41 results about "Glutamine synthetase gene" patented technology

The invention relates to the field of genetic engineering, and specifically discloses a gene expression system for producing recombinant protein by using CHO cells, and an eukaryotic expression vector, wherein the eukaryotic expression vector comprises a GS expression unit, and the GS expression unit comprises a TK promoter gene sequence aligned along a 5' to 3' direction, a glutamine synthetase gene sequence and a SV40polyA gene sequence. The expression system comprises the eukaryotic expression vector and CHO cells. With the expression system, integration and efficient expression of exogenous gene in CHO cell genome can be achieved, and broad application prospects are provided in the field of protein.

The invention discloses LAMP (loop-mediated isothermal amplification) primer composition for detecting colletotrichum gloeosporioides and an application of the LAMP primer composition. A GS (glutamine synthetase) gene is used as a target gene, primers which have high specificity and high sensitivity and can be used for LAMP detection of the colletotrichum gloeosporioides are designed and screened out, and the LAMP primer composition consists of a forward outer primer GS-F3 shown in SEQ ID NO.2, a reverse outer primer GS-B3 shown in SEQ ID NO.3, a forward inner primer GS-FIP shown in SEQ ID NO.4, a reverse inner primer GS-BIP shown in SEQ ID NO.5, a loop primer GS-LF shown in SEQ ID NO.6 and a loop primer GS-LB shown in SEQ ID NO.7. The primer composition is mainly used for quickly detecting the colletotrichum gloeosporioides, and only 2 hours are spent on detection each time.

The invention relates to a special plant expression vector pH2-35S-PrbcS-GS1 which comprises an arabidopsis thaliana cytoplasm glutamine synthetase gene GS1 and can improve the utilization rate of a plant nitrogen element. A method of RT-PCR is used for cloning the GS1 gene from the arabidopsis thaliana of a model plant, a photoinduction type promotor (the promotor of a a small subunit Rubisco) is used for controlling the excessive expression of the GS1 gene in a plant leaf and a leaf disc conversion method is used for transferring the GS1 gene into a pPZP221-PrbcS-Dof1 type transgene tobacco. An experiment result shows that the GS1 gene can be normally transferred in the transgene tobacco; under the nutrition condition of low nitrogen and the growing conditions of indoor irradiation for 24 hours of 2000LUX and 25 DEG C, the growing situation of the plant transferred with the single gene of Dof1 is (the expression of the gene is controlled by the photoinduction type promotor Prbcs) is a little better than that of a contrast tobacco (a wild type without transgene); after being transferred under the natural growing condition of a green house, the growing situation of the tobacco which is simultaneously transferred with the GS1 gene and the Dof1 gene shows remarkable growing advantages than that of the contrast plant; and therefore, simultaneously and excessively expressing the GS1 gene and the Dof1 gene, can improve the efficiency of the GS / GOGAT (glutamine synthetase / glutamic acid synthetase) approaches in the leaf more extensively, thereby improving the utilization rate of the plant nitrogen element. The vector can be broadly applied to the molecule breeding of crops, improving the utilization rate of the plant nitrogen element thereof and the durability to the nutrition condition of low nitrogen and being capable of obtaining a higher yield under the conditions of applying less fertilizers and even not applying the fertilizers.

The invention provides high-nucleoside-yield bacillus subtilis engineering bacterium as well as a construction method and application thereof The invention also provides a method for producing nucleoside by fermentation. The method comprises the following steps of (1) enhancing a glutamate synthase gene gltA for encoding an NCBI reference sequence WP-009967365.1 on a bacillus subtilis chromosome;and / or enhancing a glutamine synthetase gene glnA for encoding an NCBI reference sequence WP-003231737.1 on the chromosome of the bacillus subtilis; and (2) applying the gene-enhanced strain obtainedin the step (1) to fermentation production of nucleoside. The bacillus subtilis engineering bacterium provided by the invention is a high-nucleoside-yield strain, can effectively accumulate nucleoside, improves the yield of nucleoside, and lays a foundation for industrial production of nucleoside.

The invention discloses a glutamine synthetase and its special-purpose expression project bacterium and its allocation, which is to provide a glutamine synthetase and its special-purpose expression project bacterium and its allocation. The said glutamine synthetase is prepared by abrupt change of the 405th amino acid residue of amino end to phenylalanine corynebacterium glutamicum. The engineering bacteria prepare the recombination bacteria by importing the recombination expression carrier containing the said glutamine synthetase gene into host bacteria. The glutamine synthetase produced by the engineering bacteria can specially act on NH4+ and aminoglutaric acid, the durability to the high concentration ammonium ion has been distinctively improved because of the dissolution of adenosine acylation modification, the major bottleneck of glutamine with enzymatical production has been solved, and the concentration of the obtained glutamine can reach 47.6g / L. The invention has higher practicability and flackery and has wide application foreground in glutamine commercial manufacture.

The invention discloses a special plant expression vector pK2-35S-Prbcs-*T-GS2 which contains an arabidopsis chloroplast type glutamine synthetase gene (GS2) and can improve the metabolic ability of plant nitrogen. A RT-PCR method is utilized to clone a GS2 gene from a model plant Arabidopsis, a promoter of a Rubisco small subunit with the light-inducible promoter is used to control the overexpression of the GS2 gene in plant leaves, and the GS2 gene is transformed into wild tobacco through a leaf disk transformation method. The experimental result shows that the GS2 gene can perform normal transcription in transgenic tobacco and the growth condition of a strain with the GS2 gene is better than that of a check plant (un-transformed wild type) under low nitrogen nutrition condition, which indicates that the overexpression of the GS2 gene can improve the efficiency of a GS / GOGAT (glutamine synthetase / glutamate synthetase) way to a great extent, thereby improving the capability of the assimilation of the plant nitrogen. The special vector has the advantages that the special vector can be used for molecular breeding of crops, improves the utilization rate of nitrogen and the tolerance to low nitrogen stress, and obtains higher yield under the condition of small nitrogenous fertilizer application or even no nitrogenous fertilizer application.

The invention belongs to the technical field of gene engineering, and particularly relates to a glutamine synthetase gene with glufosinate tolerance and application thereof. The sequence and coding region of the glutamine synthetase gene exiGS with glufosinate tolerance are disclosed, the nucleotide sequence of the gene is shown in SEQ ID NO:1, and 446 amino acids are coded. The sequence of protein coded by the gene is shown in SEQ ID NO:2. Biological function verification proves that the gene is high in activity, resistant to low temperature and high in high-temperature stability. The functions and the potential application to anti-glufosinate crops of the glutamine synthetase gene with glufosinate tolerance are verified preliminarily. The gene can be widely applied to different herbicide-resistant transgenic plants.

The invention discloses a GS-DHFRmut double-gene screening expression vector, and a preparation method and an application thereof, and belongs to the biotech field. The expression vector contains a promoter controlling the expression of a glutamine synthetase gene, the glutamine synthetase gene, a promoter controlling the expression of a mutant dihydrofolate reductase gene, and the mutant dihydrofolate reductase gene, and the downstream of each of two screening genes has a PolyA signal sequence. The invention also discloses the preparation method of the expression vector and the application of the expression vector. The GS-DHFRmut double-gene screening expression vector improves the expression level of an exogenous protein, enlarges the use resource of host cell strains in the genetic engineering, and has an important application value in the industrialized development of the genetic engineering.

The invention relates to a bacillus subtilis glutamine synthetase gene as well as an encoding protein and cloning method thereof. The bacillus subtilis glutamine synthetase gene is from bacillus subtilis, and the sequence of the bacillus subtilis glutamine synthetase gene is a base sequence shown in SEQ ID NO. 1. Glutamine synthetase encoded by the gene can be used for remarkably improving the glutamine synthesis capability of model organism escherichia coli. The disclosed full-length gene and the amino acid sequence have application values in improvement of the utilization rate of nitrogen of organisms and increase of the content of nitrogen in the organisms in genetic engineering.

The invention relates to the field of biotechnology, and in particular, relates to a gene expression system for producing a recombinant protein by using CHO cells and a eukaryotic expression vector. The invention provides the pHLX201 eukaryotic expression vector including a GS expression unit, the GS expression unit has the sequence comprising an SV40L promoter gene sequence and a glutamine synthetase gene sequence which are arrayed successively in a direction from 5' to 3', wherein the SV40L promoter gene sequence is shown in SEQ ID NO:2, and the glutamine synthetase gene sequence is shown in SEQ ID NO:3. The expression system provided by the invention can realize integration and efficient expression of exogenous genes in a CHO cell genome. The preparation method of the system mainly includes that the eukaryotic expression vector is constructed, is suitable for transient expression of a target gene in the CHO cells and is suitable for screening target gene high-expression cell lines. The CHO expression system can be used for realizing integration and expression of the recombinant protein, a monoclonal antibody and the like in the CHO cells, and has wide application prospects.

Cells and cell lines are disclosed that are able to produce therapeutic proteins, antibodies, vectors, and viral vectors such as lentiviral vectors and adeno-associated viral (AAV) vectors. The cells and / or cell lines can have mutations or deletions in either one or both of the endogenous di-hydrofolate reductase (DHFR− / −) or glutamine synthetase (GS− / −) genes such that DHFR and / or GS expression or function is substantially reduced or eliminated.

The invention discloses a gastrodia elata Glutamine Synthetase (GS) gene. The nucleotide sequence of the gene is shown as SEQ ID NO: 1; and 342 amino acid residues are encoded; a prokaryotic expression vector of the gastrodia elata GS gene is constructed, and prokaryotic expression analysis shows that the gastrodia elata glutamine synthetase gene is soluble protein, and the molecular weight of the gene is about 59.94kDa. The eukaryotic overexpression vector of the gastrodia elata glutamine synthetase gene is constructed, gastrodia elata symbiotic bacteria armillaria mellea is transfected through an agrobacterium method, the armillaria mellea transfected with the gastrodia elata glutamine synthetase gene is obtained, the GS genetically engineered armillaria mellea is cultured at 13 DEG C and 28 DEG C, the growth vigor of the GS genetically engineered armillaria mellea is better than that of wild type armillaria mellea, and the growth speed is high. Therefore, the gastrodia elata glutamine synthetase gene is beneficial for shortening the culture time of the armillaria mellea and improving the cold resistance and high-temperature growth resistance of the armillaria mellea. The gastrodia elata glutamine synthetase gene provides an experimental basis for enhancing the cold resistance of gastrodia elata, expanding the planting distribution range of gastrodia elata, enhancing the nitrogen metabolism capacity of gastrodia elata and increasing the yield of gastrodia elata.

The present disclosure is directed to genetically engineered bacteria strains with enhanced biofuel-producing capabilities from cellulosic substrates. The bacteria strains of the present disclosure comprise an inactivated Type I glutamine synthetase gene. The present disclosure is also directed to methods of producing biofuels from cellulosic biomass using the genetically engineered bacteria strains.

The invention provides a glutamine synthetase exogenous expression method based on bacillus megaterium in the technical field of genetic engineering. Glutamine synthetase gene sequences obtained through sequencing are amplified by a primer sequence and connected into an expression vector to obtain a recombinant expression vector, the recombinant expression vector is guided into escherichia coli further, and genetically modified recombinant bacteria containing target genes are screened and obtained. According to the glutamine synthetase exogenous expression method based on the bacillus megaterium, the problem that the using effect is limited severely due to the fact that the expression quantity of glutamine synthetase gene sequences in natural bacillus megaterium is very small is solved; and genetically engineered bacteria are used for expressing the glutamine synthetase gene sequences, so that the expression quantity of the gene is increased greatly.

The invention provides a new glutamine synthetase gene having a nucleotide sequence shown as SEQ ID NO: 8 and further provides a eukaryotic expression vector containing the same. The glutamine synthetase gene can be used in a GS gene amplification system to promote expression of target protein and screening of high-expression cell lines.

The invention discloses a mutant site of a glufosinate-ammonium resisting Eleusine indica species. The mutant site is located at a 175th position of a nucleotide sequence shown in SEQ ID NO: 1 of a target enzyme, i.e., glutamine synthetase gene EiGS1-S cloned from glufosinate-ammonium sensitive Eleusine indica, and generated mutation is A-G. The invention further discloses a primer dCAPs for detecting a gene mutation site of the glufosinate-ammonium resisting Eleusine indica species, a detection method for the glufosinate-ammonium resisting Eleusine indica species and application of the primerin preparation of a kit or biological preparation for detecting the glufosinate-ammonium resisting Eleusine indica species.

The invention relate to a micrococcus glutamine synthetase gene, and encoding protein and a clone method thereof. The gene provided by the invention is derived from micrococcus bacteria, and has a basic group sequence as shown in SEQ ID NO. 1. The gene encodes glutamine synthetase, and can obviously improve the capacity of mode creature colibacillus in the aspect of synthesizing glutamine, which foreshows that the disclosed full-length gene and the amino acid sequence have application value in the gene engineering aspect of improving the nitrogen utilization efficiency of organisms and increasing nitrogen element content of the organisms.

The invention provides a rapid and efficient locating method of a tea tree plasmid type glutamine synthetase gene. The method includes the steps of putting a fresh and tender tea tree leaf sample intoa PBS buffer solution on the same day of a test, slicing the sample into sections with the thickness of 50-100 micron in an oscillator slicer after the sample is embedded in and fixed by sepharose gel, hybridizing the obtained tea tree leaf transverse sections with the two antibodies (tea tree CsGS2 special antibody and commercial antibody), and finally conducting photographing under a laser scanning confocal microscope. By means of the cytology gene locating method, the gene can be rapidly, efficiently and precisely located; the gene locating can reach the cytology level; the cycle requiredin the test process is short; the heterology troubles caused by the tea tree gene locating on a model plant (arabidopsis thaliana) are avoided; safety, operability and high repeatability are realized.

The invention discloses a glutamine synthetase and its special-purpose expression project bacterium and its allocation, which is to provide a glutamine synthetase and its special-purpose expression project bacterium and its allocation. The said glutamine synthetase is prepared by abrupt change of the 405th amino acid residue of amino end to phenylalanine corynebacterium glutamicum. The engineering bacteria prepare the recombination bacteria by importing the recombination expression carrier containing the said glutamine synthetase gene into host bacteria. The glutamine synthetase produced by the engineering bacteria can specially act on NH4+ and aminoglutaric acid, the durability to the high concentration ammonium ion has been distinctively improved because of the dissolution of adenosine acylation modification, the major bottleneck of glutamine with enzymatical production has been solved, and the concentration of the obtained glutamine can reach 47.6g / L. The invention has higher practicability and flackery and has wide application foreground in glutamine commercial manufacture.

The invention relates to an Arthrobacte glutamine synthetase gene, coding protein and cloning method thereof. The gene of the present invention is derived from Acinetobacter bacteria, and SEQ ID NO1 shows the base sequence. The gene coding glutamine synthetase can perform the functions of glutamine synthetase in the Escherichia Coli, and can recover the capability of Escherichia Coli mutant for synthesizing glutamine, and simultaneously indicates application values in the high efficiency utilization aspect of biological nitrogen.

The invention relates to a glutamine synthetase gene of acinetobacter as well as encoding protein and a cloning method of the glutamine synthetase gene. The gene is from acinetobacter bacteria and adopts a base sequence represented by SEQ ID NO.1. The gene encodes glutamine synthetase, can play functions of glutamine synthetase in escherichia coli and can recover glutamine synthesis capacity of escherichia coli mutants, and application values of gene in efficient utilization of other biological nitrogen are further predicted simultaneously.

Disclosed herein are methods and compositions for inactivating the glutamine synthetase (GS) gene using a fusion protein comprising a zinc finger protein and a cleavage domain or half-domain. Also provided is a polynucleotide encoding the fusion protein, a cell comprising the polynucleotide and the fusion protein.

The invention relates to the field of molecular biology, in particular to a bacterial glutamine synthetase and a preparation method thereof. The preparation method of bacterial glutamine synthetase comprises the following steps of: 1) amplifying a base sequence shown in SEQ ID NO.2 by using a PCR (polymerase chain reaction) technology; 2) cloning the base sequence shown in SEQ ID NO.2 into a vector plasmid to construct an expression vector of bacterial glutamine synthetase; and 3) transforming the expression vector into competent thalli of Escherichia coli to obtain recombinant thalli, and obtaining bacterial glutamine synthetase protein through the sequential processes of expanding culture, inducing expression, collecting thalli, crushing the thalli and purifying protein. The method for inducing and expressing bacterial glutamine synthetase protein by using an Ecoli BL21-pET21a-GS recombinant strain is simple and easy to operate, and finds a novel glutamine synthetase gene sequence, thereby providing a new research direction for the research of glutamine synthetase.

The invention discloses a tea tree glutamine synthetase gene SSR molecular marker (SSR1-GS, SSR2-GS) primer and application of the tea tree glutamine synthetase gene SSR molecular marker primer. Researches show that the method for developing the SSR primer on the whole genome of the tea tree according to specific genes is feasible, and the obtained primer is high in polymorphism and good in repeatability and has good feasibility and pertinence. Therefore, the inventor develops research on genetic polymorphism of tea tree germplasm resources. In conclusion, the research method, the SSR molecular marker and the primer of the SSR molecular marker can be widely applied to tea tree variety identification, genetic structure and resource diversity analysis, genetic map construction, functional gene positioning and QTL positioning, seed purity identification and molecular marker-assisted seed selection research, and tea tree theanine synthesis and tea tree genetic polymorphism can be further researched conveniently; deep development of tea tree resources is promoted.

The invention discloses a glutamine synthetase gene derived from achromobacter xylosoxidans and an application of the gene. The glutamine synthetase gene comprises 1413 basic groups and 471 coded amino acids. The nucleotide sequence of the gene is as shown in SEQ ID NO 1, and the sequence of the coded amino acids is as shown in SEQ ID NO 2. The glutamine synthetase gene derived from the achromobacter xylosoxidans has high glufosinate-ammonium tolerance and can be used for cultivating glufosinate-ammonium resistant transgenic crops.

The present disclosure is directed to genetically engineered bacteria strains with enhanced biofuel-producing capabilities from cellulosic substrates. The bacteria strains of the present disclosure comprise an inactivated Type I glutamine synthetase gene. The present disclosure is also directed to methods of producing biofuels from cellulosic biomass using the genetically engineered bacteria strains.

The invention discloses cloning and application of a poa pratensis glutamine synthetase gene PpGS1.1 promoter, relates to the field of gene engineering, and aims to fill the blank of research on a poa pratensis PpGS1.1 promoter sequence in existing resources and lay a foundation for improving poa pratensis nitrogen absorption and utilization capacity and improving the turf use quality of poa pratensis. According to the primer disclosed by the invention, the specific primer is SP1: 5 '-GCGGCCATGACAAGAATAAGAAG-3'; sP2: 5 '-GCGATGATCTTCTTCGGTGGTGTT-3', and SP3: 5 '- SP3: 5 '-AGATCGGTGAGGAGCGCCCATAA-3', and SP3: 5 '- And the SP4 is 5 '-TCGAACCGATGTACCGTATATGG-3', and the SP4 is 5 '- The nucleotide sequence of the promoter fragment of the PpGS1.1 gene is as shown in SEQ ID NO. 1.

The present invention provides a method for integrating multiple exogenous genes at specified sites in the host cell genome. The present invention also provides a method for modifying the host cell genome, the method comprising: a. integrating an exogenous gene at a designated site in the host cell genome; and / or b. knocking out fucose transfer in the host cell genome Enzyme 8 gene and / or glutamine synthase gene and / or α2,3-sialyltransferase gene 4 and / or α2,3-sialyltransferase gene 6. The present invention also provides host cells prepared according to the methods of the present invention. The present invention also provides the use of the host cell for producing the target protein. The cell genome of the CHO strain provided by the present invention integrates genes capable of improving protein expression at a specific point, which is of great significance for the development of novel antibodies and high-end imitation antibodies.

The present disclosure provides polynucleotides and related polypeptides related to urea uptake. The disclosure provides genomic sequences for urea transporter, urease and glutamine synthetase genes. Urea transporters, urease and glutamine synthetase are responsible for controlling nitrogen utilization efficiency in plants. Urea transporter, urease or glutamine synthetase sequences are provided for improving grain yield and plant growth. The disclosure further provides recombinant expression cassettes, host cells and transgenic plants.