36 results about "UDPglucose pyrophosphorylase" patented technology

The subject invention pertains to novel mutant polynucleotide molecules that encode enzymes that have increased heat stability. These polynucleotides, when expressed in plants, result in increased yield in plants grown under conditions of heat stress. The polynucleotide molecules of the subject invention encode maize endosperm ADP glucose pyrophosphorylase (AGP) and soluble starch synthase (SSS) enzyme activities. Plants and plant tissue bred to contain, or transformed with, the mutant polynucleotides, and expressing the polypeptides encoded by the polynucleotides, are also contemplated by the present invention. The subject invention also concerns methods for isolating polynucleotides and polypeptides contemplated within the scope of the invention. Methods for increasing yield in plants grown under conditions of heat stress are also provided.

Compositions and methods for improving plant growth are provided herein. Polynucleotides encoding ADP-glucose pyrophosphorylase small subunit (AGPaseSS) proteins, polypeptides encompassing AGPaseSS proteins, and expression constructs for expressing genes of interest whose expression may improve agronomic properties including but not limited to crop yield, biotic and abiotic stress tolerance, and early vigor, plants comprising the polynucleotides, polypeptides, and expression constructs, and methods of producing transgenic plants are also provided.

The invention discloses a corn endosperm ADP-glucose pyrophosphorylase mutant, encoding genes thereof and application of the same two. The protein can be the protein (1) formed by the first amino acid sequences showed as in a sequence table and can also be the protein (2) which is formed by substituting and / or deleting the first amino acid residue sequences showed as in a sequence table and / or adding one or several amino acid residues to the first amino acid residue sequences, has ADP-glucose pyrophosphorylase activity and is derived from the protein (1). The corn endosperm ADP-glucose pyrophosphorylase mutant and the encoding thereof play important roles in the corn variety improvement.

The invention concerns the use of duplex oligonucleotides about 25 to 30 base pairs to introduce site specific genetic alterations in plant cells. The oligonucleotides can be delivered by mechanical (biolistic) systems or by electrpoporation of plant protoplasts. Thereafter plants having the genetic alteration can be generated from the altered cells. In specific embodiments the invention concerns alteration in the gene that encode acid invertase, UDP-glucose pyrophosphorylase, polyphenol oxidase, O-methyl transferase, cinnamyl alcohol dehydrogenase, ACC synthase and ACC oxidase or etr-1 or a homolog of etr-1, and plants having isolated point mutations in such genes.

The invention relates to the field of a genetic engineering technology, and particularly relates to a kelp uridine diphosphate (UDP)-glucose pyrophosphorylase (UGPase) gene. The nucleotide sequence of the gene and the amino acid sequence of the coding protein are SEQ ID No.1 and SEQ ID No.2 respectively. According to the invention, the gene sequence is cloned through the gene cloning technology, and a prokaryotic expression vector is established; and enzyme activity detection on recombinant protein proves that the gene has the function of catalyzing UDP-glucose and pyrophosphoric acid to form glucose-1-phosphoric acid and UTP and belongs to a key enzyme coding gene of the synthesis path of agar-agar, starch, cellulose, trehalose, sucrose and the like. The gene has an important application value in increasing the content of economic components including algae agar-agar, starch, cellulose, trehalose, sucrose and the like.

The invention relates to an engineering bacterium with overexpressed uridine diphosphoglucose pyrophosphorylase gene and establishment thereof, belonging to the technical fields of gene engineering and microbial fermentation. By cloning the key enzyme uridine diphosphoglucose pyrophosphorylase gene in the polysaccharide flocculant synthesis route, an Escherichia coli-Bacillus shuttle plasmid is utilized to establish a recombinant expression vector. The recombinant plasmid is transformed into Bacillus licheniformis by electric transformation to establish recombinant Bacillus licheniformis HN301-2; and compared with the initial strain, the polysaccharide flocculant yield of the recombinant Bacillus licheniformis is enhanced by 15.6%, and the fermentation liquid flocculation activity is enhanced by 70%. The engineering bacterium is hopeful to be used in industrial production of polysaccharide flocculants, and can enhance the yield and lower the cost.

The invention provides an ADP-glucose pyrophosphorylase mutant and a screening method and application thereof. The screening method includes the steps of firstly, synthesizing first chain cDNA; secondly, performing PCR amplification; thirdly, building AGPase large and small subunit cDNA prokaryotic expression vectors; fourthly, performing active screening. The method has the advantages that distant hybridization is performed on AGPase large subunit genes and small subunit genes from different species, and the recombinant gene expression vectors are obtained in a highly-controllable manner; escherichia coli glgC mutant strains are converted and inoculated to a Conberg enrichment solid culture medium, and iodine-potassium iodide dyeing is used to screen the high-activity ADP-glucose pyrophosphorylase mutant. The screening method is simple and practical and stable and reliable. The enzyme activity and glycogen content of the ADP-glucose pyrophosphorylase mutant are evidently higher than those of wild corn AGPase, evident change of the substrate affinity of the ADP-glucose pyrophosphorylase mutant is avoided, and the sensitivity of the ADP-glucose pyrophosphorylase mutant to activating agent is increased.

The invention discloses a maize-endosperm ADP-glucose pyrophosphorylas mutant and sieving method and application, which comprises the following steps: 1) mutating cDNA of large and small subunit of maize-endosperm AGPase through alcaine hydroxylamine mutation agent randomly; 2) transmitting mutated cDNA and wild-type cDNA into escherichia coli glgC mutation germ; sieving positive monoclonal; 3) seeding positive monoclonal line on the Conberg rich solid board to culture under 35-39 deg.c; 4) dyeing; comparing the color; obtaining deep color mutation as maize-endosperm ADP-glucose pyrophosphorylas mutant.

The invention discloses a method for improving germination and growth of corn seeds and relates to the field of crop planting. The method aims to solve the technical problems of low germination rate and long germination period of existing corn seeds. The method comprises the steps of 1, sterilizing the surfaces of the seeds; 2, preparing a seed soaking agent; 3, performing vibrated seed soaking treatment. The method can shorten the average time of seed germination and increase the germination rate, germination vigor, germination index and vigor index of the seeds. The growth and elongation ofa root system and germs are boosted; the activity of hexokinase and uridine diphosphate glucose pyrophosphorylase is obviously increased, the activity of aldolase is reduced, the activity of phosphofructokinase is maintained at a relatively normal level, the glycolysis metabolism and the biosynthesis process of cell walls are promoted, and metabolic products and energy are provided for the germination and growth of the seeds. The method is used for promoting the germination and growth of the corn seeds.

The invention discloses a transgenic method for enhancing sink strength in female reproductive organs. It involves the expression of nucleic acids encoding acid invertase or ADP-glucose pyrophosphorylase floral tissue. The invention also includes expression constructs used in the method.

The invention discloses a method for synthesizing UDP-galactose and a method for synthesizing a galactosyl compound, and belongs to the technical field of bioengineering. The invention relates to a method for generating UDP-galactose by catalyzing UTP, ATP and D-galactose through a thermophilic enzyme method two-step reaction. The method comprises the following two steps: i) taking D-galactose and ATP as substrates, and catalytically synthesizing 1-P-Gal (1-phosphate-galactose) by using thermophilic galactokinase TTHA0595; and (ii) by taking the 1-P-Gal and the UTP as the substrates, carrying out catalytic synthesis on the UDP-galactose by using the thermophilic UDP-glucose pyrophosphorylase TTE0732. The synthesized UDP-galactose is used as a glycosyl donor, and thermophilic galactosyl transferase TON1857 can be used for further catalyzing substrate compounds such as vanillin, p-nitrophenol or silybin to synthesize the galactosyl compound, so that the method for synthesizing the galactosyl compound by the thermophilic enzyme method is constructed.

The invention provides for sequencing a nucleic acid molecule based on the detection of base incorporation by the release of pyrophosphate (PPi) using a new enzyme system comprising adenosine diphosphate (ADP)-glucose pyrophosphorylase (AGPase) and its substrate ADP-glucose.

The subject invention pertains to novel mutant polynucleotide molecules that encode enzymes that have increased heat stability. These polynucleotides, when expressed in plants, result in increased yield in plants grown under conditions of heat stress. The polynucleotide molecules of the subject invention encode maize endosperm ADP glucose pyrophosphorylase (AGP) and soluble starch synthase (SSS) enzyme activities. Plants and plant tissue bred to contain, or transformed with, the mutant polynucleotides, and expressing the polypeptides encoded by the polynucleotides, are also contemplated by the present invention. The subject invention also concerns methods for isolating polynucleotides and polypeptides contemplated within the scope of the invention. Methods for increasing yield in plants grown under conditions of heat stress are also provided.

The invention discloses an overexpressed UGPase (UDP-glucose pyrophosphorylase) gene and a method for constructing recombinant lactobacillus acidophilus thereof. The method is characterized by comprising the specific steps as follows: (1) a UGPase gene LBA0625 is amplified with lactobacillus acidophilus genome DNA as a template; an expression vector pMG36e is subjected to single restriction digest with restriction enzyme Hind III, and after dephosphorylation and gel recovery are performed, a recombinant expression vector is constructed by inserting a PCR amplification product into the expression vector pMG36e; (2) an overexpression plasmid subjected to extraction and concentration is imported into lactobacillus acidophilus through electrotransformation, the lactobacillus acidophilus is resuscitated at 37 DEG C for 2 h, applied to an erythrocin resistant plate and cultured at 37 DEG C for 36 h for screening transformants; the transformants are subjected to PCR verification, and the recombinant lactobacillus acidophilus PLY127-1 of the overexpressed UGPase gene is obtained. The freeze-drying survival rate of the lactobacillus acidophilus can be increased substantially.

The invention relates to construction and application of an engineering strain for biologically synthesizing momordica grosvenori glucoside V by taking mogrol as a substrate. The method comprises the following steps: trans mogrol by a recombinant cell containing a glycosyl transferase gene or a recombinant cell containing the glycosyl transferase gene and a UDP-glucose pyrophosphorylase gene to obtain the momordica grosvenori glycoside V. The recombinant cell expresses the glucose glycosyl transferase gene, the UDP-glucose pyrophosphorylase gene, the UGT enzyme and the UDP-glucose pyrophosphorylase. According to the invention, exogenous species UGT enzyme gene and UDP-glucose pyrophosphorylase gene are introduced through a gene recombination technology, the exogenous species UGT enzyme gene and UDP-glucose pyrophosphorylase gene are transformed into genetically engineered bacteria for expression, a new biosynthesis path of the momordica grosvenori glycoside V is reconstructed. Through overexpression of the UGT enzyme gene and the UDP-glucose pyrophosphorylase gene, the mogroside V is biologically synthesized by taking mogrol as a substrate, and the synthesis yield is high.

Disclosed herein are a novel sucrose-inducible promoter sequence and a 5′ untranslated region which are derived from sweetpotato ADP-glucose pyrophosphorlyase gene (ibAGPl) (SEQ ID NO: 1). Also disclosed are expression vectors using the same sequences and a transgenic plant using the same vectors. The promoter and 5′ untranslated region according to the present invention can confer a high level of sucrose-inducible expression in plants, particularly in plant storage roots which contain sucrose in relatively large quantities to accumulate starch in large quantities in plants. Therefore the present invention may be useful for the generation of transgenic plants to produce useful proteins in large quantities in plant storage roots.

The invention discloses a paulownia uridine diphosphate glucose pyrophosphorylase gene and application thereof. Amino acid residue sequence of the paulownia uridine diphosphate glucose pyrophosphorylase gene is shown in SEQ ID No. of 1. The information can be used for constructing the paulownia uridine diphosphate glucose pyrophosphorylase gene, or antisense gene and RNA disturbing mass of the gene, which can be used for improving plant materials and constructing new materials with higher or lower cellulose content.

The invention provides an ADP-glucose pyrophosphorylase mutant and a screening method and application thereof. The screening method includes the steps of firstly, synthesizing first chain cDNA; secondly, performing PCR amplification; thirdly, building AGPase large and small subunit cDNA prokaryotic expression vectors; fourthly, performing active screening. The method has the advantages that distant hybridization is performed on AGPase large subunit genes and small subunit genes from different species, and the recombinant gene expression vectors are obtained in a highly-controllable manner; escherichia coli glgC mutant strains are converted and inoculated to a Conberg enrichment solid culture medium, and iodine-potassium iodide dyeing is used to screen the high-activity ADP-glucose pyrophosphorylase mutant. The screening method is simple and practical and stable and reliable. The enzyme activity and glycogen content of the ADP-glucose pyrophosphorylase mutant are evidently higher than those of wild corn AGPase, evident change of the substrate affinity of the ADP-glucose pyrophosphorylase mutant is avoided, and the sensitivity of the ADP-glucose pyrophosphorylase mutant to activating agent is increased.

The invention concerns the use of duplex oligonucleotides about 25 to 30 base pairs to introduce site specific genetic alterations in plant cells. The oligonucleotides can be delivered by mechanical (biolistic) systems or by electroporation of plant protoplasts. Thereafter plants having the genetic alteration can be generated form the altered cells. In specific embodiments the invention concerns the alteration in the gene that encode acid invertase, UDP-glucose pyrophosphorylase, polyphenol oxidase, O-methyl transferase, cinnamyl alcohol dehydrogenase, ACC synthase and ACC oxidase or etr-1 or homolog of etr-1, and plants having isolated point mutations in such genes.

The invention relates to the field of microbial synthesis, in particular to construction and application of an engineering strain for biosynthesizing resveratrol glucoside by taking resveratrol as a substrate. The genetically engineered bacterium expresses a glycosyl transferase (UDP-glucose transferase, UGT) gene and a UDP-glucose pyrophosphorylase gene, a UGT enzyme and a UDP-glucose pyrophosphorylase are produced, a beta-glucosidase gene is knocked out at the same time, and beta-glucosidase is not expressed. The method has the advantages of simple operation, no environmental pollution, high yield, high purity and low cost, and provides a basis for industrial production of resveratrol glucoside.

The invention discloses a corn endosperm ADP-glucose pyrophosphorylase mutant, a screening method and application thereof. The screening method comprises the following steps: 1) using hydroxylamine hydrochloride mutagen to randomly mutate the large and small subunit cDNA of corn endosperm AGPase under the condition of 70-80°C warm bath; The wild-type maize endosperm AGPase large and small subunit cDNAs were transformed into Escherichia coli glgC mutant strains, and positive single clones were screened; 3) The positive clones were streaked and inoculated on a Conberg enriched solid culture plate, and cultured at 35-39°C; 4) Use iodine-potassium iodide solution to stain the single colony grown on the Conberg enriched solid culture plate, and compare the color with the control colony. The mutant strain is darker than the control color. After sequencing verification, the corn endosperm ADP- Glucose pyrophosphorylase mutants. The maize endosperm ADP-glucose pyrophosphorylase mutant and its screening method of the present invention will play an important role in the improvement of maize varieties.

The invention discloses genetically engineered bacteria and application thereof to preparation of rebaudioside A. The genetically engineered bacteria capable of producing stevia rebaudiana glycosyl transferase UGT76G1 and UDP-glucose pyrophosphorylase UGPase are obtained through construction, and perform overexpression on the UDP-glucose pyrophosphorylase UGPase on the basis of performing constitutive expression on recombinant bacteria of the stevia rebaudiana glycosyl transferase UGT to increase the glycosyl donor quantity of glycosylation reaction. A whole-cell catalytic reaction system is established, and stevioside is converted into the rebaudioside A. The dosage concentration of the bacteria is 1 to 10 g / L, the dosage of glucose is 20 to 120 g / L and the dosage of the substrate stevioside is 1 to 20 g / L. In the catalytic reaction process, a cell catalyst is subjected to surfactant treatment, the cell permeability is improved and the yield of the rebaudioside A is greatly increased. A synthetic route of intracellular UDPG is utilized, so the glycosyl donor quantity of the intracellular UDPG is increased, additional addition is avoided and regeneration cycle is realized; and cultivation of yeast and expression of recombinase are conducted simultaneously, so the cycle of the cultivation of the recombinant bacterial and the expression of the recombinase is shortened.

The invention discloses recombinant saccharomyces cerevisiae for producing ginsenoside CK by metabolizing glycerol and a construction method thereof, and the construction method comprises the following steps: (1) introducing a glycosyl transferase UGT1 gene expression cassette, a glucose phosphate mutase PGM2 gene expression cassette and a UDP glucose pyrophosphorylase UGP1 gene expression cassette into saccharomyces cerevisiae capable of synthesizing protopanoxadiol to obtain recombinant bacteria 1; (2) introducing a glycerol transport protein CjFPS1 gene expression cassette, a glycerol dehydrogenase OpGDH gene expression cassette and a dihydroxyacetone kinase DAK1 gene expression cassette into the recombinant bacterium 1 to obtain a recombinant bacterium 2; (3) introducing a 3-hydroxy-3-methylglutaryl coenzyme A reductase HMGr gene expression cassette into the recombinant bacterium 2 to obtain a recombinant bacterium 3; experiments prove that the yields of the ginsenoside CK of the recombinant bacteria 1, 2 and 3 are 109.84 mg / L, 152.12 mg / L and 330.94 mg / L in sequence.