44results about How to "Improve enzyme production efficiency" patented technology

The invention relates to a construction method for co-expression hemoglobin VHb and cellulase protein in pichia pastoris, in particular to cellulase protein gene codon optimization and construction of a cellulase protein and vitreoscilla hemoglobin protein (VHb)-co-expressed pichia pastoris system. According to the construction method, codon bias optimization is performed on the nucleotide sequence of EG II (GenBank Accession No.DQ178347.1) through Gene Designer (DNA2.0, Menlo Park, CA, USA) software, a pPIC9K-eg2 expression vector is constructed, and a recombined pichia pastoris strain is obtained by taking Pichiapastoris GS115 as a host through electrotransformation. In addition, the nucleotide sequence of the VHb (GenBank Accession No.M30794.1) is obtained from NCBI and artificially synthesized into a gene, then a pPICZalphaA-vgb expression vector is constructed, and the co-expressed pichia pastoris strain is obtained by taking the recombined pichia pastoris strain containing the EG II gene as a host through electrotransformation. Detection shows that the co-expressed strain is improved on the aspects of bacterial concentration growth and enzyme activity, wherein the OD600 value is increased by 7.2%, and the enzyme activity is improved by 2.2%.

The present invention is production process of UV and NTG mutagenizing Bacillus subtilis TQS6 to culture high efficiency expressing chitosanase strain TQK and fermentation producing high activity chitosanase. The chitosanase strain is first slant cultured and then fermentation cultured while introducing oxygen-rich air to increase dissolved oxygen to 30-80 %; and after culturing for 18-24 hr, average enzyme activity may reach 718 mu / ml. High activity chitosanase may be obtained through fermentation in optimized enzyme producing condition. The process of the present invention can obtain chitosanase with high yield, high stability and high activity, and the produced chitosanase is applied in converting chitosan into chitin oligose effectively and economically.

The invention discloses a bacillus licheniformis and a method for utilizing the sludge as raw material for producing alkaline protease. The bacillus licheniformis is gram-positive bacteria and is shaped like a straight rod, and the collection number is CGMCC 1970. The method for utilizing the bacillus licheniformis for producing the alkaline protease includes (1) the conditioning of a culture medium; (2) shaking culture; (3) the fermentation by a fermentation tank; and (4) the purification of the protease. The bacterial strain of the invention is utilized for producing the alkaline protease, which can not only dispose sludge, but also obtain the alkaline protease product with high additional value and lower cost. The method uses urban waste sludge to replace the traditional food culture medium for producing the alkaline protease product, which not only provides a new way for the resource disposal of urban sludge, but also can greatly reduce the price of the product and promote the industrial production and application of protease preparations.

The invention belongs to the technical field of enzyme engineering and particularly relates to a method for obtaining sucrose isomerase through site-directed mutation so as to improve the product specificity of the sucrose isomerase, and a method for preparing immobilized sucrose isomerase and achieving separation of the immobilized enzyme through a non-aqueous phase catalytic system. The content of isomaltulose in a catalytic product of the sucrose isomerase is substantially increased and increased to 99.16% from 90.28%, and the enzyme preparation process is easy to implement and is high in enzyme producing efficiency; due to the sucrose isomerase immobilization technology, the sucrose isomerase can easily keep high catalytic activity and low enzyme amount loss in the repeated catalysis process, multi-batch continuous use can be performed, operating difficulty can be greatly reduced, and economic cost is saved.

The invention discloses a high-temperature-resistant high-yield cellulase bacillus subtilis and application thereof. A cellulase production strain is separated from surface soil of a corn straw pile in a Jiangjin area of Chongqing, and the preservation number of the bacillus subtilis is CCTCC NO: M 2020002; a strain Z2 with the best enzyme production performance is finally obtained through enrichment culture, a CMC-Na plate experiment, a lignin plate experiment and an enzyme production experiment, and the obtained strain is subjected to 16S rRNA sequence comparison and identification to determine that the strain is bacillus subtilis Z2. By optimizing the culture medium and the culture conditions, the filter paper enzyme activity, the CMC enzyme activity and the beta-glucosidase activity under the optimal enzyme production conditions of 0.800 U / ml, 5.20 U / ml and 2.07 U / ml are obtained respectively. Cellulase produced by the strain has relatively high activity and stability under the conditions of a buffer solution with the pH value ranging from 4.0 to 10.0 and the temperature of 30 DEG C and 80 DEG C. Cellulase produced by the strain is mixed with commercial xylanase, then a saccharification experiment is carried out on pretreated straw stalks, and 84.27 mg / ml of total reducing sugar can be obtained.

The invention aims to construct recombinant pichia pastoris for heterologous expression of EG II, EG IV and EG V proteins. A method comprises the following specific steps: (1) obtaining of trichodermareesei RNA and CDNA: obtaining mycelia by using a solid induction culture medium, and extracting total RNA and cDNA by using a kit; (2) cloning of a target gene: amplifying a target gene by a reasonable primer PCR; (3) constructing an expression vector: taking Ppic9k as a vector, inserting the target gene into the downstream of a promoter AOX1, wherein the 5 ' and 3 ' end enzyme digestion sites are EcoR I and Not I respectively; (4) obtaining of a recombinant strain: taking bgl I as a linearization site of an expression vector to realize linearization and electrotransformation of the expression vector to obtain recombinant yeast; (5) induction and enzyme production of the recombinant strain: carrying out shaking flask fermentation, and inducing enzyme production by using appropriate methanol; and (6) recombinant protein activity detection: detecting the expression and activity of recombinant protein by SDS-PAGE and Congo red-CMC. The recombinant strain constructed by the method can efficiently express the protein with a his label, and a high-efficiency single enzyme component can be obtained.

The invention relates to a bacterial strain for producing a gel breaking enzyme and an application thereof. The bacterial strain for producing the gel breaking enzyme is the Bacilluslichenformis M1-4 bacterial strain, and the preservation number is CGMCC NO.4511. The M1-4 bacterial strain is used for producing the gel breaking enzyme and preparing a bacteria liquid preparation. The bacterial strain obtained by the invention is high in enzyme production efficiency, and the enzyme output under the industrial fermentation condition is greater than 240 U / ml. The recovery method employed by the invention is relatively high in enzyme yield. The produced fracturing fluid gel breaking enzyme has a high activity within a wide range of temperature and is convenient in operation and less in usage amount. The fracturing fluid gel breaking enzyme has a rapid and even viscosity breaking property which enables the enzyme to completely degrade a guar gum with fewer residues, and the viscosity of a fracturing liquid after gel breaking is less than 5.0 mPa.s. Based on the characteristics of the gel breaking enzyme for cleaning fracturing fluid, the disclosed bacterial strain in the invention can not only be used for the conventional fracturing gel breaker construction operation, but also can be used for a remedial measure of a fracturing operation failure.

The invention relates to an inulase mutant and a preparing method. According to the inulase mutant and the preparing method, directed evolution is carried out on an encoding gene inuI of inulase in vitro with a codon optimization technology, an error-prone PCR technology and a site-directed mutagenesis technology, and the mutant with the inulase activity remarkably improved is screened out. The K<cat> value of the mutant is 3.1 times that of the inulase of an original strain; in addition, the preparing process of the inulase is easy to implement and high in enzyme production efficiency, and the enzyme activity of fermentation liquor in a 30-m<3> system can be 60,000 U / ml. Meanwhile, the invention discloses a DNA sequence, an expression vector and a host cell of the encoding inulase mutant. According to the inulase mutant, the preparing method and the DNA sequence, the expression vector and the host cell of the encoding inulase mutant, high efficiency and simple and convenient operability are achieved.

The invention discloses a method for increasing the output of alkaline pectinase produced by fermentation by adding sorbitol, relating to the technical field of application of the carbon source adding policy in the optimization of fermentation. In the method, recombinant Pichia pastoris GS115 is adopted as fermentation strains, and sorbitol is added into the fermentation liquid at a constant speed or accelerated speed during fermentation induction phase to improve the activity of cells, weaken the toxicity of methanol, and increase the output of the alkaline pectinase. The lower adding amount of sorbitol can not inhibit the enzyme production effect of methanol to the strains, on the other hand, the toxicity to the strains from the methanol can be reduced to a certain extent, and the efficiency of enzyme production is further increased. When the sorbitol is added at a varying speed (0.9g / h-7.2g / h), the activity of enzyme is increased from 930U / ml to 1541U / ml. The invention is simple and effective in process, capable of being applied in the industrial production of pectinase in the future, and instructive to the optimization of fermentation of other Pichia pastoris.

The invention relates to an inulase mutant and a preparing method. According to the inulase mutant and the preparing method, directed evolution is carried out on an encoding gene inuI of inulase in vitro with a codon optimization technology, an error-prone PCR technology and a site-directed mutagenesis technology, and the mutant with the inulase activity remarkably improved is screened out. The K<cat> value of the mutant is 3.1 times that of the inulase of an original strain; in addition, the preparing process of the inulase is easy to implement and high in enzyme production efficiency, and the enzyme activity of fermentation liquor in a 30-m<3> system can be 60,000 U / ml. Meanwhile, the invention discloses a DNA sequence, an expression vector and a host cell of the encoding inulase mutant. According to the inulase mutant, the preparing method and the DNA sequence, the expression vector and the host cell of the encoding inulase mutant, high efficiency and simple and convenient operability are achieved.

The invention belongs to the technical field of microorganisms and discloses a compound microorganism culture for preparing carotenoid. The compound microorganism culture comprises bacillus subtilis,rhodopseudomonas capsulate and rhodotorula glutinis. The compound microorganism culture increases yield of carotenoid and lowers culture cost.

The invention discloses a fungus-derived laccase gene, and relates to the technical field of biology. The laccase gene is lac1, and the cDNA sequence of the lac1 is as shown in SEQ ID NO.2, or the lac1 has a base sequence for encoding an amino acid sequence as shown in SEQ ID NO.3. The invention also provides a recombinant pichia pastoris engineering strain carrying a laccase gene expression vector and an application of the recombinant pichia pastoris engineering strain. The fungus-derived laccase and the recombinant pichia pastoris engineering bacteria thereof have the beneficial effect that the laccase Lac1 newly identified in Trametes sp. Is expressed in a heterologous manner. The obtained recombinant laccase has good thermal stability and good tolerance to metal ions, organic solvents and inhibitors, can catalyze decoloration of indigo blue and other dyes, and has a good application prospect in industry.

The invention provides instant berry powder and a making method thereof and belongs to the field of food processing. The instant berry powder is prepared from the following raw materials in parts by weight: 35-40 parts of blueberry, 25-30 parts of mulberry, 10-15 parts of raspberry, 10-12 parts of strawberry, 6-10 parts of gooseberry, 3-5 parts of ficus carica, 8-12 parts of grape, 3-5 parts of kiwi fruit, 4-6 parts of carambola, 5-8 parts of papaya, 3-5 parts of psidium guajave and 4-8 parts of passiflora edulis. The making method comprises the following steps: freezing the raw materials, pressing, then adding an active substance for enzymatic hydrolysis, filtering an enzymatic hydrolysate and mixing with an auxiliary material and maltodextrin, wherein the auxiliary material is prepared by radix puerariae, poria cocos, radix angelicae, semen nelumbinis and peppermint leaves; drying to obtain the instant berry powder. The instant berry powder provided by the invention is powdery and granular, convenient to eat, better in dissolubility and mouth feeling, specific in berry aroma and sweetness, rich in nutrition and difficult for moisture absorption in air and has the healthcare effects of resisting aging, reducing blood pressure and the like.

The invention relates to a composition for catalyzing degradation of uric acid in intestinal tracts. The composition comprises cordyceps militaris powder or dry enzyme powder, which contains urate oxidase which can be used for catalyzing degradation of uric acid. The invention further provides a preparation method and application of the composition for catalyzing degradation of uric acid in intestinal tracts. According to the composition, uric acid is catalyzed to be degraded into allantoin, the content of uric acid in intestinal tracts can be reduced, the proportion of uric acid, secreted andexcreted to the intestinal tracts, in blood can be increased, the concentration of uric acid in the blood can be reduced, and hyperuricemia and gout diseases can be prevented and treated; and excretion of uric acid in urea can be reduced, and the incidence rate of uric acid calculus can be reduced.

The invention belongs to the technical field of biology and discloses a method for producing carotenoid. The method comprises the following steps: centrifuging a compound microorganism culture; removing supernatant and collecting thalli; then adding acetone and carrying out ultrasonic wall breaking; then standing for 30min; then centrifuging and collecting supernatant and sediment; adding acetonewith the volume the same as that of the sediment into the sediment; carrying out ultrasonic treatment and collecting supernatant; combining the supernatant of the two times; then evaporating and drying at low temperature by utilizing a rotary evaporator; dissolving residues with ethyl ether; then adding a KOH methanol solution and standing for 10 to 30min; then adding a NaCl solution and layeringa solvent; collecting an upper-layer ethyl ether phase and evaporating and drying the ethyl ether phase to obtain a carotenoid product. According to the method disclosed by the invention, the purity of a prepared carotenoid product is relatively high and reaches 94 percent or above, and is converted to the yield which reaches 90 percent or above.

The invention discloses a method for inducing coriolopsis gallica to express laccase by using food waste. According to the method, the food waste is used as a fermentation substrate, provides a carbon source, a nitrogen source and a biological induction factor for the coriolopsis gallica, and promotes mycelial growth and laccase induced expression; and the method mainly comprises the following steps: (1) pretreatment of food processing waste; (2) preparation of a coriolopsis gallica seed solution; (3) preparation of a liquid enzyme production culture medium; and (4) production of the laccase through liquid fermentation. The method for inducing the coriolopsis gallica to express the laccase by using the food waste, which is provided by the invention, is high in enzyme production efficiency, short in fermentation period and low in cost, does not need to additionally add any chemical inducer, meanwhile, is beneficial for promoting the resource utilization of the food waste, and has obvious economic benefits and social benefits.

The invention provides a method for producing neutral phytase through immobilized bacillus fermentation. The method comprises the following steps: (1) preparing a seed solution; (2) carrying out pretreatment of a carrier: modifying a ceramsite carrier with an alkali liquor; (3) preparing immobilized cells: filling the carrier into a fixed bed reactor, and immobilizing thallus cells on the carrierthrough an adsorption effect; (4) carrying out pre-culturing: culturing the immobilized cells for 30-40 hours by using a fermentation culture medium; and (5) carrying out continuous fermentation culture: enabling the fermentation culture medium to flow into a fixed bed reactor at a certain dilution rate for continuous fermentation. The bacillus is immobilized by adopting the alkali-modified carrier, and the method has the advantages of being low in cost, stable in property, non-toxic and good in adsorption effect. According to the method, the immobilized cell technology is combined with the continuous fermentation process, so that the purpose of continuously producing neutral phytase at high intensity is achieved, the fermentation efficiency and the phytase yield are greatly improved, andindustrial production is facilitated.

The invention discloses Sartorya Vuill W-10 with a high cellulase yield. A preservation number of the Sartorya Vuill W-10 is CGMCC No.11991. By fermenting the Sartorya Vuill W-10 disclosed by the invention, high-yield cellulose crude enzyme liquid can be obtained; cellulase filter paper enzyme activity can reach 1.26U / ml; the cellulase obtained from fermentation can be applied to preparing bioethanol, a straw residue enzymolysis rate and an ethanol yield can be effectively improved, and certain application value is achieved. The Sartorya Vuill W-10 disclosed by the invention not only has low production cost, high ethanol yield and favorability for commercial application and implementation of biological ethanol, but also provides a way for comprehensively utilizing biological waste.

The invention relates to the technical field of microorganisms, in particular to bacillus velezensis and an application thereof. The preservation number of the bacillus velezensis Lpl-wx is CGMCC No.17045. The bacterial strain can be applied to preparation of astaxanthin esterases. The bacillus velezensis belongs to prokaryotes, so that the fermentation process is quick, enzyme production efficiency is high, and the astaxanthin esterases can be quickly and efficiently obtained. The bacterial strain can be further applied to preparation of an astaxanthin single body and can effectively degradeastaxanthin esters, the astaxanthin single body can be obtained, a green efficient catalyzing way few in by-products is provided, and a technical basis is provided for preparation and promotion of theastaxanthin single body.

The invention provides an excrement treatment composition, a preparation method and application thereof, and a method for treating anhydrous ecological toilet excrement by using the excrement treatment composition. The excrement treatment composition comprises a complex microbial inoculant and a biological padding, the complex microbial inoculant comprises, by mass, 1-3 parts of aspergillus, 3-7 parts of bacillus subtilis, 5-9 parts of saccharomycetes and 1-4 parts of trichoderma, and the biological padding comprises crop straw. By combining the characteristics of the biological padding and the excrement, the preparation of the complex microbial inoculant is optimized, organic macromolecular substances such as protein in the excrement can be efficiently decomposed, the fermentation periodis shortened, and the frequency of cleaning toilets is reduced. When the excrement treatment composition is used for excrement treatment, ammonia gas and other peculiar smells can be effectively removed, and the excrement treatment composition is suitable for indoor use and has important significance in application and popularization of waterless ecological toilets.

The invention relates to a preparation technology of isomaltulose, and specifically to a preparation technology for producing isomaltulose. The preparation technology comprises the following steps ofpreparing mother liquid, preparing immobilized enzyme-producing bacteria, preparing crude enzyme liquid and isomaltulose mother liquid, and preparing an isomaltulose finish product with the purity being greater 95%; the preparation technology for producing isomaltulose provided by the invention abandons the traditional technology of directly producing isomaltose through microbial fermentation (lowyield), produces sucrose isomerase through microbial immobilized fermentation, enables the sucrose isomerase to perform enzymolysis on sucrose in vitro, wherein the percent conversion reaches 95% orabove, the production cost is reduced, a product completely meets the newest international standards and Chinese national standards, and the production cycle is shortened.

The invention discloses a Sartorya Vuill W-10 with high cellulase production, with a preservation No. CGMCC No. 11991. The invention can obtain high-yield cellulose crude enzyme liquor by fermentationof Sartorya Vuill W 10, wherein the filter paper enzyme activity of the cellulase can reach 1.26 U / ml; The cellulase from fermentation can be used in the production of bioethanol, which can effectively improve the enzymatic hydrolysis rate of straw residue and ethanol yield, and has a certain application value. The invention not only has low production cost and high ethanol yield, which is favorable for commercial application and implementation of bioethanol, but also provides a way for comprehensive utilization of waste organisms.

The invention discloses a transglutaminase fermentation culture medium pretreated by acidolysis or enzymolysis and a preparation method and application thereof. According to the method, fish meal components in the culture medium are separately subjected to acid hydrolysis and enzymatic hydrolysis, correspondingly nutrient components in the fish meal components can be utilized by a transglutaminaseproducing strain, the utilization rate of the nutrient components by the transglutaminase producing strain is increased, and the enzyme production efficiency is thus improved. The method has the advantages of simple and convenient operation, high efficiency, low cost and easy application in industrial production.

The invention provides a processing method and application of fish meal in a glutamine transaminase fermentation culture medium, discloses a fish meal hydrolysis method and application of the method in fermentative production of glutamine transaminase, and mainly solves the problems of insufficient fish meal hydrolysis and low fish meal utilization rate of an existing fish meal hydrolysis process.Improvement is performed on the basis of an existing fish meal hydrolysis process, wherein the fish meal is subjected to pulverization, impurity removal, acid hydrolysis, alkali hydrolysis, enzymatichydrolysis and the like so that nutrients in the fish meal can be better utilized, and the fish meal utilization rate is increased. The hydrolysis process is also applied to the fermentative production of the glutamine transaminase, which can significantly improve the utilization rate of the fish meal by glutamine transaminase bacteria, and the produced glutamine transaminase has the advantages of high enzyme activity, high stability and the like. The method is simple, convenient, efficient, low in cost and easy to use in industrial production.

The invention relates to the technical field of microorganisms, in particular to a bacillus velei and its application. The preservation number of Bacillus Velez Lpl-wx in the present invention is CGMCC No.17045. The bacterial strain can be applied to the preparation of astaxanthin esterase, and because the Bacillus velaisi belongs to prokaryote, the fermentation process is fast and the enzyme production efficiency is high, and the astaxanthin esterase can be obtained quickly and efficiently. The strain can be further applied to the preparation of astaxanthin monomers, which can effectively degrade astaxanthin esters and obtain astaxanthin monomers, providing a green and efficient catalytic pathway with less by-products, which is a good catalyst for the production of astaxanthin monomers. Preparation and promotion provide the technical basis.

The invention discloses a pichia pastoris engineering bacterium for heterologous expression of a cellulase gene EGV and application of the pichia pastoris engineering bacterium. The engineering bacterium is Pichia pastoris-eg5 and is preserved in China General Microbiological Culture Collection Center (CGMCC) in August 2019, with the preservation number of CGMCC No.18422. An endoglucanase gene (EGV) is obtained from trichoderma reesei through a PCR method, cloned and inserted into a pichia pastoris integrated expression vector pPIC9K, and a pPIC9K-eg5 expression vector is obtained. The vectoris introduced into pichia pastoris GS115 through electrotransformation to obtain recombinant pichia pastoris strains, and the recombinant pichia pastoris strains comprising high-copy integrated plasmids are screened by using antibiotics G418 with different concentrations. The optimal induction condition is determined through primary optimization of shake flask fermentation, and meanwhile, the enzymatic properties of the purified recombinant protease are analyzed.

The invention discloses a method for increasing the output of alkaline pectinase produced by fermentation by adding sorbitol, relating to the technical field of application of the carbon source adding policy in the optimization of fermentation. In the method, recombinant Pichia pastoris GS115 is adopted as fermentation strains, and sorbitol is added into the fermentation liquid at a constant speed or accelerated speed during fermentation induction phase to improve the activity of cells, weaken the toxicity of methanol, and increase the output of the alkaline pectinase. The lower adding amountof sorbitol can not inhibit the enzyme production effect of methanol to the strains, on the other hand, the toxicity to the strains from the methanol can be reduced to a certain extent, and the efficiency of enzyme production is further increased. When the sorbitol is added at a varying speed (0.9g / h-7.2g / h), the activity of enzyme is increased from 930U / ml to 1541U / ml. The invention is simple and effective in process, capable of being applied in the industrial production of pectinase in the future, and instructive to the optimization of fermentation of other Pichia pastoris.