21 results about "Acetosyringone" patented technology

The invention belongs to the technical field of genetic engineering of edible mushrooms, and particularly relates to a construction method of an efficient and stable genetic transformation system of flammulina velutipes. The construction method comprises the following steps: culturing in an MM culture medium by taking a flammulina velutipes strain 0747 as a receptor; the pFgnpt vector is converted into agrobacterium AGL1 (AGL1); inoculating hyphae into an IM liquid culture medium containing acetosyringone and MES, mixing with agrobacterium tumefaciens, culturing, and transferring the hyphae to an IM solid flat plate paved with filter paper; hyphae are transferred to a PDA resistance plate containing G418 and cephalosporin for resistance screening, and an efficient and stable flammulina velutipes genetic transformation system is obtained. The new selection marker G418 is developed and applied, limitation of a traditional selection marker is broken, more choices are provided for flammulina velutipes genetic transformation, a new thought is provided for genetic transformation research of other edible mushrooms, and the application field of an agrobacterium tumefaciens-mediated transformation technology is expanded.

This invention provides a method for transient transformation of oil palm zygote embryos based on Agrobacterium-mediated transformation, which employs the Agrobacterium-mediated approach and is based on the optimal Agrobacterium concentration (OD) for stable transformation of oil palm. 600 The optimal Agrobacterium infection time for zygotic embryos was determined by measuring the concentration of acetylsuccinone and co-culture time. The transient GUS expression staining rate was compared between zygotic embryos of different oil palm varieties and at different growth stages. The method described in this invention optimizes infection conditions based on stable transformation parameters, clarifies the optimal explant materials and treatment process, and effectively improves transient transformation efficiency and experimental repeatability. This study establishes for the first time an Agrobacterium-mediated transient transformation system for oil palm zygotic embryos and obtains zygotic embryos most suitable for transient transformation, achieving efficient transient expression of exogenous genes in oil palm zygotic embryos. This provides a rapid and low-cost technical means for studying oil palm gene function.

The application provides a method for Agrobacterium-mediated genetic transformation of oil palm, and the method uses embryogenic callus of oil palm as explants, adopts Agrobacterium-mediated mode, and selects the most suitable Agrobacterium liquid concentration, immersion time, co-culture time, acetosyringone concentration and antibiotic concentration in the transformation process. An expression cassette containing an EPSPS mutant gene is used as a screening marker, and glyphosate is used as a screening agent to select glyphosate-resistant callus. The screening efficiency of the application is higher than that of the traditional hpt / hygromycin screening system. Meanwhile, the effectiveness of the EPSPS / glyphosate screening system in the genetic transformation of oil palm is proved, which marks the first application of the system in the genetic transformation of oil palm and provides a valuable tool for promoting the research of the important economic crop.

The invention belongs to the technical field of edible mushroom genetic engineering, and relates to a method for constructing a mushroom genetic transformation system by using a G418 resistance selection marker, which comprises the following steps: taking a mushroom strain Shenxiang 215 cultured in a PDA (potato dextrose agar) culture medium as a receptor to obtain hypha blocks; the method comprises the following steps: transferring a pLgnpt plasmid into agrobacterium tumefaciens GV3101; agrobacterium tumefaciens is inoculated into an IM liquid culture medium containing acetosyringone and MES, hypha blocks are subjected to ultrasonic treatment and co-incubation with the hypha blocks, then the hypha blocks are transferred to an IM solid culture medium laid with filter paper, and dark culture is carried out; and transferring the hyphae subjected to dark culture to a PDA resistant plate culture medium containing G418 and Cef for resistance screening to obtain the shiitake transformant. The method not only verifies the applicability of the novel selection marker in flammulina velutipes, but also constructs a multi-marker technology platform parallel to the existing hygromycin system, and lays a foundation for subsequent complex operations such as multi-gene superposition editing, gene loop construction and the like.

The invention discloses a fraxinus mandshurica somatic embryo genetic transformation method, which comprises the following steps: carrying out genetic transformation on fraxinus mandshurica embryonic cells through ultrasonic and vacuum assisted agrobacterium mediation, co-culturing for 3 days, removing bacteria, carrying out resistance screening, irradiating by using an LED red light source, illuminating for 16 hours every day, darkness for 8 hours every day, and culturing for 4-8 weeks at the temperature of 25 DEG C to obtain the somatic embryo. LED red and blue light combination (16 red and 9 blue) is used for irradiation, illumination is carried out for 16 h every day, darkness is carried out for 8 h every day, the temperature is 25 DEG C, resistant somatic embryo regeneration seedlings are obtained after 4-6 weeks, and GUS detection proves that genetic transformation succeeds. According to the method, ultrasonic and vacuum assistance is used in agrobacterium mediation, so that the addition of acetosyringone medicine is avoided, and the cost is saved. Red and blue light irradiation is utilized in somatic embryo induction and regeneration, light signals are accurately controlled, and compared with full-spectrum irradiation of white light, energy is saved, and cost is low. The invention provides a new way for genetic transformation of fraxinus mandshurica.

The invention discloses grape pomace pretreatment liquid and a method for preparing microcrystalline cellulose from grape pomace. The grape pomace pretreatment liquid is prepared from laccase, polygalacturonase, acetosyringone, sodium dihydrogen phosphate, disodium hydrogen phosphate, dithiothreitol, fatty alcohol-polyoxyethylene ether, sorbitol and the balance of water. The method for preparing the microcrystalline cellulose from the grape pomace comprises the following steps: S1, cleaning the grape pomace with normal hexane, then cleaning with water, drying and crushing; s2, carrying out mixed reaction with the pretreatment liquid, and then separating out pretreatment residues; s3, adding a sodium carbonate solution for premixing, then adding a tetramethylammonium hydroxide solution and a sodium borohydride solution, and filtering after reaction to obtain filter residues; s4, adding a citric acid solution, adding carboxymethyl chitosan, adsorbing, and filtering to obtain cellulose residues; and S5, hydrolyzing and depolymerizing by using dilute acid, neutralizing, separating and washing an acidolysis product, and drying to obtain the microcrystalline cellulose. According to the method, impurity components which are difficult to treat in the grape pomace can be overcome, impurity removal and purification are completed, cellulose is reserved to the maximum extent, loss is reduced, and the yield is remarkably increased while high-purity microcrystalline cellulose is obtained.

The invention relates to a construction method of an Agrobacterium rhizogenes mediated acacia melanoxylon genetic transformation system, which comprises the following steps: by taking an acacia melanoxylon SR17 sterile single plant as an explant, selecting an Agrobacterium rhizogenes strain Ar1193, infecting for 45 minutes in a heavy suspension of which the bacterial liquid concentration OD600 is equal to 0.9 and the acetosyringone concentration is 100 [mu] mol.L <-1 >, co-culturing for 3 days, and then transferring into a bacteriostatic culture medium to induce hairy roots; and taking the transgenic hairy roots obtained by induction as explants, and inoculating the explants into a WPM culture medium containing 0.05 mg.L <-1 > TDZ and 200 mg.L <-1 > casein hydrolysate to induce regeneration of adventitious buds so as to obtain complete transgenic plants. The screening concentrations of the four antibiotics, namely kanamycin, hygromycin, glyphosate and glufosinate-ammonium, are determined; a stable and efficient acacia melanoxylon genetic transformation system is established, the hairy root induction rate reaches 63.17%, the transgenic positive rate reaches 15.42%, and the adventitious bud regeneration rate reaches 52%.

The application relates to the field of plant genetic engineering, and specifically discloses a high-efficiency gene transformation system based on the stem tip meristem of a yellowing seedling of Zhongmian 113. The core components of the system include L1 medium suitable for Zhongmian 113, Agrobacterium GV3101 carrying a target gene, a special resuspension solution containing acetosyringone and Silwet-L77, and special primers for overexpression and gene editing; and the preparation method is as follows: seed disinfection and germination are performed to obtain a yellowing seedling, the stem tip is cut in an X shape to expose the meristem, Agrobacterium infection is performed, and then dark culture and light culture are performed, and after transplanting and cultivation to the true leaf stage, positive plants are screened and identified through PCR; the system can be used for gene transformation of Zhongmian 113 and improvement of high yield, high quality, disease resistance and the like, and has the advantages of strong adaptability, high transformation efficiency and short cycle; in addition, the preparation method of the application does not need callus regeneration, is simple and convenient to operate, has a standardized process, is good in repeatability and stability, and can quickly realize transgenic seed acquisition.

The application belongs to the field of biology and modern agricultural technology, and discloses a rooting aid invasion agent, and a formula thereof is as follows: B5 culture medium 3.0-3.5 g / L, sucrose 28-32 g / L, MES 3.8-4.0 g / L, acetyl-syringone 38-42 mg / L, 6-BA 1.5-1.8 mg / L, gibberellin 0.02-0.03 mg / L, and surfactant Silwet L-77 100 ml / L, and pH is 5.4. The application also discloses a soybean hairy root induction and transformation method mediated by Agrobacterium rhizogenes by using the rooting aid invasion agent, and the method comprises the following steps: disinfection and germination of soybean seeds; resuspension of Agrobacterium rhizogenes with a target expression plasmid in the rooting aid invasion agent to obtain an Agrobacterium rhizogenes aid invasion liquid; infection; co-culture; and screening of positive roots.

The invention discloses a tissue-culture-free agrobacterium rhizogenes-mediated genetic transformation system method for crape myrtle hairy roots, and relates to the technical field of molecular biology.The method comprises the steps that an overexpression vector carrying an RUBY visual tag is constructed, agrobacterium rhizogenes K599 is introduced, and a positive engineering strain is obtained through resistance screening and molecular verification; propagating the positive engineering strain, centrifugally collecting thalli, resuspending by using a buffer solution containing acetosyringone, adjusting the concentration of a bacterial solution, and standing and activating to form a bacterial suspension for infection; the method comprises the following steps: selecting current-year semi-lignified branches of crape myrtle to prepare cutting slips, retaining two pairs of bud points, and performing incision treatment; the cutting slips are completely immersed in the bacterial suspension for infection, and agrobacterium tumefaciens-mediated genetic transformation is completed in combination with vacuum-assisted treatment; taking out the cutting slips, temporarily dipping the cutting slips in a rooting aqueous solution, and then cutting the cutting slips in a sterile substrate formed by mixing vermiculite and perlite; and culturing for 24 hours under a dark condition, and continuously culturing for 30 days in a high-temperature and high-humidity environment.

The invention belongs to the technical field of agricultural biology, and discloses a non-tissue culture dependent transformation method for promoting effective transformation of potatoes by using growth factors. The invention discloses a reagent composition. The reagent composition comprises an infection solution and a bud inducing screening solution, wherein the infection liquid is prepared from acetosyringone, dithiothreitol and plant hormones; the bud inducing screening liquid comprises plant hormones, glufosinate-methyl and regeneration factors. The invention provides a reagent composition. The reagent composition can be used for non-tissue culture dependent genetic transformation of potatoes. By using the reagent composition, the non-tissue culture dependent genetic transformation step of the potatoes can be simplified, the conditions of chimera and false positive of transformants are reduced, and the transformation efficiency is remarkably improved.

The invention provides an asparagus microspore induction culture medium and a culture method, and relates to the technical field of crop cultivation. The asparagus microspore induction culture medium comprises the following components, 50 to 1500 mg / L of KNO3, 300 to 340 mg / L of NH4H2PO4, 155 to 195 mg / L of CaCl2. 2H2O, 18 to 224 mg / L of MgSO4. 7H2O, 60 to 700 mg / L of K2SO4, 7.3 to 9.5 mg / L of MnSO4. 4H2O, 2.0 to 4.0 mg / L of ZnSO47H2O, 2.0 to 4.0 mg / L of H3BO3, 0.5 to 1.0 mg / L of KI, 2.0 to 2.4 mg / L of acetylcholine, 20 to 30 mg / L of spermidine, 5.0 to 6.0 mg / L of proline, 1.5 to 2.5 mg / L of glycine, 20 to 240 mg / L of glutathione, 80 to 100 mg / L of inositol, 2.4 to 3.6 mg / L of acetosyringone-220, 2 to 4 mg / L of The addition of acetylcholine, spermidine, proline, glutathione, acetosyringone, epicatechin, hydrocortisone and other additives can promote the induction of the microspore callus.

The present invention belongs to the technical field of plant genetic engineering, and particularly relates to a genetic transformation method for perennial Elymus nutans. The present invention provides a genetic transformation method for Elymus nutans. Using mature embryos as explants for inducing callus formation has the characteristics of convenient material collection and simple and easy operation, avoiding the supply problem of explant material collection. At the same time, the consistent physiological state of mature embryos results in good repeatability of experiments. Additionally, the infection solution contains acetosyringone, and acetosyringone (AS, Acetosyringone) can promote the growth of Agrobacterium and enhance its Agrobacterium activity, thereby improving the transformation efficiency. The bactericidal treatment effectively inhibits the browning phenomenon of callus, enabling the callus to resume normal growth. Transgenic Elymus nutans plants are obtained using the technical solution of the present invention, which can greatly promote the research of researchers on the gene functions of Elymus nutans.

The invention relates to the technical field of plant genetic engineering and agricultural biology, and discloses a method for improving drought resistance of plants based on agrobacterium tumefaciens K599 and application of the method. The method comprises the following steps: cutting off radicles of germinating plant seeds, immersing wound sections into an infection bacterium solution containing a K599 strain and acetosyringone, transplanting, and culturing to obtain the transgenic hairy roots. Through optimization, the peanut transformation efficiency reaches 78.33%, and the method is suitable for various plants such as pelargonium odoratissimum and bryophyllum pinnatum. Taking the peanut AhCYP76b6 gene as an example, the leaf withering degree of an overexpressed plant subjected to drought treatment is obviously lower than that of a control group, and the drought resistance of the overexpressed plant is proved to be obviously enhanced. The method is easy and convenient to operate, high in transformation efficiency and high in cross-species applicability, and an effective tool is provided for plant drought-resistant gene function research and germplasm innovation.

The invention discloses an establishment method of a grape genetic transformation system, and relates to the technical field of plant genetic engineering.The method comprises the steps that grape seeds are placed in distilled water at the temperature of 55 DEG C to be soaked for 15 min and then stand for 24 h at the normal temperature; placing the soaked grape seeds on gauze in a first culture dish, and performing dark culture for 10-15 days until the grape seeds crack and expose embryos; the method comprises the following steps: preparing agrobacterium, activating the agrobacterium, obtaining thalli in the activated agrobacterium, re-suspending the thalli until OD600 is equal to 0.9 through a 1 / 2 MS liquid culture medium, and adding 100 mM of acetosyringone into the 1 / 2 MS liquid culture medium to obtain an agrobacterium infection solution; and transferring the cracked and embryo-exposed grape seeds into a second culture dish containing 20ml of agrobacterium infection liquid for shallow culture for 2d, and screening to obtain agrobacterium transformed grape plants. The method is easy to operate, the tedious processes of seed cutting and plant tissue culture are avoided, damage to embryos caused by seed coat cutting is also avoided, the consistency of genetic transformation materials is achieved, and meanwhile the transformation efficiency is improved.

The application discloses a kind of Agrobacterium-mediated transgenic methods based on Anthracene callus.The method includes callus induction and proliferation, Agrobacterium infection, co-culture, and resistant callus screening.The application discloses callus induction medium, subculture medium, co-culture medium and screening medium and other formulations, Agrobacterium infection time and bacterial liquid concentration, and acetyl-syringone concentration, first establish Anthracene genetic transformation system with callus as receptor, which is conducive to molecular directed breeding, accelerates excellent foreign gene transfer to Anthracene, improves Anthracene variety, provides new ideas for Anthracene industrialization development, and has important application prospect.

An agrobacterium-mediated transgenic method on the basis of Hippeastrum calli, which method comprises the steps of the induction and proliferation of calli, agrobacterium infection, co-culture and the screening of resistant calli. Provided are formulas such as a callus induction medium, a subculture medium, a co-culture medium, and a screening medium, the time and bacterial liquid concentration of Agrobacterium infection, and the concentration of acetosyringone. A Hippeastrum genetic transformation system using a callus as a receptor is established for the first time, which facilitates molecular directed breeding, accelerates the transfer of excellent exogenous genes into Hippeastrum, and improves the variety of Hippeastrum. The method provides a new idea for the industrial development of Hippeastrum and has important application prospects.

The invention relates to the technical field of plant genetic transformation, and particularly discloses an agrobacterium rhizogenes-mediated efficient genetic transformation method for citrus. The composition is mainly an infection solution and comprises an MS liquid culture medium, agrobacterium rhizogenes K599 containing target genes and acetosyringone. The preparation method comprises the following steps: culturing citrus seedlings, preparing an agrobacterium infection solution containing acetosyringone, trimming the roots of the seedlings, making wounds, wrapping the wounds with cotton slivers soaked with the infection solution, inducing hairy roots through dark culture and light culture, and finally extracting hairy root nucleic acid to identify transgenic plants. The method can be used for citrus genetic improvement, functional gene verification and metabolic engineering research, and a transgenic hairy root material can be efficiently obtained; in addition, the preparation method is easy and convenient to operate, acceptor materials are good in adaptability, the agrobacterium infection stability can be guaranteed, the problems that traditional transformation is low in efficiency and long in period are solved, and the quality of transgenic materials can be accurately confirmed in the identification link.

The invention relates to a method for constructing a gene editing system of hybrid larch, aiming at solving the problem of difficulty in gene editing of the hybrid larch and optimizing the gene editing system of the hybrid larch. The method comprises the following steps: 1, taking a hybrid larch callus growing for about 10 days as a gene editing transformation material; 2, preparing an infection solution, and resuspending the target gene-containing agrobacterium by using the infection solution to obtain an infection solution containing the agrobacterium; 3, placing the hybrid larch callus in an infection solution containing agrobacterium tumefaciens, and transferring the hybrid larch callus into a shaking table for infection; 4, filtering out the callus of the hybrid larch from the infection solution, and laying the callus on a culture medium containing acetosyringone; and 5, carrying out bacterium removal treatment on the callus which is placed for about 24 hours, and then laying the callus on a culture medium containing cephalosporin for subsequent screening. The CRISPR / Cas9 editing system of the hybrid larch callus, which is constructed by the method, is used for transforming larch genes, so that the CRISPR / Cas9 edited transgenic larch callus can be obtained. The method is used for constructing the CRISPR / Cas9 editing system of the hybrid larch.

The invention provides a construction method of an agrobacterium tumefaciens-mediated wheat genetic transformation system and a culture medium of the agrobacterium tumefaciens-mediated wheat genetic transformation system, and belongs to the technical field of genetic transformation. The method comprises the following steps: firstly, taking a Fielder immature embryo as an explant, optimizing the influence of the embryo age, the concentration and infection time of acetosyringone, the concentration of glufosinate-ammonium and the concentrations of ABA and KT on callus induction, differentiation and transformation efficiency, and carrying out applicability evaluation on Yangmai 20, Zheng mai 7698 and Kinong 199 by utilizing an optimized system. Under an optimized system, the transformation efficiency of Fielder reaches 14.83% and is improved by 81.7% compared with that before optimization, and the transformation efficiency of materials such as Yangmai 20, Zheng mai 7698 and Konnong 199 which are difficult to genetically transform reaches 7.32%-9.03%. The system is optimized, the wheat genetic transformation efficiency is improved, the method has good applicability to wheat varieties with different genetic backgrounds, and support is provided for wheat functional gene analysis and molecular breeding.

The invention discloses a method for establishing a transgenic system of bilberry mediated by Agrobacterium tumefaciens. The method comprises: taking bilberry stems and leaves as explants and inducing them to grow a thin layer of callus tissue; placing the explant material with the thin layer of callus tissue into an Agrobacterium infection solution containing auxin IBA, cytokinin ZT, and acetosyringone for infection and cultivation; performing ultrasonic fragmentation and vacuum infiltration on the material during the infection and cultivation process to promote the entry of the Agrobacterium plasmid into the plant material; then co-culturing the infected explant material and then transferring the transgenic material to a screening medium for screening, ultimately obtaining magenta (rose-red) bilberry callus tissue expressing a reporter gene (betaine synthesis gene). The invention uses a specific callus induction medium, Agrobacterium infection treatment, co-cultivation medium, co-cultivation time, and resistant callus screening medium to construct the transgenic system of bilberry callus mediated by Agrobacterium tumefaciens, obtaining successfully transgenic rose-red callus tissue, and increasing the conversion rate to 4%.