司 | High yielding cellulase engineering bacterium and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| 张建军 等: "提高蛋白质在米曲霉中表达量的策略", 《中国生物工程杂志》 * |
| 朱凤妹 等: "β-葡萄糖苷酶基因克隆及在丝状真菌中的高效表达的研究进展", 《食品安全质量检测学报》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106591259A (en) * | 2016-11-25 | 2017-04-26 | 山东隆科特酶制剂有限公司 | Alkaline lipase prepared from aspergillus niger and production method of alkaline lipase |
| CN106591259B (en) * | 2016-11-25 | 2020-06-23 | 山东隆科特酶制剂有限公司 | Alkaline lipase produced from aspergillus niger and production method thereof |
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