18 results about "Chitin deacetylase" patented technology

The invention relates to the technical field of food, in particular to geobacillus stearothermophilus cKM-Geo, application of the geobacillus stearothermophilus cKM-Geo, a shrimp shell oligosaccharide peptide compound and a preparation method of the shrimp shell oligosaccharide peptide compound. The invention provides geobacillus stearothermophilus cKM-Geo, the strain can grow in an oxygen-free environment, ventilation is not needed in the process, and proteolytic enzyme, cellulase, lipase and chitin deacetylase can be highly secreted and expressed; the invention further provides a preparation method for preparing the shrimp shell oligosaccharide peptide by utilizing geobacillus stearothermophilus cKM-Geo fermentation coupling steam explosion treatment, chitin and protein in the shrimp shell are converted into water-soluble oligopeptide and oligosaccharides, the water-soluble oligopeptide and oligosaccharides are separated from calcium carbonate in the shrimp shell, acid and alkali do not need to be used, in the process, waste water is little, pollution is little, and the method is suitable for industrial production. The resource utilization rate is high.

The present invention provides a recombinant chitin deacetylase, its preparation method, and application. The inventors have isolated and obtained a novel chitin deacetylase that not only catalyzes the deacetylation of chitin but also effectively inhibits pathogenic bacteria. The chitin deacetylase of the present invention has good adaptability to low temperatures, can be expressed under prokaryotic expression conditions, and exhibits relatively high low-temperature activity. Furthermore, the present invention also optimizes the recombinant expression method of the low-temperature chitin deacetylase, thereby enabling its efficient expression in host cells.

The invention discloses a high-deacetylation-degree chitosan preparation method, which comprises: suspending chitin in a solvent to prepare a chitin suspension; adding a compound enzyme into the obtained suspension liquid to carry out enzymatic reaction, so as to prepare the chitosan with high deacetylation degree; wherein the compound enzyme is prepared from chitin deacetylase, chitinase, beta-N-acetylglucosaminidase and protease. According to the method, four enzymes with complementary functions are creatively combined to form a unique technical path of four-stage synergistic interaction of'removal-deconstruction-assistant-deacetylation ', so that a natural chitin crystal barrier can be efficiently deconstructed, the protein shielding effect is overcome, and the effect of directly preparing high-deacetylation-degree chitosan from chitin is achieved.

The invention relates to the field of genetic engineering, in particular to a non-methanol-induced chitin deacetylase pichia pastoris expression vector and engineering bacteria as well as a construction method and application of the non-methanol-induced chitin deacetylase pichia pastoris expression vector and engineering bacteria. According to the invention, a carrier containing a constitutive strong promoter is adopted as a skeleton, a chitin deacetylase gene coding sequence is inserted into the carrier, methanol is not required to be used as an inducer when the obtained recombinant plasmid is applied, and the produced chitin deacetylase is efficient, green and environment-friendly, and can be applied to the fields of beauty industry, agriculture, biomedicine and the like. The invention establishes a non-methanol-induced chitin deacetylase pichia pastoris expression vector and engineering bacteria, which are used for high-efficiency expression of chitin deacetylase, and solves the problem of low enzyme yield of chitin deacetylase from natural sources.

The invention relates to the technical field of microbial fermentation and enzyme engineering, and in particular relates to a strain Precottula equi GXAS 17-2 for high yield of chitin deacetylase as well as a fermentation culture medium and application thereof, the preservation number of the strain Precottula equi GXAS 17-2 is GDMCC NO: 67488, and the strain is separated from mangrove forest soil and can efficiently produce enzyme in the culture medium taking N-acetylglucosamine (GlcNAc) as an inducer. The enzyme activity is remarkably improved by optimizing culture medium components (3% of colloidal chitin, 10% of yeast powder and 0.3% of GlcNAc) and culture conditions (30 DEG C, 180 rpm, pH 7 and 48 h). The method has the advantages of high induction efficiency, green process, stable enzyme activity and the like, and is suitable for chitin degradation and chitosan green preparation industry.

The invention discloses a method for synchronously preparing chitosan and citric acid through a kitchen waste lobster shell fermentation method, belongs to the field of solid waste resource utilization, and aims to solve the problems of difficult treatment, low utilization rate and pollution caused by impurity wrapping in shellfish kitchen waste treatment. According to the method, kitchen waste containing grease, chitin, calcium carbonate and protein is used as a raw material, and two-stage treatment mainly including fermentation is carried out. In the first stage, citric acid is produced through fermentation, and chitin is exposed through in-situ deoiling, decalcification and deproteinization; in the second stage, enzymolysis is mainly carried out, and after the pH is adjusted, chitinase and chitin deacetylase are exogenously added to convert chitin into chitosan; and finally, separating the feed liquid to obtain citric acid and chitosan. According to the method, fermentation is combined with enzymolysis segmented treatment, so that the problem of substrate wrapping is solved, the conversion rate and purity of chitosan are remarkably improved, high-valued co-production of waste is realized, and the process is mild and environment-friendly.

The invention belongs to the technical field of marine bioengineering, and discloses N-acetylated chitosan oligosaccharide for inducing plant salt-resistant activity, and a preparation method of the N-acetylated chitosan oligosaccharide comprises the following steps: reacting all N-acetylated chitosan oligosaccharide with chitin deacetylase ArCE4, desalting after the reaction, and freeze-drying to obtain the N-acetylated chitosan oligosaccharide with the plant salt-resistant activity. According to the method disclosed by the invention, N-acetyl groups in the chitosan oligosaccharide and at a non-reducing end are removed at fixed points by adopting a chitin deacetylase method, so that the complexity and uncertainty of directly separating chitosan oligosaccharides with different polymerization degrees and a mixture of chitosan oligosaccharide isomers with different sequence arrangements are avoided, and a chitosan oligosaccharide target product with a specific sequence structure can be directly prepared. The enzyme reaction condition is mild, generation of by-products and environmental pollution are avoided, and the obtained chitosan oligosaccharide with the specific sequence does not contain any nucleic acid and protein, is high in purity, good in water solubility and high in plant induced resistance activity and can be used for development of agricultural biological products.

The invention belongs to the technical field of green biosynthesis of multi-enzyme catalysis, and relates to a method for preparing chitobiose by a double-enzyme catalysis method, which comprises the following steps: 1) obtaining a substrate containing chitin; 2) preparing chitin hydrolase and chitin deacetylase in a recombinant expression manner; and (3) carrying out enzyme reaction on the chitin-containing substrate obtained in the step (1), the chitin hydrolase obtained in the step (2) and chitin deacetylase obtained in the step (2) in a buffer system to obtain chitobiose. The invention provides the method for preparing the chitobiose by the double-enzyme catalysis method, which can be used for remarkably improving the yield and the purity of the chitobiose and effectively reducing the environmental pollution.

The application belongs to the technical field of marine biological engineering, and particularly relates to a preparation method of specific N-acetylated chitosan (N-acetylated chitosan with different acetylation degrees). Specifically, the method comprises the following steps: taking fully deacetylated chitosan as a substrate, and performing reaction on the substrate in sodium acetate buffer solution through chitin deacetylase (PesCDA) with a specific deacetylation mode; and after the reaction, salt is removed, concentrated and freeze-dried, thereby obtaining chitosan with a specific N-acetylation degree. Compared with a traditional method for preparing chitosan with a specific N-acetylation degree, the method is green, environmentally friendly and pollution-free.

The application discloses a chitin deacetylase mutant with improved acid resistance, a coding gene, a recombinant expression plasmid, a genetically engineered bacterium and a preparation method. The amino acid sequence of the chitin deacetylase AnCDA is subjected to the following mutations: the 119th leucine (Leu) is mutated into threonine (Thr); and / or, the 139th phenylalanine (Phe) is mutated into glycine (Gly); and / or, the 166th tyrosine (Tyr) is mutated into tryptophan (Trp). The chitin deacetylase mutant obtained in the application has significantly improved stability under acidic conditions, the half-life t1 / 2 is prolonged by nearly 3 times at pH 5.0, the optimal catalytic pH is reduced from 6.0 to 5.0, and the chitin deacetylase mutant can be applied to catalyzing chitin deacetylation under acidic conditions to prepare chitosan and other chitin products, and has a good industrial application prospect.

The invention belongs to the technical field of microorganisms, and discloses a method for improving the activity of chitin deacetylase through mixed fermentation, which is characterized in that rhodococcus qingshengii with the preservation number of CGMCC (China General Microbiological Culture Collection Center) No.34439 is used as an enzyme producing bacterium, bacillus zhangzhouensis with the preservation number of CGMCC No.34440 is used as a companion bacterium, and mixed fermentation is carried out to produce the chitin deacetylase. The chitin deacetylase obtained by adopting the method disclosed by the invention is high in activity and high in deacetylation efficiency, a potential production strain is provided for chitin biodegradation, and a theoretical basis is provided for application of the CDA enzyme generated by the strain.

The invention provides application of brown planthopper chitin deacetylase CDA, and belongs to the field of gene engineering. According to the dsRNA preparation of the NlCDA gene provided by the invention, the dsRNA preparation is introduced into a brown planthopper body, so that the brown planthopper can be effectively killed and prevented from damaging rice. The nilaparvata lugens insecticide is specifically used for nilaparvata lugens, is harmless to mammals, fishes and shrimps, natural enemy insects and pollination insects, and has the advantages of quick effect, high fatality rate, environment friendliness and the like.

The invention relates to the technical field of agricultural biology, in particular to a tomato leaf miner chitin deacetylase CDA8 gene and application of the tomato leaf miner chitin deacetylase CDA8 gene in prevention and control of tomato leaf miner. According to the invention, the CDA8 gene is cloned from tomato leaf miner, and the expression quantity of the CDA8 gene is obviously reduced after target gene double-stranded RNA (Ribonucleic Acid) is fed; after the dsCDA8 is fed, both larvae and pupae of the tomato leaf miner have phenotypic deformity, ecdysis failure, epidermis deformity and abdomen darkening of the larvae, pupae ecdysis failure and partial tissue deletion of the pupae, peritrophic membranes are obviously damaged, structural integrity is lost, and breakage is obvious. The result shows that the CDA8 gene plays a key role in the growth and development process of the tomato leaf miner larva. RNAi-mediated CDA8 silencing enhances the sensitivity of the tomato leaf miner to the insecticide Bt, and a method basis is provided for reducing the harm of the tomato leaf miner through related researches of chitin deacetylase in the future.

The invention discloses an application of a chitin deacetylase gene PbCDA in prevention and control of rape clubroot. According to the invention, a chitin deacetylase gene CDA (Chitin Deacetylase, CDA) which possibly has important influence on the pathogenic process of plasmodiophora brassicae is screened out from a gene bank of plasmodiophora brassicae ZJ-1, and an RNAi carrier of the chitin deacetylase gene CDA is constructed by using an HIGS technology and is transformed into double 11 in rape. The PbCDA-RNAi transgenic rape is subjected to a plasmodiophora brassicae disease resistance experiment, and the result shows that the PbCDA-RNAi transgenic rape has remarkable resistance to plasmodiophora brassicae, so that the gene can provide a new gene resource for green prevention and resistance of rape clubroot.

This invention discloses a chitin deacetylase product and its application in agricultural disease prevention. Through codon optimization, the *Rhodococcus* chitin deacetylase gene (SEQ ID NO:1) was cloned into the *Pichia pastoris* expression vector pGAPZa A, constructing the recombinant vector pGAPZaA-CDA-His, which was then transformed into the *Pichia pastoris* SMD1168 host, achieving efficient secretory expression of the enzyme. The resulting recombinant enzyme exhibits high activity and good thermostability. This invention includes a universal enzymatic deacetylation process characterized by deacetylation efficiency, applicable to chitin deacetylases from various sources. This invention also provides a method for further enzymatic hydrolysis to produce N-acetylglucosamine monomers or their oligosaccharides, and the application of the recombinant enzyme and its product in the preparation of biopesticides and the treatment of chitin-containing biomass materials.

Insect control nanobodies are provided. Accordingly there is provided a nanobody which specifically binds to an insect polypeptide selected from the group consisting of: a polypeptide comprising a chitin binding domain (CBD), V-ATPase subunit c, trehalase, cytochrome p450 monooxygenase, chitin deacetylase, chitin synthase and NPC1 sterol transporter, wherein binding of the nanobody to the insect polypeptide confers an insect control activity to the nanobody. Also provided are polynucleotides encoding the nanobody, host cells expressing the nanobody and methods of using it.

We provide insect control nanobodies. [Solution] A nanobody is provided that specifically binds to an insect polypeptide selected from the group consisting of a chitin-binding domain (CBD), V-ATPase subunit c, trehalase, cytochrome p450 monooxygenase, chitin deacetylase, chitin synthase, and NPC1 sterol transporter, wherein the binding of the nanobody to the insect polypeptide confers insect control activity to the nanobody. A polynucleotide encoding the nanobody, a host cell expressing the nanobody, and its applications are also provided.

The invention discloses a primer group for judging the cocooning time of bombyx mori after feeding pyriproxyfen-polluted mulberry leaves and application of the primer group, and a method for judging the cocooning time of bombyx mori by utilizing the primer group. The method comprises the following specific steps: (1) establishing a correlation between the relative transcription expression quantity of a normal silkworm chitin deacetylase 1 gene and the development time of five-instar silkworm larvae; (2) determining the CDA1 gene relative transcription expression quantity (T to be detected) of to-be-detected bombyx mori, calculating the relative transcription expression quantity (T normal) of the normal bombyx mori according to the development period of the to-be-detected bombyx mori by comparing the correlation obtained in the step (1), comparing the CDA1 gene transcription expression quantity of the to-be-detected bombyx mori with the CDA1 gene transcription expression quantity of the normal bombyx mori in the same period, and calculating the ratio R = T to be detected / T normal, and determining the cocooning time of the silkworms according to whether the ratio is greater than 1. According to the method, the accuracy, the efficiency and the operability of judging the cocooning time after the silkworm eats the mulberry leaves polluted by pyriproxyfen are improved.