Agrobacterium-mediated Soybean Transformation Of GmUBC2Using Cotyledonary Nodes

In this study, soybean varieties wuxing2was used as material to optimize the systemof agrobacterium-mediated transformation. The genomic DNA extraction method indifferent growth periods with different soybean varieties was optimized. Based on thisresult, the genetic transformation of resistance Gene GmUBC2was studied. Using the PCRpositive plants of T1generation as experimental materials, preliminary discussed theefficiency of the LAMP technology detection of transgenic soybean. Results are asfollows:1. The effect of different supersonic combined with vacuum processing, infectionspeed and DTT on transformation efficiency and tryed to find out appropriate screeningconcentration were discussed. The results showed that supersonic treatment was30scombined with vacuum processing was3min, infection speed was190rpm and theconcentration of DTT was1.0mmol/L in infection stage, the average number of multipleshoots was highest and the first two treatment efficiency reached2.56%and2.25%respectively. The appropriate screening concentration of Basta was8.0mg/L on screeningmultiple shoot clumps stage.2. Different methods for the genomic DNA extraction were used. The results indicatedthat the DNA extracted with the modified CTAB method had higher quality and purity infresh leaves period. The most of polysaccharides and polyphenolic compounds could beremoved efficiently by utilizing new modified CTAB method and the new method wassuitable for extracting high qualified DNA in flowering stage. This study laid thefoundation for soybean molecular researches.3. Based on the optimized system, GmUBC2transformation by agrobacterium-mediated was investigated. The result showed that the PCR positive rate of transformedplants was highest, which was73.15%at stem elongation for21d. T0generationtransgenic plants by PCR detection and acupuncture Basta screening, positive plants were61and the PCR positive rate was0.61%. The number of T1generation plants by PCR andRT-PCR detection were two. The result showed that the GmUBC2had already integrated tothe genome of Wuxing2soybean. 4. LAMP technology compared with PCR detection, which was higher sensitivity andspecificity, the operation was convenient, fast and accurate. LAMP have good consistencywith RT-PCR. It was expected to develop into an effective method for rapid detection oftransgenic soybeans.