31 results about "Pseudomonas denitrificans" patented technology

Recombinant organisms are provided comprising genes encoding aquacobalamin reductase, cob(II)alamin reductase, cob(I)alamin adenosyltransferase, glycerol dehydratase and 1,3-propanediol oxidoreductase activities useful for the production of 1,3-propanediol from a variety of carbon substrates. More specifically the following nucleotide sequences are provided: btuR, encoding the E. coli cob(I)alamin adenosyltransferase enzyme; cobA, encoding the Salmonella typhimurium cob(I)alamin adenosyltransferase enzyme; cobO, encoding the Pseudomonas denitrificans cob(I)alamin adenosyltransferase enzyme; dhaB1, encoding the α subunit of the Klebsiella glycerol dehydratase enzyme; dhaB2, encoding the β subunit of the Klebsiella glycerol dehydratase enzyme; dhaB3, encoding the γ subunit of the Klebsiella glycerol dehydratase enzyme; dhaT, encoding Klebsiella oxidoreductase enzyme; the yciK gene isolated from E. coli; the glucose isomerase promoter sequence from Streptomyces; and the N-terminal amino acid sequence for cob(II)alamin reductase from Pseudomonas denitrificans.

According to the invention, through amplifying a strong promoter in Ensifer adhaerens and carrying out bioinformatics analysis and functional verification, a strong promoter is obtained, and the strong promoter can be widely used for gene expression, genetic manipulation and strain improvement in Sinorhizobium meliloti, Zymomonas mobilis, Caulobacter crescentus, Pseudomonas denitrificans, Agrobacterium tumefaciens, Brucella abortus, Pseudomonas fluorescens, Rhizobium leguminosarum, Ensifer adhaerens and other alpha-proteobacteria and has a nucleotide sequence of SEQ ID NO: 1. The invention also relates to a plasmid vector containing the strong promoter, a method for constructing a genetic engineering strain by using the strong promoter, and an application of the corresponding strain and ahost cell in starting expression of a target gene.

The invention relates to a compound microorganism bacterium agent used for sludge reduction, a preparation method and application of the compound microorganism bacterium agent. The compound microorganism bacterium agent is a liquid bacterium agent prepared by fermenting denitrified pseudomonas, nocardia coralline, candida utilis, rhodobacter sphaeroides and aspergillus sojae. According to the invention, the metabiosis action between microbe consortia is utilized to form a biodegradation system in which the activities of various required enzymes are all high, so dead or decrepit thalli in activated sludge can be effectively damaged and decomposed, and meanwhile, organic matters can be decomposed; and the generation quantity of excess sludge is reduced through reducing the quantity of the sludge in the process without influencing the quality of yielding water. On the premise of not greatly changing the traditional sewage treatment process, the sludge reduction ratio can reach 30%-70% in the sewage treatment process by adopting the compound microorganism bacterium agent, and the comprehensive running cost of the sludge treatment can be reduced, so that the compound microorganism bacterium agent is of great significance on the aspects of economic, environmental and social benefits.

The invention relates to a culture medium for producing vitamin B12 by fermenting pseudomonas denitrificans and a fermentation method thereof, and a method for producing vitamin B12 by the same. The culture medium comprises a primary seed culture medium, a secondary seed culture medium and a fermentation culture medium all of which contain artificial molasses composed of maltose, thiamine, riboflavin and biotin. By substituting the artificial molasses composed of maltose, thiamine, riboflavin and biotin for beet molasses and optimizing the formula of the culture medium, the invention solves the problem that the fluctuation range of fermentation titer is large because the quality of beet molasses is unstable, and acquires a method for stably and effectively producing vitamin B12. The invention saves the doses of raw and auxiliary materials, lowers the comprehensive cost and reduces the waste water discharge.

This invention consists of a consortium of plant-growth promoting microorganisms including Pseudomonas stutzeri, Pseudomonas denitrificans, Pseudomonas resinovorans, Pseudomonas brassicacearum, Pseudomonas fluorescens, Shimwellia blattae and Klebsiella oxytoca. The bacterial consortium along with a vehicle suitable for agricultural application, form a biofertilizer useful to enhance agricultural yield when applied to cultivation plants. The biofertilizer described present a long shelf life and preserve a high cell density along the time.

The invention belongs to the technical field of biological fermentation, and discloses a method for producing vitamin B12 by using Pseudomonas aeruginosa based on pH value control. The method for producing the vitamin B12 by using the Pseudomonas aeruginosa based on the pH value control comprises the steps that a production strain of vitamin B12, namely the Pseudomonas aeruginosa is inoculated into a fermentation medium, and fermentation is carried out at a temperature of 25-30 DEG C; the composition of the fermentation medium comprises 70-85g / L of cane molasses, 90-110g / L of yeast extract, 0.05-0.2 g / L of 5,6-dimethylbenzimidazole, 0.1-1g / L of glycerophosphoric acid, 0.05-2g / L of cobalt chloride, 0.05-0.2g / L of urea, 10-30g / L of betaine, 0.05-0.2 g / L of arginine, 0.01-0.5g / L of thiamine and the like; the pH value of fermentation broth is controlled at 7 by the cane molasses and the urea; and the purified vitamin B12 is separated from the fermentation broth. The yield of the vitamin B12 is improved by controlling the pH in the fermentation process.

The present invention relates to a promoter system inducing expression of 3-hydroxypropionic acid (3-HP) and a method of biologically producing 3-HP using the same. To improve production of 3-HP in a biological manner, continuous synthesis of new enzymes having enzyme activity is necessary. As a result of screening 3-HP reactive transcription regulators and 3-HP reactive promoters from several microorganisms including Pseudomonas denitrificans, it was confirmed that the transcriptions regulations and promoters are composed of LysR proteins and particular gene nucleotide sequences binding to the LysR proteins. Therefore, the 3-HP inducible system is expected to be effectively used to regulate 3-HP metabolic pathways.

The invention belongs to the technical field of biological fermentation and discloses a multi-stage rotating speed regulating policy for improving pseudomonas denitrificans fermentation for productionof vitamin B12. A pseudomonas denitrificans strain is fermented and cultured in a fermenting culture medium. One litter of the culture medium contains 70-85g of saccharose, 40-55g of corn steep liquor, 10-20g of glycine betaine, 0.02-0.05g of auxin, 0.01-0.05g of tryptophan and 0.05-0.1g of glutamine and the like; if the fermentation time is 0-25h, the stirring rotating speed is 150-280 rpm; if the fermentation time is 26-90 h, the stirring rotating speed is 300-400 rpm; if the fermentation time is 91-185h, the stirring rotating speed is 100-200 rpm; vitamin B12 is separated and purified fromfermentation liquor. By controlling the stirring rotating speed and the components of the fermenting culture media, the output of vitamin B12 is improved.

The invention relates to a microbial agent applied to wax and corrosion prevention of an oil well. The microbial agent comprises 10.0 to 18.0 percent of industrial salt, 2.0 to 4.0 percent of inorganic ammonium salt, 4.0 to 6.0 percent of nitrate, 1.0 to 2.0 percent of inorganic phosphorus salt, 0.5 to 1.0 percent of inorganic sulfate, 8.0 to 12.0 percent of white oil, 5.0 to 10.0 percent of microbial wax-preventing culture, 5.0 to 10.0 percent of denitrifying culture and 37.0 to 64.5 percent of water, wherein the microbial wax-preventing culture comprises pseudomonas, corynebacterium and micrococcus; and the denitrifying culture comprises pseuomonas denitrifican and paracoccus denitrificans. The invention further provides a preparation method of the microbial agent. The microbial agent can control corrosion of the oil well (hydrogen sulfide corrosion generated by sulfate reducing bacteria) while controlling wax precipitation of the oil well, and can avoid various problems caused by the use of the traditional chemical wax cleaning and preventing agent and chemical sterilizer; and the microbial agent is multi-purpose, high in reliability, high in economic benefit, convenient to useand good in effect.

The invention provides a strong promoter and application thereof in vitamin B<12>-producing strains. According to the invention, a strong promoter in Ensifer adhaerens is amplified; bioinformatics analysis and functional verification are carried out to obtain the strong promoter which can be widely applied to the vitamin B<12>-producing strains such as Sinorhizobium meliloti, Pseudomonas denitrificans and Ensifer adhaerens for gene expression, genetic gene operation and strain improvement, and the nucleotide sequence of the obtained strong promoter is SEQ ID NO: 1. The invention also relates to a plasmid vector containing the strong promoter, a method for constructing a gene engineering strain by using the promoter and a corresponding strain thereof, and application of a host cell in promoting expression of a target gene.

The invention discloses a salt-tolerant efficient denitrification-type microbial preparation for water treatment. The microbial preparation consists of the following components in percentage: 2.5-5% of halophilic archaea, 5-10% of bacillus subtilis, 5-10% of pseudomonas denitrificans, 5-10% of halomonas and 5-10% of halophilic bacteria, and 5-10% (w / t) of bamboo charcoal particles are added. The microbial preparation for water treatment consisting of halophilic archaea, pseudomonas denitrificans, bacillus subtilis, nitrifying bacteria, denitrifying bacteria and coryneform bacteria still has good denitrification capacity under the condition of high salt, and solves the limitation of a high salt environment on traditional biological denitrification methods. The bamboo charcoal particles have good adsorption performance because of large specific surface area, can be in direct contact with microorganisms better, and can adsorb organic matters on the surface, so that the formation of a surface biological membrane is promoted; and on the other hand, a large amount of oxygen can be stored in the bamboo charcoal particles, so that oxygen-containing components in a water area are improved, and a water area recovery effect is improved.

The invention relates to the technical field of biology fermentation, in particular to a method for promoting pseudomonas denitrificans to generate vitamin B12. The method for promoting pseudomonas denitrificans to generate vitamin B12 comprises the following steps of through a mother bottle culture medium, a shake flask culture medium and a supplemented medium optimized through massive experiments, under the optimized fermentation condition, production bacterial strains of vitamin B12 are cultured. In 0-39h of the front period of the fermentation process, the temperature is controlled to be at a relatively-low level, namely 26-30 DEG C, the strains are controlled to be at appropriate growth rate and favorable mushroom shapes are maintained. In the middle and later periods of the fermentation process, the temperature of the fermentation system is increased and maintained to be at a relatively-low level, namely 30-34 DEG C, the growth of thalli is promoted, starting and accumulation ofsynthesis of the vitamin B12 are promoted, and further, after the fermentation is finished, the fermentation unit of the vitamin B12 is as high as 198mg / L, so that the yield and the production efficiency of the vitamin B12 are increased, the energy consumption is reduced, and the cost is reduced.

The invention discloses pseudomonas denitrificans having the preservation number of CGMCC No.10845. The invention also discloses a microbial inoculum comprising the pseudomonas denitrificans. The invention further discloses an application of the pseudomonas denitrificans and the microbial inoculum in degradation of quinclorac. The invention also discloses a method for degradation of quinclorac. The pseudomonas denitrificans is characterized in that: (1) only an inorganic salt culture medium is provided, and the pseudomonas denitrificans can survive and grow in a culture environment containing quinclorac as a sole carbon source, and has the advantages of strong vital force, fast growth speed, and large living weight; (2) during preparation, only a preservation strain needs to be activated, the operation is simple and the cost is low; and (3) the pseudomonas denitrificans has high degradation effect in water bodies and soil, in the water bodies, under a condition that the content of quinclorac is 200 mg / L, the bacteria are cultured for 24 h, the degradation rate is as high as 93.4%, and in the soil, under a condition that the content of quinclorac is 20 mg / kg, the degradation rate is as high as 88.0%.

The invention discloses a strong promoter from ensifer adhaerens as well as a plasmid vector and an application of the strong promoter. Function verification is performed by amplifying one strong promoter in ensifer adhaerens, and the strong promoter which can be widely applied to gene expression, gene operation and strain improvement of alpha-proteobacteria such as sinorhizobium meliloti, zymomonas mobilis, caulobacter crescentus, pseudomonas denitrificans, agrobacterium tumefaciens, brucella abortus, pseudomonas fluorescens, rhizobium leguminosarum and ensifer adhaerens is obtained and has the nucleotide sequence being SEQ ID NO:1. The invention also relates to the plasmid vector containing the strong promoter, a method for constructing a gene engineering strain by use of the promoter, the corresponding strain and an application of a host cell in starting target gene expression.

The invention discloses a xylose-induced promoter. By amplifying a xylose-induced promoter in sinorhizobium meliloti, bioinformatics analysis and function verification are carried out, the xylose-induced promoter which can be widely applied to gene expression, genetic gene operation and strain improvement in alpha-proteobacteria such as Sinorhizobium meliloti, Zymomonas mobilis, Caulobacter crescentus, Pseudomonas denitrificans, Agrobacterium tumefaciens, Brucella abortus, Pseudomonas fluorescens, Rhizobium leguminosarum and Sinorhizobium adhaerens is obtained, and the nucleotide sequence of the promoter is SEQ ID NO: 1 or SEQ ID NO: 2. The invention also relates to a plasmid vector containing the xylose-induced promoter, a method for constructing a genetic engineering strain by using thepromoter, a corresponding strain, and application of a host cell to induction and starting of expression of a target gene under xylose inducible conditions.

The invention discloses a composition used for killing pests. The composition is composed of a component A and a component B in a weight ratio of (1-20):(1-10), wherein the component A is validamycin,and the component B is tea saponin. The invention also discloses another composition; and on the basis of the above composition, a component A is composed of validamycin and pseudomonas denitrificans, and a weight ratio of the validamycin to the pseudomonas denitrificans is 3:1. The compositions provided by the invention can improve targeting on pests, reduce damage to natural enemies, slow downgeneration of resistance, and achieve a synergistic effect; and the compositions can promote crop growth and improve crop quality.

The present invention relates to a promoter system inducing expression of 3-hydroxypropionic acid (3-HP) and a method of biologically producing 3-HP using the same. To improve production of 3-HP in a biological manner, continuous synthesis of new enzymes having enzyme activity is necessary. As a result of screening 3-HP reactive transcription regulators and 3-HP reactive promoters from several microorganisms including Pseudomonas denitrificans, it was confirmed that the transcriptions regulations and promoters are composed of LysR proteins and particular gene nucleotide sequences binding to the LysR proteins. Therefore, the 3-HP inducible system is expected to be effectively used to regulate 3-HP metabolic pathways.

This invention consists of a consortium of plant-growth promoting microorganisms including Pseudomonas stutzeri, Pseudomonas denitrificans, Pseudomonas resinovorans, Pseudomonas brassicacearum, Pseudomonas fluorescens, Shimwellia blattae and Klebsiella oxytoca. The bacterial consortium along with a vehicle suitable for agricultural application, form a biofertilizer useful to enhance agricultural yield when applied to cultivation plants. The biofertilizer described present a long shelf life and preserve a high cell density along the time.

The invention discloses pseudomonas denitrificans having the preservation number of CGMCC No.10845. The invention also discloses a microbial inoculum comprising the pseudomonas denitrificans. The invention further discloses an application of the pseudomonas denitrificans and the microbial inoculum in degradation of quinclorac. The invention also discloses a method for degradation of quinclorac. The pseudomonas denitrificans is characterized in that: (1) only an inorganic salt culture medium is provided, and the pseudomonas denitrificans can survive and grow in a culture environment containing quinclorac as a sole carbon source, and has the advantages of strong vital force, fast growth speed, and large living weight; (2) during preparation, only a preservation strain needs to be activated, the operation is simple and the cost is low; and (3) the pseudomonas denitrificans has high degradation effect in water bodies and soil, in the water bodies, under a condition that the content of quinclorac is 200 mg / L, the bacteria are cultured for 24 h, the degradation rate is as high as 93.4%, and in the soil, under a condition that the content of quinclorac is 20 mg / kg, the degradation rate is as high as 88.0%.

The invention relates to a culture medium for producing vitamin B12 by fermenting pseudomonas denitrificans and a fermentation method thereof, and a method for producing vitamin B12 by the same. The culture medium comprises a primary seed culture medium, a secondary seed culture medium and a fermentation culture medium all of which contain artificial molasses composed of maltose, thiamine, riboflavin and biotin. By substituting the artificial molasses composed of maltose, thiamine, riboflavin and biotin for beet molasses and optimizing the formula of the culture medium, the invention solves the problem that the fluctuation range of fermentation titer is large because the quality of beet molasses is unstable, and acquires a method for stably and effectively producing vitamin B12. The invention saves the doses of raw and auxiliary materials, lowers the comprehensive cost and reduces the waste water discharge.

A brewing method of soy sauce containing vitamin B12 comprises the following steps: inoculating a pseudomonas denitrificans strain into a seed liquid culture medium, and culturing to obtain a pseudomonas denitrificans seed solution; cooking extruded soybean meal to obtain a koji material; and mixing the koji material, aspergillus oryzae spore powder and the pseudomonas denitrificans seed solution, fermenting to obtain koji, preparing mature sauce mash, and filtering to obtain the soy sauce containing vitamin B12. According to the present invention, the prepared soy sauce is red brown in color and pure in sauce fragrance; and the fermented soy sauce contains the vitamin B12 produced by metabolism of pseudomonas denitrificans, and is a fermented soy sauce for strengthening the nutrition of the vitamin B12.

The construction method of the pseudomonas denitrificans lacking malonic semialdehyde bypass metabolic pathway comprises the following steps: (1) starting from the constructed pseudomonas denitrificans ZAP01, respectively knocking out mmsAI, mmsAII, mmsAIII and BAPAT genes in genomic DNA (Deoxyribonucleic Acid) of the strain independently and in a combined manner to obtain a plurality of mutant strains; (2) by constructing a plasmid pUCPK-AM, overexpressing a 3-HP synthesis path related enzyme to convert acetic acid to produce 3-HP; and (3) screening on an LB plate containing 30mg / L kanamycin to obtain a target mutant recombinant bacterium. According to the method disclosed by the invention, malonic semialdehyde dehydrogenase and beta-alanine-pyruvate transaminase are knocked out from pseudomonas denitrificans, so that a malonic semialdehyde branch metabolic pathway is completely blocked, and more malonic semialdehyde is used for synthesizing 3-HP, so that the yield of 3-HP synthesized by acetic acid is increased.

The invention relates to a composite material for removing nitrogen and phosphorus in wastewater, which comprises composite particles containing Pseudomonas denitrificans ATCC 13867 and Acinetobacter johnsonii ATCC 17909; the composite particles include the following components in parts by weight: 25-45 parts of iron tailings, 15-30 parts of steel slag powder, 15-30 parts of pine bark powder and 10-15 parts of clay. The present invention also relates to a method for treating wastewater by using the above-mentioned composite material. The invention utilizes the composite material processed by solid waste iron tailings, steel slag, etc. to simultaneously remove nitrogen and phosphorus in wastewater, and the processed composite material can be used to make organic fertilizer or seedling substrate, and simultaneously realize wastewater treatment and solid waste The dual purpose of resourcing.

The present invention relates to the field of soil remediation, and discloses an in-situ remediation preparation for quinclorac contaminated soil, the in-situ remediation preparation for quinclorac contaminated soil comprises fungi and / or bacteria with quinclorac degrading capability; the fungi are at least one of Aspergillus niger, rhizomucor variabilis and cunninghamella bertholletiae, and the bacteria are at least one of Pseudomonas denitrificans, Burkholderia caribensis and Serratia marcescens. The in-situ remediation preparation is used for in-situ bioremediation of the quinclorac contaminated soil, the problems that photochemical degradation method is vulnerable to the restriction of light conditions and secondary chemical reagent pollution of farmlands is easy to cause are overcome, the fungi and / or the bacteria are fully used for decomposition metabolism of residual quinclorac in the soil, the problems that the highly residual quinclorac in the soil seriously endanger food safety can be overcome, the micro-ecology of the soil is improved, the farming ability of the soil is faster to recover, and the method is low cost, and has broad application prospects.

The invention discloses metabolic engineering pseuomonas denitrifican for producing succinic acid as well as a construction method and the application of the pseuomonas denitrifican and belongs to thefields of genetic engineering and fermentation engineering. The construction method comprises the following steps: knocking out genes sdhAB and / or pta-ackA from genome DNA of pseuomonas denitrifican,over-expressing acetyl coenzyme A synthetase genes acs derived from the pseuomonas denitrifican in the pseuomonas denitrifican, and performing aerobic fermentation on the constructed pseuomonas denitrifican by taking acetic acid as a unique raw material, so as to obtain the succinic acid. According to the method disclosed by the invention, the acetic acid serves as the raw material, and the succinic acid is biologically synthesized by utilizing an acetate metabolism intermediate product acetylcoenzyme A. Compared with the prior art, the metabolic engineering pseuomonas denitrifican disclosedby the invention has the advantages that the acetylcoenzyme A serves as a metabolic intermediate, the raw material source is wide, and the cost is low.

The invention provides a microbial agent and a preparation method and application thereof. The microbial agent comprises the ingredients: Microbacterium sp. CICC 23615, Pseudomonas denitrificans CCTCC M91003, Deinococcus sp. CICC 10697, Bacillus sp. CICC 21937, Pichia pastoris ACCC 21021, and Aspergilus flavus CGMCC 3.2823. The microbial agent provided herein can remove ammonia and nitrogen in wastewater effectively in a biological treatment stage, up to about 97% in removal rate; in early startup stage of a system or in case of unstable operation, the microbial agent has excellent impact resistance. The microbial agent is free of pathogens, heavy metals and toxic and harmful chemicals, and solid products of the microbial agent are nontoxic and nonirritating.

In the present invention, by amplifying a strong promoter in S. cohescens, bioinformatics analysis and functional verification are carried out to obtain a promoter that can be widely used in Sinorhizobium meliloti, Zymomonas mobilis, Caulobacter crescentus, and Pseudomonas denitrification. Strong promoters for gene expression, genetic manipulation and strain improvement in α-Proteobacteria such as A. The nucleotide sequence is SEQ ID NO:1. The present invention also relates to a plasmid vector containing the strong promoter, a method for constructing a genetically engineered bacterial strain using the promoter, a corresponding bacterial strain, and the application of host cells in promoting the expression of an objective gene.

The invention relates to a preparation method of a cobamamide crude product. A process comprises the following steps: carrying out spray drying on pseudomonas denitrificans fermentation liquor to prepare cobamamide mycelium powder, performing extraction by using an acetone composite solvent, performing centrifugal separating and concentrating under reduced pressure to obtain a cobamamide concentrated solution I, performing adsorbing by using macroporous resin, performing desorbing by using an acetone aqueous solution, performing concentrating under reduced pressure to obtain a cobamamide concentrated solution II, and finally, carrying out flocculation filtration, acetone crystallization, suction filtration and vacuum drying to obtain the cobamamide crude product. According to the invention, the pseudomonas denitrificans fermentation liquor is subjected to the spray drying to obtain the cobamamide mycelium powder; the acetone composite solvent is used for the extraction, so that the situation that when the pseudomonas denitrificans fermentation liquor is abnormal and plate-frame filtration is difficult is avoided, cobamamide is recycled to the maximum extent, and the product qualityand yield are improved. The method has the characteristics that: the process operation is simpler and more convenient, the production schedule is easy to guarantee, the production operation coherenceis high, the generated wastewater amount is small, and the like.

The invention provides a microbial agent for treating sludge of a sewage plant. The microbial agent is prepared from the following components in parts by weight: 50-60 parts of clostridium thermophile, 25-30 parts of pseudomonas denitrificans, 25-30 parts of rhodospirillum rubrum, 15-20 parts of denitrifying bacteria and 15-20 parts of saccharomycetes through strain activation, triangular flask culture medium preparation, culture and fermentation. The microbial agent disclosed by the invention is easy to obtain and can efficiently treat sludge of a sewage plant.