391 results about "Agrobacterium" patented technology

The invention discloses a method for improving the inductivity and genetic transformation efficiency of hairy roots of toona ciliata. According to the method, agrobacterium rhizogenes carrying RUBY-TcGRF8-GIF2 chimeric protein are injected into stems and petioles of aseptic seedlings of the toona ciliata, and the TcGRF8-GIF2 chimeric gene is over-expressed, so that the induction efficiency and the genetic transformation efficiency of the toona ciliata hairy roots are remarkably improved, and the transformation period of the toona ciliata hairy roots is remarkably shortened. Specifically, after overexpression of the TcGRF8-GIF2 gene, the induction time of the hairy roots is shortened from 70 days of a RUBY-XbaI no-load plasmid control group to 25 days, the induction rate and conversion rate of the hairy roots mediated by agrobacterium tumefaciens are improved, the operation efficiency of genetic transformation of the toona ciliata is effectively improved, and the method has important significance for promoting genetic improvement of the toona ciliata.

The invention relates to the field of rice gene engineering, and discloses an application of a GSE3.1 protein and a coding gene thereof in regulating and controlling the grain length, the grain width and the grain weight of rice seeds. According to the invention, a GSE3.1 gene is edited based on a CRISPR / Cas9 technology, a japonica rice variety Zhonghua 11 (ZH11) is introduced by using an agrobacterium-mediated method, and a knockout mutant is obtained through screening. The homozygous knockout mutation of the GSE3.1 gene in the invention leads to reduction of the grain width and grain length of rice seeds and reduction of thousand grain weight. According to the invention, the constructed plant overexpression vector ProActin: GSE3.1 is expressed in wild type ZH11, so that compared with the wild type plant, the seed grain length and grain width of the transgenic plant are obviously increased, and the thousand grain weight is obviously increased. Therefore, the GSE3.1 gene and the encoding protein thereof can regulate and control the size and the weight of the rice seeds, and are of great significance to cultivation of high-yield rice varieties.

The invention relates to the field of plant molecular biology and forest genetic engineering, and particularly provides a PagRAP2.3 protein point mutant and application thereof in drought resistance of poplar. The mutant PagRAP2.3 MA is obtained by mutating methionine at the first site and cysteine at the second site of a wild type PagRAP2.3 protein into methionine and alanine. The invention further discloses a preparation method of the mutant PagRAP2.3. An expression vector containing the PagRAP2.3 MA gene is constructed, poplar 84K is transformed through an agrobacterium-mediated method, and a transgenic line with stable expression is obtained. Functional verification results show that overexpression of PagRAP2.3 MA can significantly increase the plant height of the poplar and promote plant growth, but the sensitivity to moisture is enhanced under drought stress. The mutant can be used for regulating and controlling the growth and development of forest trees and evaluating the drought resistance, and has a forestry breeding application prospect.

The invention discloses a plant immunomodulatory gene RDP as well as an expression vector and application thereof. The inventor finds a plant immunomodulatory factor gene RDP from potatoes, after a corresponding sequence is obtained through cloning, agrobacterium competent cells are transferred into nicotiana benthamiana, it is verified that the plant immunomodulatory factor gene RDP can be successfully expressed in other plants, and it is verified that the pathogenic effects of various phytophthora are good. The gene is proved to be capable of effectively improving the disease resistance of the nicotiana benthamiana after being expressed. Through objective evaluation of an agrobacterium tumefaciens-mediated transient expression system, it is found that the plant immunomodulatory factor RDP can induce plants to generate resistance, the gene can be directly introduced into the plants through a transgenic technology due to the powerful function of the gene, disease-resistant varieties are developed and cultivated, and the gene has great significance in prevention and treatment of plant diseases.

The invention discloses a sugarcane ScPP2C49 gene and application thereof, and relates to the technical field of plant biology, the nucleotide sequence of the ScPP2C49 gene is as shown in SEQ ID No.1, and the amino acid sequence of protein coded by the ScPP2C49 gene is as shown in SEQ ID No.2. The gene is induced to express by drought and abscisic acid, and the encoded protein of the gene is positioned in a cell nucleus. Functional studies show that after overexpression of the ScPP2C49 gene in the plant, the drought resistance of the plant is negatively regulated through various mechanisms of inhibiting an abscisic acid signal channel, hindering stomatal closure under drought stress, weakening active oxygen scavenging ability, reducing photosynthetic efficiency, inhibiting root growth and the like. The invention also provides a recombinant overexpression vector containing the gene and a method for obtaining a transgenic plant with reduced drought resistance by using the vector through an agrobacterium-mediated method. According to the invention, a new gene resource is provided for deep analysis of a plant drought-resistant molecular mechanism, and an important target gene is provided for cultivation of high-stress-resistance crop varieties through a reverse genetics means (such as gene knockout).

The invention belongs to the technical field of gene editing, and particularly relates to a construction method and application of a needle mushroom CRISPR / Cas9 gene editing vector. The CRISPR / Cas9 gene editing vector for flammulina velutipes is characterized in that the vector takes pFgnpt as a skeleton vector, and further comprises a Cas9 expression cassette and an sgRNA expression cassette; the nucleotide sequence of the Cas9 expression cassette is as shown in SEQ ID NO. 10. The nucleotide sequence of the sgRNA expression cassette is as shown in SEQ ID NO. 11. According to the present invention, with the agrobacterium-mediated transformation technology, the stable expression of Cas9 and sgRNA in the flammulina velutipes body is achieved, the efficient and stable CRISPR / Cas9 gene editing system is established, and the technical support is provided for the flammulina velutipes variety improvement, the functional gene research and the industrial upgrading.

The invention relates to the field of plant genetic engineering, and discloses an application of OsAAT2 protein and a coding gene thereof in regulating and controlling the salt tolerance of rice. The gene of the OsAAT2 protein is edited on the basis of a CRISPR / Cas9 technology, a rice ZH11 variety is introduced by utilizing an agrobacterium-mediated method, and a knockout mutant is obtained through screening; tests prove that the rice mutant with the OsAAT2 gene knocked out is rehydrated for 4 days after being subjected to stress treatment of 150 mM NaCl salt, and the survival rate is remarkably higher than that of a wild type ZH11 plant. And a simple and effective technical means is provided for rapidly creating a new salt-tolerant rice strain. Therefore, the OsAAT2 protein and the coding gene thereof disclosed by the invention can regulate and control the salt tolerance of the rice, and are of great significance to the cultivation of salt-tolerant transgenic rice.

The application provides a method for increasing the length of a main panicle of rice, the number of secondary branches of the main panicle, the number of grains on the main panicle of rice and / or improving the yield of rice, which comprises overexpressing a cystathionine beta-lyase gene OsCBL in rice by using a recombinant vector containing a CDS coding sequence of the cystathionine beta-lyase gene OsCBL as shown in SEQ ID NO: 2 to obtain a transgenic plant. The transgenic plant obtained by the method has an increased number of grains per panicle and an increased yield per plant compared with normal plants. The method can be used for treating rice plants to improve the yield of rice and provides a new gene resource and a new idea for breeding rice with increased yield.

The invention discloses an oilseed rape gene BnaKNAT1s and an application thereof in creation of droop-type silique oilseed rape, and belongs to the technical field of plant genetic engineering. The gene comprises two copies of BnaA03.KNAT1 and BnaC03.KNAT1, and the CDS (Coding Sequence) sequences of the gene are respectively shown as SEQ ID NO: 1 and SEQ ID NO: 2. Targeted sgRNA is designed according to homologous regions of first exons of the two genes, nucleotide sequences of the sgRNA are shown as SEQ ID NO: 3 and SEQ ID NO: 4 respectively, CRISPR / Cas9 editing vectors are constructed, agrobacterium tumefaciens is transferred, and then rape is infected, so that a transgenic plant is obtained. And screening a homozygous mutant with two genes subjected to frameshift mutation and without a transgenic element, hybridizing the homozygous mutant with a good variety, and carrying out multi-round backcross and selfing to obtain a new germplasm of the sagittaria silique rape. According to the germplasm, the canopy structure is optimized, the light energy utilization rate and ventilation performance are improved, high-density planting is adapted, and gene resources and core materials are provided for high-yield breeding of oilseed rape.

The invention relates to a cultivation method of an overexpressed EVM0003075 gene isatis tinctoria plant, and belongs to the technical field of plant cultivation. The method comprises the following steps: firstly, transforming an agrobacterium strain through a constructed agrobacterium transformation vector for expressing the isatis tinctoria EVM0003075 gene, then transforming an explant of the isatis tinctoria by using a screened recombinant agrobacterium strain, regenerating a seedling of the isatis tinctoria, and screening and culturing the seedling of the isatis tinctoria with stable overexpression of the EVM0003075 gene, so as to obtain an isatis tinctoria plant with increased isatin content in the isatis tinctoria. The expression level of the EVM0003075 gene of the transgenic isatis tinctoria is improved to 5-9 times, the content of the isatis tinctoria is remarkably improved, and the character that the accumulation period of the isatis tinctoria is short is shown. By adopting the method disclosed by the invention, the content of the isatin in the isatis tinctoria can be increased to 10-66 times. The specific information content of the patent of the method for improving the content of the isatin in the isatis tinctoria through overexpression of the EVM0003075 gene is shown in the specification.

The invention relates to the technical field of preparation of L-tyrosine, and discloses a method for enhancing generation of L-tyrosine in nicotiana benthamiana and application. In order to relieve product negative feedback inhibition of a key enzyme in a production path of L-tyrosine in tobacco and to improve the yield of L-tyrosine in tobacco, DAHPS enzyme derived from microorganisms is subjected to L175Q site mutation and then is connected with a signal peptide AtRs1A; shikimic acid mutase / prebenzoate dehydrogenase TyrAfbr and tyrosine aminotransferase TyrB which are derived from microorganisms are introduced into the tobacco chassis after being connected with the signal peptide Rs1A, the expression has orthogonality, and transient expression is performed in the Bensi tobacco plant chassis by utilizing agrobacterium infection, so that the synthesis capability of L-tyrosine in tobacco chassis cells can be improved; a large amount of L-tyrosine can be directly generated in the tobacco chassis plastid.

The application provides a plasmid containing modified satellite RNA and a construction method and application, and belongs to the field of plant virology and molecular biology. Each copy number of the modified satellite RNA is composed of a modified satRNA TA-Tb fragment and an HDV ribozyme fragment, and the copy number of the modified satellite RNA in the plasmid is 10. The nucleotide sequence of the modified satellite RNA is shown in SEQ No. 3. After the plasmid of the application is inoculated by agrobacterium injection, the symptoms caused by cucumber mosaic virus on different strains and the same strain (CMV Fny ) on different hosts can be significantly reduced. An effective control measure for field control of cucumber mosaic virus is provided.

This invention discloses PagUBC10a The application of genes in regulating the growth and development of poplar trees belongs to the field of agricultural bio-genetic engineering technology. PagUBC10a The application of genes in regulating the growth and development of poplar trees, the PagUBC10a The nucleotide sequence of the gene is shown in SEQ ID NO:1, and is utilized using the... PagUBC10a Genetically modified poplar trees are cultivated using the following methods: cloning poplar trees. PagUBC10a Genes; the aforementioned PagUBC10a The gene is linked to a vector to obtain an overexpression vector; the overexpression vector is transformed into Agrobacterium to obtain Agrobacterium bacterial suspension; the Agrobacterium bacterial suspension is used to inoculate poplar leaves to obtain... PagUBC10a Gene overexpression in poplar. This invention provides a new option for regulating the growth and development of poplar trees and offers a potential target for molecular breeding improvement of poplar traits.

The invention discloses a PVX expression vector pWX for efficiently expressing foreign protein and application of the PVX expression vector pWX. In the construction process of optimizing potato X virus infectious clone, after a single base mutation NruI-gene sequence (CCGCGA) is inserted into an upstream regulation region of a PVX coat protein subgene promoter, a novel PVX expression vector capable of promoting foreign protein expression is obtained, and the novel PVX expression vector is named as pWX. Experiments prove that compared with a control vector, the accumulation amount of GFP (Green Fluorescent Protein) generated by the pWX vector through transient expression of agrobacterium in tobacco is obviously increased. The invention further applies a pWX vector to successfully express an important target protein HPV16L1 researched and developed by an HPV vaccine, and the expression quantity of the important target protein HPV16L1 in tobacco is 5.25 micrograms per gram of fresh heavy leaves. The invention provides a novel vector based on a PVX genome for high-efficiency expression of a plant source target protein, and the vector has a good application prospect in preparation of plant source vaccines.

The invention relates to the technical field of biology, in particular to a method for regulating and controlling plant sodium ion homeostasis and salt tolerance by a light-operated pump, which comprises the following steps: constructing systems of rhodopsin light-sensitive pumps KR2-T1.0, KR2-T2.0, KR2-T3.0 and KR2-T4.0 with different cell membrane localization and membrane loading efficiencies through gene optimization, connecting the systems with a retinaldehyde production gene Ret, then inserting into a plant expression vector pCAMBIA3300, and carrying out photosynthesis to obtain the plant sodium ion homeostasis and salt tolerance control plant sodium ion homeostasis and salt tolerance control plant sodium ion homeostasis and salt tolerance control plant sodium ion homeostasis and salt tolerance control plant sodium ion homeostasis and salt tolerance control plant sodium ion homeostasis. Transient expression of nicotiana benthamiana and stable expression of rape are achieved through agrobacterium tumefaciens mediation, the pump is activated through irradiation of green light of 530 nm + / -5 nm, active excretion in plant cells is driven, and accurate, dynamic and reversible regulation and control of the sodium ion steady state are achieved. According to the method, the technical bottlenecks that KR2 is poor in plant cell membrane localization and low in membrane loading efficiency are broken through, gradient regulation and control of efflux efficiency are achieved, the method is high in response speed, light energy serves as power, cell ATP is not depended on, exogenous chemical intervention is avoided, and the plant salt tolerance can be remarkably improved.

The invention discloses application of apple MdMYB41L in regulation and control of seed size, and belongs to the technical field of plant genetic engineering. According to the invention, an MdMYB41L gene is cloned from apple GL-3, the CDS nucleotide sequence of the MdMYB41L gene is shown as SEQ ID NO: 1, and a plant overexpression vector is constructed. A transgenic plant is obtained by heterologous transformation into arabidopsis thaliana through an agrobacterium tumefaciens-mediated method. Compared with wild arabidopsis thaliana, the weight, the length, the width and the surface area of the transgenic plant seeds are obviously improved. In addition, the cotyledon area of a transgenic plant is obviously higher than that of a wild type. Therefore, the MYB transcription factor MdMYB41L provides a new thought in the aspects of improving the plant seed size and molecular breeding.

The invention belongs to the technical field of plant genetic engineering, and relates to a GhAL5 gene and application thereof in cultivation of heat-resistant cotton and rice germplasm. Researches find that expression of the AL5 gene is up-regulated by using an agrobacterium tumefaciens-mediated genetic transformation technology, so that heat resistance of cotton and rice can be enhanced, plant anther fertility can be enhanced, and leaf damage degree can be reduced. The invention provides a new approach and method for improving the heat resistance of cotton and rice, and has wide agricultural application prospect and market value.

The invention discloses application of agrobacterium tumefaciens C53 in converting cow dung water into efficient liquid microbial fertilizer, and relates to the field of agricultural environment microbial technology and organic waste resource utilization, agrobacterium tumefaciens C53 bacterial liquid is inoculated into cow dung water, fermentation is carried out under the aerobic condition, and the liquid microbial fertilizer is obtained. The inoculation volume of the C53 bacterial liquid accounts for 20% of the total volume of the cow dung water. According to the strain C53, high-load cow dung water is transformed into a liquid fertilizer with better nutrient form and remarkable growth promoting effect, after the efficient liquid microbial fertilizer is applied to corn, the leaf weight and the root weight of the corn can be increased by about 48.9% and 58.8% respectively compared with blank control, and a feasible microbial technical scheme is provided for green and high-value resource utilization of breeding dung.

The application belongs to the field of new application of functional proteins and mutants thereof, and discloses application of sugarcane ScCBS protein and key domain deletion mutants thereof. The nucleotide sequence of the sugarcane ScCBS protein is shown as SEQ ID NO:1, and the amino acid sequence is shown as SEQ ID NO:2. The key domain deletion mutants of the ScCBS protein are ScCBS△motif1, ScCBS△motif3, ScCBS△motif4 and ScCBS△motif9, which respectively lack 28, 50, 49 and 29 amino acids. The application proves that the ScCBS protein can interact with SCSMV P1 and inhibit the RNA silencing suppressor activity by means of yeast two-hybrid, two-molecular fluorescence complementary experiment and agrobacterium transient expression system; the four mutants all lose the interaction and inhibition activity. Therefore, the ScCBS protein and the key domain deletion mutants thereof can be used for cultivating excellent SCSMV-resistant materials, and have wide application prospects in sugarcane disease-resistant molecular breeding.

The invention relates to a non-tissue-culture-dependent hevea brasiliensis high-altitude layering genetic transformation method, and relates to the technical field of plant genetic engineering. The method comprises the following steps: S1, providing an agrobacterium rhizogenes K599 bacterial solution containing a target gene, and loading the target gene in a pCAMBIA2301 vector; s2, preparing an infection material (infection liquid or lawn) by using the agrobacterium rhizogenes K599; s3, carrying out girdling treatment on stems of the rubber tree seedlings, and carrying out pretreatment on girdling wounds; s4, the pretreated girdling wound is infected with the infection material; s5, covering and mounting the girdling wound with a wet culture medium so as to keep the wound wet and incubate the strain; wherein the target gene is a GUS gene, an RUBY gene or a JcFT gene. The method can avoid a series of complicated operations such as tissue culture, and is simple, convenient, rapid, low in cost, not limited by seasons, easy to operate and independent of genotypes; particularly, by means of the method, the high positive rate can be obtained.

The present invention relates to a method for preparing a MAGEA8-Fc transgenic plant. Through a platform using a tobacco plant, a recombinant protein MAGEA8-Fc, in which a human IgG Fc fragment is fused to a MAGEA8 protein, can be stably expressed. The method of the present invention enables high-level expression efficiency and stable protein production by introducing a target gene into a plant genome using an Agrobacterium binary vector. By directly isolating and purifying the recombinant protein from the transgenic tobacco plant, large-scale production can be easily achieved. Compared to conventional microbial or animal cell-based expression systems, the method provides economic efficiency and scalability, and thus can be usefully applied as a platform for producing fusion proteins.

The invention discloses a Chinese cabbage clubroot-resistant gene BrRLP31 and application of the Chinese cabbage clubroot-resistant gene BrRLP31. The nucleotide sequence of the Chinese cabbage clubroot-resistant gene BrRLP31 is as shown in SEQ ID No. 1. Functional verification results show that after the expression of the endogenous BrRLP31 of the Chinese cabbage is inhibited by utilizing a VIGS technology, the resistance of the plant to clubroot is obviously reduced, and the disease index and the root swelling severity of the plant are obviously higher than those of a control material. On the other hand, the transgenic flowering cabbage plant over-expressed by the BrRLP31 is constructed by an agrobacterium-mediated method, and disease resistance identification proves that the heterologous over-expression of the BrRLP31 obviously improves the resistance of the plant to plasmodiophora brassicae. The results show that the BrRLP31 gene plays a key positive regulation role in the clubroot resistance process of the Chinese cabbage, the disease resistance of cruciferae crops and other horticultural plants is improved by utilizing the gene and the homologous sequence thereof, and the gene and the homologous sequence thereof have important application value.

The application belongs to the field of plant genetic engineering, and particularly relates to application of a rice-derived OsDREB1 gene in enhancing the low-phosphorus stress tolerance of soybeans. A recombinant expression vector of the OsDREB1 gene is constructed, and the soybean plants are transformed by using an agrobacterium-mediated method to obtain a transgenic soybean line with stable heredity. It is found through experiments that overexpression of the OsDREB1 gene in soybeans can enhance the low-phosphorus tolerance of the soybeans, and under the same conditions, the number of pods per plant and the weight of 100 seeds of the transgenic plants are higher than those of a wild control group. The growth performance of soybeans in low-phosphorus soil is improved by using genetic engineering technology, and a new strategy for reducing the dependence of agriculture on phosphorus fertilizer is provided.

The application belongs to the technical field of biology, and discloses application of CmPHL1 transcription factor in inhibition of citrus leaf oil cell formation and regulation of volatile substances. In view of the problem of insufficient research on existing citrus oil cell formation and essential oil synthesis, a pGBI-CmPHL1 overexpression vector is constructed, and positive strains are obtained by means of Agrobacterium-mediated transformation of citrus. Experiments prove that overexpression of CmPHL1 can significantly reduce the number of oil cells and the content of monoterpenes, sesquiterpenes and other volatile substances. CmPHL1 targets the molecular regulation network of oil cell development, accurately regulates and is heritable, provides a new gene target for citrus quality improvement and breeding, is in line with the direction of green agriculture, and has important economic and ecological benefits.

The invention discloses application of an OsRPP13 gene in regulation and control of heat resistance and photosynthetic performance of rice. According to the invention, a CRISPR-Cas9 carrier is constructed, and rice OsRPP13 gene knockout mutant strains (osrpp13-2, osrpp13-18 and osrpp13-20) are obtained by using an agrobacterium tumefaciens-mediated transformation method. The inventor discloses the function of the OsRPP13 gene in the aspect of regulating and controlling the temperature tolerance and the photosynthetic performance of a rice plant by improving a culture temperature simulation experiment. Through the research, a gene resource is provided for high-temperature-resistant and high-photosynthetic-efficiency molecular breeding of rice. The rice can survive and grow in a temperature range exceeding the original adaptive temperature range, the photosynthetic efficiency is improved, and a gene resource is provided for high-photosynthetic-efficiency molecular breeding and genetic improvement of the rice.

The invention belongs to the technical field of plant genetic engineering, and particularly relates to a transcription factor BnC05.PLATZ01 gene for regulating and controlling sclerotinia rot resistance of rape and application of the transcription factor BnC05.PLATZ01 gene. A transcription factor BnC05.PLATZ01 gene capable of enhancing sclerotiniose resistance of rape is screened, and the nucleotide sequence of the transcription factor BnC05.PLATZ01 gene is as shown in SEQ ID NO: 1; the sequence of the protein coded by the gene is as shown in SEQ ID NO: 2. According to the method, a pGWB418-BnC05. PLATZ01 overexpression strain is obtained by virtue of agrobacterium tumefaciens mediated transformation. Sclerotinia sclerotiorum inoculation identification on a transgenic material finds that the sclerotinia rot resistance of a BnC05. PLATZ01 overexpression strain is enhanced, which indicates that the BnC05. PLATZ01 is a positive regulation factor of the sclerotinia rot resistance of the rape and has a positive regulation effect in the sclerotinia rot resistance of the rape, and the sclerotinia rot resistance of the rape can be remarkably improved by overexpressing the gene.

The invention discloses a method for detecting subcellular localization of a functional gene by using a tea tree leaf, a target gene is expressed in the tea tree leaf through an agrobacterium transformation method, the method has the advantages of short period, rapidness, high conversion rate, low cost, easiness in operation and the like, and the localization condition of the target gene in the tea tree leaf can be more intuitively and accurately observed; and important technical support is provided for tea tree gene function research. Compared with mode plants such as tobacco and arabidopsis thaliana, the novel subcellular localization experimental material has the advantages that the tea tree leaves are used for observing subcellular localization, and a more intuitive and more accurate mode is provided for gene function research in tea trees. In addition, the method is short in conversion period, fluorescence signals can be observed 36-48 hours after conversion, and the method is fast and efficient.

The invention relates to the technical field of gene editing, and particularly provides a method for creating fluazifop-p-butyl herbicide resistant rice through gene editing of an OsACCase gene promoter region. The method comprises the following steps: constructing a crRNA plasmid library covering an OsACCase gene promoter region on the basis of a CRISPR / LbCas12a system, performing agrobacterium-mediated transformation on rice callus, and performing resistance screening, fluazifop-p-butyl herbicide screening and molecular identification to obtain a fluazifop-p-butyl resistant rice strain. The strain is subjected to-4bp / -5bp double allelic deletion mutation in an upstream 1865-site region of an initiation codon ATG of the OsACCase gene, and a homozygous mutant strain and a double allelic deletion mutant strain can normally grow after being sprayed with 6 times of field recommended dose of fluazifop-p-butyl. The herbicide resistance is realized by editing the gene promoter region, the growth defect of coding region mutation is avoided, the breeding technology is enriched, and weed prevention and control are assisted.