36 results about "Chorismic acid" patented technology

The invention relates to a marker-free gene deletion attenuated mutant strain of an Edwardsiella tarda wild strain. The marker-free gene deletion attenuated mutant strain is an attenuated live vaccine of an Edwardsiella tarda virulent strain, which deletes the chorismic acid synthase gene aroC of the Edwardsiella tarda virulent strain, three types of secretion system response element genes of eseB, escA, eseC and eseD and an endogenous plasmid, preferably, the Edwardsiella tarda virulent strain is an Edwardsiella tarda virulent strain EIB202 with the preservation number of CCTCC No:M208068; the endogenous plasmid is a plasmid of pEIB202; and the marker-free gene deletion attenuated mutant strain of the Edwardsiella tarda virulent strain is an attenuated strain WED with the preservation number of CCTCC No:M2010278. The invention also provides relevant preparations and application of the marker-free gene deletion attenuated mutant strain. The attenuated mutant strain or relevant preparations eliminate the potential environment and safety risk of products existing in the traditional attenuated live vaccines generally and is a safe, effective and economic vaccine aiming at Edwardsiella tarda diseases of cultured fishes.

A method for producing an L-amino acid is described, for example, L-phenylalanine and L-histidine, by fermentation using a bacterium of the Enterobacteriaceae family, wherein the bacterium has been modified by attaching a DNA fragment able to be transcribed encoding the peptide represented in SEQ ID NO: 2, or a variant thereof, particularly a portion of the ssrA gene, to the 3′-end of gene encoding for the bacterial enzyme, which influences on the L-amino acid biosynthesis, such as chorismate mutase / prephenate dehydrogenase or phosphoglucose isomerase.

The invention discloses recombinant corynebacterium glutamicum capable of being used for highly yielding L-phenylalanine and a method for producing the L-phenylalanine (Phe) by fermentation and belongs to the field of metabolic engineering. According to the invention, a corynebacterium glutamicum engineering strain C.glutamicum19AF / 99TP capable of being used for highly yielding the L-Phe is obtained in a corynebacterium glutamicum type strain ATCC13032 by carrying out induction expression on the following four genes: a 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase gene, a chorismate mutase / prephenate dehydratase gene which has resistance to feedback inhibition, a transketolase gene and a phosphoenolpyruvic acid synthetase gene. A shikimic acid metabolic pathway of an L-Phe synthetic metabolic pathway is over-expressed in the C.glutamicum ATCC13032; a key enzyme gene of a chorismic acid metabolic pathway enables the yield of the L-Phe to reach 3.47g / L; two enzyme genes which combines an expression center metabolic pathway to improve a precursor enables the highest yield of the L-Phe to reach 4.86g / L; and the original strain ATCC13032 which is used as a contrast strain cannot detect accumulation of the L-Phe in the integral fermenting process.

The invention relates to the technical scheme of organisms, in particular to gene modification application and a bacterial strain obtained through gene modification. MK3-MEP123, sourced from B.subtilis168 and established in a laboratory, is used as an original strain, at first, glycerinum-3-phosphate dehydrogenase (GlpD) is subjected to overexpression, the transformation from glycerinum-3-phosphate to dihydroxyacetone phosphate is improved, and the influence on MK-7 synthesis is inspected; then, dhbB genes are knocked out, consumption of isochorismate is reduced, and the influence on MK-7 synthesis is inspected; and finally the influence of the supply of an oxygen and carbon and nitrogen source on MK-7 synthesis is inspected through 2L shake flask fermentation. A fundamental research and theoretical basis is provided for establishing a high-yield MK-7 bacterial strain of bacillus natto by a metabolic engineering method industrially.

The invention discloses a gene engineering strain capable of safely and efficiently producing Phenazino-1-carboxylic acid. The gene engineering strain is Pseudomonas aeruginosa with a collection number of CCTCC No. M2015040. The gene engineering strain is prepared according to the following steps: performing pathogenic gene knockout on a strain PA1201; then, performing Shenqinmycin metabolizing gene knockout and metabolize metabolizing gene knockout; then, performing point mutation to reduce the bioactivity of PheA; performing isozyme substitution method to reduce the bioactivity of chorismic acid / pyruvic acid lyase; increasing a phzC gene copy number in a genome; replacing an aroG promoter with a strong promoter Ptac so as to enhance the gene expression level of DAHP synthase; and enhancing the Phenazino-1-carboxylic acid biosynthetic gene cluster expression level and Phenazino-1-carboxylic acid efflux pump MexGHI-OpmD expression level by a promoter replacement method. The obtained strain is capable of safely and economically producing Phenazino-1-carboxylic acid with superhigh yield.

Provided is a phenol-producing transformant constructed by transferring a gene which encodes an enzyme having chorismate-pyruvate lyase activity and a gene which encodes an enzyme having 4-hydroxybenzoate decarboxylase activity into a coryneform bacterium as a host. Also provided is a process for producing phenol, which comprises a step of allowing the transformant to react in a reaction mixture containing a saccharide under reducing conditions, and a step of collecting phenol from the reaction mixture.

An objective of the present invention is to provide a transformant altered so as to produce a secondary metabolite in which a benzene ring of the secondary product is modified at the para-position with a functional group containing a nitrogen atom. A transformant according to the present invention is a transformant of an organism producing a secondary metabolite having a benzene ring skeleton without substitution at the para-position with a functional group containing a nitrogen atom, said transformant being transformed by introducing genes involved in a biosynthetic pathway from chorismic acid to p-aminophenylpyruvic acid, including a gene encoding an amino acid sequence (SEQ ID NO: 2) having 4-amino-4-deoxychorismic acid synthase activity, a gene encoding an amino acid sequence (SEQ ID NO: 4) having 4-amino-4-deoxychorismic acid mutase activity and a gene encoding an amino acid sequence (SEQ ID NO: 6) having 4-amino-4-deoxyprephenic acid dehydrogenase activity, so as to produce a secondary metabolite having a benzene ring skeleton substituted at the para-position with a functional group containing a nitrogen atom. Another objective of the present invention is to provide a novel gene involved in the biosynthetic pathway from chorismic acid to p-aminophenylpyruvic acid. A novel gene according to the present invention comprises genes encoding the amino acid sequences of SEQ ID NOs: 2, 4 and 6 or modified sequences thereof.

The invention discloses the method of improving the content of the salicin in the plant and the appropriative carrier. The carrier is the plant expressing carrier containing the separating offset acid composing enzyme gene and the separating offset acid-the pyruvic splitting dividing enzyme gene. The method uses the salicin composing approach and guides the different ICS gene and the IPL gene into the plant. The invention guides the said carrier into the plant group or the cell to gain the turning gene plant of the content of the salicin improved. The transferring gene test indicates that the dissociating state SA content in the transferring gene rice is mostly nine times than the wildness type. The method of the invention and the appropriative carrier can improve the content of the salicin in the plant greatly and provides the new method of amending the plant breed and improving the crop.

Provided is a method for producing an aniline derivative by fermentation from a carbon source such as glucose. The method comprises the following steps: production of microorganisms capable of producing 1.8 g / L or more of 4-aminophenylalanine (4APhe) under prescribed culture conditions by introducing at least three exogenous genes into microorganisms having the ability to biosynthesize 4-aminophenylpyruvic acid from chorismic acid; and production of at least one aniline derivative selected from the group consisting of 4-aminophenylalanine (4APhe), 4-aminocinnamic acid (4ACA), 2-(4-aminophenyl)aldehyde, 4-aminophenylacetic acid, and 4-aminophenethylethanol (4APE) by bringing these microorganisms into contact with a carbon source under conditions suited to the growth and / or maintenance of these microorganisms.

The invention discloses rapid sputum dissolving and storing liquid. The rapid sputum dissolving and storing liquid comprises the following components in parts by weight: ethanol 10-20 parts, glycerine 10-20 parts, vegetable oil 1-2 parts, chorismic acid 0.1-1 part, disodium EDTA (ethylene diamine tetraacetic acid) 0.01-0.2 part, SDS (sodium dodecyl sulfonate) 0.001-0.1 part, thiosugar 2-10 parts, and distilled water 50-80 parts. The rapid sputum dissolving and storing liquid can rapidly dissolve sputum and prevent mycobacterium tuberculosis from cracking; and meanwhile, the storage time of the dissolving and storing liquid is long, the rapid sputum dissolving and storing liquid can still be used after being continuously stored for 3-5 years, and the activity is not weakened obviously.

The nucleotide sequence of a 992 bp region of cDNA and the nucleotide sequence of a 1973 bp (or a 1913 bp) of genomic DNA of the Gr-cm-1 gene were determined for G. rostochiensis. PCR primers and probes specific for G. rostochiensis and G. pallida were generated. PCR assays, including a real-time TaqMan PCR were used to identify G. rostochiensis and G. pallida and to differentiate G. rostochiensis from G. pallida. Transgenic hairy roots expressing Gr-cm-1 dsRNA were generated. There was a 52% reduction in the average number of females per root in the Gr-cm-1 dsRNA transgenic lines when compared with the infected control lines.

The invention discloses a chorismate mutase nucleotide sequence related to high yield of rice and application thereof. A method for screening rice with different yield traits provided by the inventioncomprises the following steps: (1) detecting the genotype based on a specific gene fragment, of the to-be-detected rice, wherein the specific gene fragment, OSCM3, is located in the rice genome, andhas two allelic forms: OSCM3_a and OSCM3_b; the OSCM3_a is shown by the sequence 1 of a sequence table; and the OSCM3_b is shown by the sequence 2 of the sequence table; and (2) judging in the following way: comparing tow genotypes in an adjoining growth condition, to determine that the average yield of the OSCM3_b homozygotic type rice population is higher than average yield of the OSCM3_a homozygotic type rice population. Experiments prove that the method provided by the invention can effectively screen the rice with different traits in yield per plant, and has the advantages of easiness inoperation and high accuracy, thereby having an important application value in rice breeding.

The invention discloses a biosynthesis method of a nicotinamide adenine dinucleotide compound. A synthesis path provided by the invention takes chorismic acid as a starting point, under the action ofPhzD protein, a transamination rearrangement reaction is carried out to generate amino deoxy isobranched acid, pyruvic acid is removed from amino deoxy isobranched acid under the catalysis of the PhzEprotein to generate 2,3-dihydro-3-hydroxyanthranilicacid, 2,3-dihydro-3- hydroxyanthranilicacid is catalyzed to generate 3-hydroxyanthranilicacid by adopting a specific enzyme, quinolinic acid is generated from 3-hydroxyanthranilicacid through an oxidation ring-opening rearrangement reaction under the action of nabC protein, and then enters an NAD remediation way to finish NAD synthesis. According to the present invention, the coupling between the NAD synthesis and the tryptophan or aspartic acid synthesis is removed, and the chorismic acid widely exists in various life forms, and has less influence on other life essential metabolic pathways.

The invention discloses engineering bacteria for producing maleic acid, and a construction method and applications thereof, and relates to the technical field of bioengineering. The provided engineering bacteria include genes that can be co-expressed and code isochorismate synthase, isochorismate pyruvate lyase, salicylic acid-5-hydroxylase, maleylpyruvate isomerase and maleylpyruvate lyase in a host. The above engineering bacteria also include the genes that are over-expressed and code shikimate kinase, DAHP synthetase, PEP synthetase and transketolase A in the host so as to realize the increased production of maleic acid. A construction method and applications of the engineering bacteria for producing the maleic acid are also provided. Through the utilization of the engineering bacteriato produce the maleic acid by taking glucose and / or glycerol as carbon source, the de novo synthesis of the maleic acid can be realized.

The present invention relates to a polypeptide which belongs to the R3R3-type MYB family and which regulates the shikimate pathway towards the production of benzenoids. The shikimate pathway is a biosynthesis pathway through which the three essential aromatic amino acids tyrosine, phenylalanine and tryptophan are synthesized in plants, bacteria and fungi. The present invention provides for the first time a regulatory protein in the shikimate pathway and a means to regulate the biosynthesis of these three essential amino acids which cannot be produced by mammals. At the same time, it opens up the way for the regulation of the biosynthesis of aromatic and non-aromatic compounds which are derived from these essential amino acids. A polypeptide or polynucleotide of the invention may be used in a method for manipulating the transcript levels of the genes of the shikimate pathway towards benzenoids for instance the genes of 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase (DAHPS), 5-enol-pyruvylshikimate-3-phosphate synthase (EPSPS), L-phenylalanine ammonia-lyase (PAL) and chorismate mutase (CM).

Provided is a microorganism that is able to efficiently produce protocatechuic acid or a salt thereof by using a saccharide as a raw material, and a method of efficiently producing protocatechuic acid or a salt thereof by using the microorganism. Provided is a transformant having protocatechuic acid producing ability, subjected to modifications of enhancement of 3-dehydroshikimate dehydratase activity; enhancement of chorismate pyruvate lyase activity; and enhancement of 4-hydroxybenzoate hydroxylase activity. Also provided is a method of producing protocatechuic acid or a salt thereof, including the step of culturing the transformant in a reaction solution containing a saccharide so as to cause the transformant to produce protocatechuic acid or a salt thereof.

The invention discloses an enzymatic activity-changed amino deoxychorismate synthetase mutant T426I and application thereof, and belongs to the field of genetic engineering and fermentation engineering. According to the enzymatic activity-changed amino deoxychorismate synthetase mutant T426I, a T426 locus of amino deoxychorismate synthetase in parent Corynebacterium glutamicum delta SSAAI (the preservation name of a strain is delta SSAAI, and the preservation number is CGMCC No 15170) is mutated into isoleucine, and the enzymatic activity of the obtained mutant is reduced by 15.2%. A genetically engineered bacterium constructed by the mutant is fermented, and the yield of L-serine is increased by 18.75%. The enzymatic activity-changed amino deoxychorismate synthetase mutant T426I effectively changes the activity of the amino deoxychorismate synthetase through point mutation, creates conditions for constructing the genetically engineered bacterium for efficiently producing the serine, and has good industrial application value and prospects.

The mutant chorismate-pyruvate lyase (A) or (B) as described below is capable of producing 4-hydroxybenzoic acid or a salt thereof with sufficient practical efficiency.(A) A mutant chorismate-pyruvate lyase obtained by replacing the valine at position 80 in a chorismate-pyruvate lyase (ubiC) from Pantoea ananatis consisting of the amino acid sequence of SEQ ID NO: 1 with one or more other amino acids.(B) A mutant chorismate-pyruvate lyase obtained by replacing an amino acid in another chorismate-pyruvate lyase, the amino acid being at a position enzymologically homologous with that of the above valine, with one or more other amino acids.

The invention provides an engineering bacterium suitable for salicin, which comprises a host and a recombinant plasmid vector, and the recombinant plasmid vector is connected with a gene for expressing and coding glucosyltransferase. The recombinant plasmid vector is also connected with a gene for coding carboxylic acid reductase and phosphoryl transferase, which is co-expressed with the gene for coding glucosyltransferase. The recombinant plasmid vector is also connected with genes for over-expressing and coding shikimic acid kinase, pyruvate kinase, transketolase, 3-deoxy-7-phosphoenanthate synthase, iso-chorismate pyruvate lyase and isochorismate synthase; and the host is also knocked out of genes for coding pykA and pykF, so that the yield of salicin is increased. The invention also provides a construction method and an application of the engineering bacterium for synthesizing salicin. By utilizing the engineering bacteria for synthesizing the salicin, the salicin is produced by using glucose and / or glycerol and other carbon sources, so that efficient biosynthesis of the salicin is realized.

A transformant constructed by introducing a gene which encodes an enzyme having chorismate-pyruvate lyase activity into a coryneform bacterium as a host is capable of efficiently producing 4-hydroxybenzoic acid or a salt thereof from a sugar. When the transformant is cultured under aerobic conditions where the transformant does not grow, 4-hydroxybenzoic acid or a salt thereof can be produced in a particularly efficient manner.

The invention discloses rapid sputum dissolving and storing liquid. The rapid sputum dissolving and storing liquid comprises the following components in parts by weight: ethanol 10-20 parts, glycerine 10-20 parts, vegetable oil 1-2 parts, chorismic acid 0.1-1 part, disodium EDTA (ethylene diamine tetraacetic acid) 0.01-0.2 part, SDS (sodium dodecyl sulfonate) 0.001-0.1 part, thiosugar 2-10 parts, and distilled water 50-80 parts. The rapid sputum dissolving and storing liquid can rapidly dissolve sputum and prevent mycobacterium tuberculosis from cracking; and meanwhile, the storage time of the dissolving and storing liquid is long, the rapid sputum dissolving and storing liquid can still be used after being continuously stored for 3-5 years, and the activity is not weakened obviously.

The invention discloses a method for increasing the yield of vitamin K2 by utilizing recombinant bacillus natto. The method comprises the following steps: overexpressing a flutalosin synthetase gene mqnA on a bacillus natto chromosome through a constitutive promoter P43; on the basis of the obtained strain NA1, the strain NA1 is obtained; the method comprises the following steps: overexpressing agene of deoxyxanthine flutalosin synthetase on a bacillus natto chromosome by using a P43 promoter; on the basis of the obtained bacterial strain NA2; a Phbs promoter is used for replacing a natural promoter of an O-succinylbenzoic acid-CoA ligase gene in bacillus natto; on the basis of the obtained strain NA3, isobranching acid genes on bacillus natto chromosomes are replaced with 1, 4-dihydroxy-6-naphthoic acid synthase genes which contain Phbs promoters and are derived from streptomyces coelicolor, and a seed solution of the recombinant bacteria is inoculated into a fermentation culture medium according to the inoculation amount of 3%-6%. According to the invention, four recombinant bacteria NA1-NA4 are constructed, wherein the NA3-NA4 significantly improves the yield of vitamin K2.

The present invention relates to a polypeptide which belongs to the R2R3-type MYB family and which regulates the shikimate pathway towards the production of benzenoids. The shikimate pathway is a biosynthesis pathway through which the three essential aromatic amino acids tyrosine, phenylalanine and tryptophane are synthesized in plants, bacteria and fungi. The present inventiont provides for the first time a regulatory protein in the shikimate pathway and a means to regulate the biosynthesis of these three essential amino acids which cannot be produced by mammals. At the same time, it opens up the way for the regulation of the biosynthesis of aromatic and-non- aromatic compounds which are derived from these essential amino acids. A polypeptide or polynucleotide of the invention may be used in a method for manipulating the transcript levels of the genes of the shikimate pathway towards benzenoids for instance the genes of 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase (DAHPS), 5-enol-pyruvylshikimate-3-phosphate synthase (EPSPS), L- phenylalanine ammonia-lyase (PAL) and chorismate mutase (CM). Through these enzymes, biosynthetic processes at lower levels may be influenced. For instance, compounds of the invention may be used in a method to regulate scent in flowers or to regulate resistance to pest insects or pathogenic organisms.

The invention discloses an engineering bacterium for microbial synthesis of piceatannol by taking glucose as a substrate as well as construction and application of the engineering bacterium. According to the invention, L-tyrosine ammonolysis enzyme, p-coumaroyl-coenzyme A ligase, resveratrol synthetase, DAHP synthetase, chorismate mutase and hydroxylase complex genes are over-expressed in the engineering strain, so that the de novo synthesis strategy of piceatannol is realized, and the content of the product piceatannol is remarkably increased; and a simple and effective implementation method is provided for safe biotransformation. The yield of the engineering bacterium for synthesizing piceatannol by using the glucose as the substrate microorganism disclosed by the invention can reach 7-8g / L.

The invention discloses recombinant corynebacterium glutamicum capable of being used for highly yielding L-phenylalanine and a method for producing the L-phenylalanine (Phe) by fermentation and belongs to the field of metabolic engineering. According to the invention, a corynebacterium glutamicum engineering strain C.glutamicum19AF / 99TP capable of being used for highly yielding the L-Phe is obtained in a corynebacterium glutamicum type strain ATCC13032 by carrying out induction expression on the following four genes: a 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase gene, a chorismate mutase / prephenate dehydratase gene which has resistance to feedback inhibition, a transketolase gene and a phosphoenolpyruvic acid synthetase gene. A shikimic acid metabolic pathway of an L-Phe synthetic metabolic pathway is over-expressed in the C.glutamicum ATCC13032; a key enzyme gene of a chorismic acid metabolic pathway enables the yield of the L-Phe to reach 3.47g / L; two enzyme genes which combines an expression center metabolic pathway to improve a precursor enables the highest yield of the L-Phe to reach 4.86g / L; and the original strain ATCC13032 which is used as a contrast strain cannot detect accumulation of the L-Phe in the integral fermenting process.

The invention relates to a marker-free gene deletion attenuated mutant strain of a Edwardsiella tarda wild strain. The marker-free gene deletion attenuated mutant strain is an attenuated live vaccine of an Edwardsiella tarda virulent strain, which deletes the chorismic acid synthase gene aroC of the Edwardsiella tarda virulent strain, three types of secretion system response element genes of eseB, escA, eseC and eseD and an endogenous plasmid, preferably, the Edwardsiella tarda virulent strain is a Edwardsiella tarda virulent strain EIB202 with the preservation number of CCTCC No:M208068; the endogenous plasmid is a plasmid of pEIB202; and the marker-free gene deletion attenuated mutant strain of the Edwardsiella tarda virulent strain is an attenuated strain WED with the preservation number of CCTCC No:M2010278. The invention also provides relevant preparations and application of the marker-free gene deletion attenuated mutant strain. The attenuated mutant strain or relevant preparations eliminate the potential environment and safety risk of products existing in the traditional attenuated live vaccines generally and is a safe, effective and economic vaccine aiming at Edwardsiella tarda diseases of cultured fishes.

The invention provides a preparation method of a formamide phenazine bio-fungicide. The preparation method comprises the following steps: firstly, implementing shake-flask culture on activated pseudomonas chlororaphis and pseudomonas aeruginosa, so as to obtain a seed solution A and a seed solution B; inoculating a fermentation medium with the seed solution A, conducting fermentation culture for 20h, inoculating the fermentation medium with the seed solution B, simultaneously refilling a nutrient solution, and continuing to implement fermentation culture for 40-50h, so that fermentation brothis obtained; centrifuging the fermentation broth at high speed, regulating pH value, and conducting extracting and concentrating, so as to a crude product; and implementing column chromatography and separating, so that the formamide phenazine bio-fungicide is obtained. The culture medium and culture conditions provided by the invention are quite suitable for growth of strains; the refilled nutrient solution contains chorismic acid; two strains undergo mixed fermentation; the content of formamide phenazine in the fermentation broth is obviously improved; based upon a series of post-treatment, afinished product that purity is higher than 99% is separated and obtained; and the finished product takes a strong inhibitory effect on pathogenic fungi of plants.

The invention provides an engineering bacterium for producing alpha-arbutin. The engineering bacterium comprises genes for co-expressing and encoding alpha-arbutin synthetase, PchB, SalABCD, and GD in a host, wherein the alpha-arbutin synthetase is one of or any combination of HGE, XgtA, and DGAS. The engineering bacterium further comprises genes for co-expressing AroL, ppsA, tktA, aroG<fbr>, and EntC from escherichia coli, so that the yield of the alpha-arbutin is increased. The invention also provides a construction method and application of the engineering bacterium for producing the alpha-arbutin. By utilizing the engineering bacterium for producing the alpha-arbutin, the alpha-arbutin is produced by using carbon sources such as maltose and / or glycerol, so that the denovo synthesis of the alpha-arbutin is realized.

The invention discloses a chorismate mutase nucleotide sequence related to high yield of rice and application thereof. A method for screening rice with different yield traits provided by the inventioncomprises the following steps: (1) detecting the genotype based on a specific gene fragment, of the to-be-detected rice, wherein the specific gene fragment, OSCM3, is located in the rice genome, andhas two allelic forms: OSCM3_a and OSCM3_b; the OSCM3_a is shown by the sequence 1 of a sequence table; and the OSCM3_b is shown by the sequence 2 of the sequence table; and (2) judging in the following way: comparing tow genotypes in an adjoining growth condition, to determine that the average yield of the OSCM3_b homozygotic type rice population is higher than average yield of the OSCM3_a homozygotic type rice population. Experiments prove that the method provided by the invention can effectively screen the rice with different traits in yield per plant, and has the advantages of easiness inoperation and high accuracy, thereby having an important application value in rice breeding.