70results about How to "Improve the efficiency of genetic transformation" patented technology

The invention relates to an agrobacterium tumfaciens-mediated transgenic soybean regenerated seedling rooting method, and belongs to the technical fields of soybean tissue culture and transgenic research. Research of soybean cotyledon node and embryo genetic transformation is performed by using an agrobacterium tumfaciens mediating method; a novel transgenic plant rooting method is adopted for the first time; and even if a double-layer rooting culture medium is adopted, the upper layer contains antibiotics and the lower layer does not contain antibiotics, the growth of agrobacterium tumfaciens can be inhibited, adverse influence of the antibiotics on transgenic regenerated plants can be greatly reduced, and the transplanting survival rates of the agrobacterium tumfaciens infected plants reach 90.70 percent and 88.10 percent respectively. The application of the method has significance for improving the agrobacterium tumfaciens-mediated soybean genetic transformation efficiency, and has good reference effect on genetic transformation of other plants.

The invention relates to a method for improving the genetic transfer efficiency of gene-transferred soybean. Wherein, it uses agricillin soybean cotyledon transfer system and germ sharp transfer system to obtain transfer adventitious bud; said adventitious buds without growing root and seed are directly made into graft, via cleft grafting method to be grafted on the soybean seed stock; the graft and the stock via combine period, recover period, fusion period and survival period can improve the gene transfer efficiency of soybean. The grafting can improve the survival rate of plant more than 90%, to shorten the gene transfer period and improve the gene transfer research.

The invention relates to the field of plant genetic engineering. Specifically, the invention relates to an efficient watermelon genetic transformation system and application. Growth regulation genes such as AtGRF5, TaGRF4, OsGIF1, ZmWUS and ZmBBM are included in watermelon genetic transformation. It is found for the first time that expression of AtGRF5, TaGRF4, OsGIF1, ZmWUS or ZmBBM genes in the watermelon transformation and/or regeneration process can promote the genetic transformation efficiency of watermelon crops, AtGRF5 can significantly improve the genetic transformation efficiency of watermelons by about 80 times, and precious gene resources are provided for genetic breeding. The invention further provides a method for using a fluorescent protein DsRed2 as a genetic transformation screening marker, and genetic transformation screening operation is simplified. When the system is expressed and used in watermelons, the genetic transformation breeding efficiency of the watermelons can be effectively improved, and the breeding cost is reduced.

The invention provides a rubber tree embryoid induction culture medium, an induction method for an embryonic callus and a culture method for regenerating resistant callus plant, and belongs to the technical field of plant tissue culture. The rubber tree embryoid induction culture medium takes an improved MS culture medium as a basic culture medium, and is prepared from Kt, NAA, 6-BA, salicylic acid, spermidine, acid hydrolyzed casein, coconut liquid endosperm coconut water, active carbon, saccharose and phytagel. The somatic embryo induction rate of the embryoid induction culture medium provided by the invention for inducing the embryonic callus of a rubber tree anther somatic embryo is increased by 2.04 to 2.16 times in comparison with that of an anther dedifferentiation callus.

The invention relates to an efficient and stable genetic transformation method for strawberries mediated by agrobacterium tumefaciens. The method comprises the steps: preculturing aseptic seedlings of strawberries, activating agrobacteria, infecting, co-culturing, delaying screening, and screening and rooting. According to the efficient and stable genetic transformation method, the genetic transformation steps are optimized, so that the transgenosis efficiency of the strawberries is increased by a large margin; the problems that a traditional agrobacterium-mediated method is low in transformation efficiency, chimeras are easy to generate, the screening is difficult and the workload is great are solved; the genetic transformation efficiency is improved enormously; and meanwhile, the false positive rate of regeneration buds is obviously lowered.

The invention discloses a method for genetic transformation of pineapple somatic embryogenic receptor kinase gene AcSERK1 and application thereof. The genetic transformation method of the invention comprises the following steps: 1) construction of a sense expression vector of pFGC5941-35s-AcSERK1; 2) preparation of callus and callus tolerance test to PPT; 3) preparation of agrobacterium infectionsolution suspended by MS; 4) pre-culture, infection, co-culture and PPT screening culture; 5) PPT resistant seedling rooting and transplanting. Transgenic plants obtained by the genetic transformationmethod of the invention are subjected to PCR and Southern blot and tested by RT-PCR, confirming that the AcSERK1 had been inserted into pineapple genomic DNA and overexpression is occurred in pineapple somatic embryos. Functional validation shows that the number of embryogenic callus particle and somatic embryos obtained by callus which overexpresses the AcSERK1 after somatic embryo induction issignificantly higher than that of wild type, indicating that overexpression of the AcSERK1 has significant promotion effect on formation of pineapple embryogenic cells and somatic embryogenesis.

The invention relates to a method for genetic transformation of eucommia ulmoides by injection. According to the method, eucommia ulmoides seedlings germinating for three days are taken, a first trueleaf between two cotyledons is pulled out to serve as a genetic transformation receptor, and agrobacterium resuspensions with different concentrations are injected between the two cotyledons of each eucommia ulmoides seedling. The method comprises the following specific operation steps: (1) preparing the genetic transformation receptor; (2) culturing agrobacterium; (3) preparing the agrobacteriumresuspensions; (4) performing genetic transformation of the eucommia ulmoides; (5) managing a transgenic plant; and (6) identifying the transgenic plant. According to the method, a tissue culture technology is not adopted, besides agrobacterium culture, sterile operation is not needed, and the method has the advantages of being easy to operate, low in cost, capable of converting large-fragment DNA, stable in heredity, low in copy number and the like, and becomes the most widely applied genetic transformation method at present.

The invention belongs to the technical field of kiwi fruit genetic engineering, and particularly relates to a culture medium and a genetic transformation method of Hongyang kiwi fruits. The genetic transformation method sequentially comprises the following steps: agrobacterium tumefaciens culturing, leaf pre-culture, infecting the leaves by the agrobacterium tumefaciens, callus screening and culturing, carrying out bud elongation culture, carrying out bud rooting culture and positive seedlings transplanting; steps of leaf pre-culture, leaf infection by agrobacterium tumefaciens, callus screening and culturing, bud elongation culture and bud rooting culture adopt the corresponding culture medium provided by the invention. According to the method, the genetic transformation time of the kiwifruits can be shortened, the genetic transformation efficiency can be improved, particularly, a culture medium formula for rapidly and efficiently obtaining positive seedlings is found, and the methodplays an important role in shortening the genetic transformation time of the Hongyang kiwi fruits and improving the genetic transformation efficiency.

The invention discloses a method for constructing genetic transformation systems of catalpa bungei. The method includes procedures of screening the critical lethal concentration of hygromycin by the aid of embryogenic calluses of the catalpa bungei; carrying out infection and transformation by the aid of agrobacterium-mediated processes assisted by ultrasonic treatment; detecting resistant regenerated embryogenic calluses and regenerated plants and the like. The embryogenic calluses of the catalpa bungei are used as transformation receptors. The method has the advantages that the embryogenic calluses of the catalpa bungei are used as the receptors by the aid of the method for the first time, accordingly, infection materials are wide in transformation population, and embryogenic cells are high in regeneration capacity; annual genetic transformation requirements can be met by long-term stable subculture storage of the embryogenic calluses; the embryogenic calluses of the catalpa bungei are infected and transformed under the assistance effect of ultrasonic treatment, and accordingly the method is high in transformation efficiency; T-DNA [thymine DNA (deoxyribonucleic acid)] regions of vectors are successfully integrated into genomes of the catalpa bungei by the aid of the method, and specific primers are designed for detecting transgenic catalpa bungei regenerated plants; the method is applicable to molecular genetic improvement and detection on the catalpa bungei, and transgenic catalpa bungei obtained from transformation events can be used as a donor, so that diversified novel transgenic varieties can be obtained.

The invention discloses a construct and an application thereof. The construct comprises a nucleotide sequence shown in SEQ ID NO: 1. The nucleotide sequence shown in SEQ ID NO: 1, namely a blue-green algae ictB gene can be successfully introduced into a receiver plant by using the construct, thus the photosynthetic efficiency of the receiver plant can be remarkably reinforced, and the yield of plants is increased.

The invention discloses a HD-Zip I type transcription factor GmHDL 57 gene with ability of improving salt resistance of leguminous plants. The sequence of the gene is a nucleotide sequence shown as SEQ ID NO:1, and a coded protein sequence is an amino acid sequence shown as SEQ ID NO: 2. The invention further discloses application of the HD-Zip I type transcription factor GmHDL 57 gene with ability of improving salt resistance of leguminous plants in lotus corniculatus. By utilizing an overexpression technique, the salt resistance of the gene in the leguminous plant lotus corniculatus is researched. Results show that compared with control plants, the salt resistance of the plants is obviously enhanced after overexpression, and the gene has great application prospects for widening a planting range of the leguminous plants in saline-alkali soil.

The invention provides application of a corn BBM1 gene for improving the plant genetic transformation efficiency. For the first time, the corn BBM1 gene can promote the genetic transformation efficiency of plants such as corn, and precious gene resources are provided for genetic breeding. The gene can improve the genetic transformation efficiency of the transformed corn variety syndrome-31 and thelike, the corn variety B73 and other transformation efficiency is improved, the corn transformation gene limit can be effectively solved, and the corn transformation acceptor varieties are enlarged.

The invention belongs to the technical field of plant genetic engineering, and provides an agrobacterium-mediated genetic transformation method using the mature embryo of sorghum as an explant to induce callus for the disadvantageous that in traditional sorghum genetic transformation systems, material drawing by using young embryos and young ears as explants is seasonally restricted and also for the tedious operation that embryo stripping is needed for other crops in which the mature crops are used as the explants. Sorghum seeds which are germinated and then appear white are utilized as objects for agrobacterium infection and the explants for callusogenesis induction, resistant callus are obtained through screening and cultivation, and then a regenerated and transformed plant is obtained through subculture, differentiation, rooting, and transplantation. The disadvantages that in an existing sorghum genetic transformation system, the material drawing by using the young embryos and youngears as the explants is seasonally restricted is avoided, the tedious operation that the embryo stripping is needed for other crops in which the mature crops are used as the explants is overcome, andthe sorghum genetic transformation efficiency is greatly improved. According to the agrobacterium-mediated genetic transformation method using the mature embryo of sorghum as the explant to induce the callus, references and technical approaches are provided for performing genetic improvement of sorghum by using biotechnology.

The invention belongs to the field of plant genetic engineering, and discloses an EMP gene for influencing differentiation and development of a rice callus and application of EMP gene in differentiation and development of the rice callus. The nucleotide sequence of the EMP gene is shown as SEQ ID NO:1; the EMP gene is related to the differentiation and development of the rice callus, and the regeneration capacity of the rice callus can be quickly detected and identified by detecting the expression quantity of the EMP gene; and moreover, through overexpression of the EMP gene, the differentiation capacity of the rice callus can be improved, the genetic transformation efficiency of rice is improved, and then the transgenosis transformation efficiency of agrobacterium-induced rice is improved, and the application is of great importance in breeding new rice varieties.

The invention discloses a kit comprising serial culture mediums and its application in soybean genetic transformation. The kit comprises recovery culture medium, screening culture medium, growth culture medium, and rooting culture medium. The recovery culture medium comprises 45-55 mg/L of L- asparagine, 45-55 mg/L L-glutamine, 28-32 mg/L EDTA salt, and 11-13 mg/L FeSO4. The screening culture medium contains 45-44 mg/L L-asparagine, 45-55 mg/L L-glutamine, 28-32 mg/L EDTA salt, and 11-13 mg/L FeSO4. The growth culture medium contains 45-55 mg/L L-asparagine, 45-55 mg/L L-glutamine, 28-32 mg/L EDTA salt, and 11-13 mg/L FeSO4. The rooting culture medium contains 45-55 mg/L L-asparagine and 45-55 mg/L L-glutamine. The application of the kit in soybean genetic transformation belongs to the protection scope of this invention. The kit helps improve soybean genetic transformation efficiency and has high application value.

The invention discloses a method for constructing a ramie genetic transformation system, which belongs to the agricultural biological technical field or the plant genetic engineering field. The method mainly comprises following steps: taking a ramie seedling cotyledon as a transformation explant to be cocultured with agrobacterium, then, picking up the cotyledon which is cocultured, transferring the ramie seedling cotyledon on selective medium to eliminate bacteria and to culture selectively, and transferring a resistant plant which is produced in the selective medium in selective elongation medium and selective rooting medium to carry out elongation culture and rooting culture successively until a resistant bud is regenerated into a complete plant. The resistant bud which is obtained is respectively checked through GUS, PCR and Southern hybridization and the result shows that a foreign gene is integrated in a ramie genome. The method of the invention has simple operation, materials are conveniently obtained, the transformation efficiency is high, and the method can be widely used in genetic engineering operations of the ramie.

The present invention belongs to the field of plant transgenic technology, and discloses a method for improving sugarcane genetic transformation efficiency by taking mannose as a screening agent. The method is based on sugarcane young heart leaves-induced yellowish and granular type II embryogenic callus as a transgenic acceptor material, employs a negative pressure assistant and improved agrobacterium tumefaciens mediated method to lead a 6-phosphate mannose isomerase gene into a sugarcane genome, and enables getting transgenic sugarcane plants after screening and differentiating on mannose-containing medium. The method uses a PMI positive selection marker system, requires a short cycle from sugarcane callusing to differentiating into seedlings, is stable in transformation effects, and helps to overcome the defects that a genetic transformation system taking antibiotics and herbicides as negative selection markers is low in transformation efficiency, long time in embryogenic callus induced differentiation, easy in sugarcane callus browning, etc. The transformed callus maintains a high regenerative capacity, and has a transformation rate of 58.97% showed by molecular detection results.

The invention discloses a method for increasing chloroplast genetic transformation efficiency through the genome editing technique. According to the method, chloroplast genome is subjected to fixed shear through the genome editing technique, thereby increasing recombination efficiency and integration efficiency of exogenous gene in the chloroplast genome, and further increasing the chloroplast genetic transformation efficiency.

The invention belongs to the technical field of gene engineering, and particularly relates to the application of cell-penetrating peptide TAT2 in transgenic cotton cultivation. The cell-penetrating peptide TAT2 comprises 18 amino acids, and the sequence is shown in SEQ ID NO.1. After cotton pistil fertilization is finished, DNA to be transformed and cell-penetrating peptide TAT2 solution which are evenly mixed are injected into an ovary through a pollen tube to be used for carrying a DNA fragment to be transformed to a cotton genome. Due to the fact that the cell-penetrating peptide can bring a foreign DNA into a fertilization cell directly and smoothly and the foreign DNA can enter a cell nucleus, genetic transformation efficiency is improved remarkably compared with the traditional pollen tube path way; besides, the method does not depend on a cotton tissue culture regeneration system, operation is convenient, expensive instruments are not needed, production cost is low, and therefore a more effective method worth application and popularization can be provided for cotton variety improvement and fundamental research.