Optimization Of Soybean Cotyledonary-node Genetic Transformation System And Idenwfication Of Soybean Isoflavone Synthase Gene Transgenic Plant

The Soybean has five thousand years history. It is important that the productionand quality of soybean were improved. The use of transgenic technology canaccelerate to select soybean varieties, they is more suitable for consumers high-qualitysoybeans. Isoflavones based on benzene propane are secondary metabolites in higherplants which are prevent diseases such as cancer, osteoporosis, cardiovascular andother common diseases.In recent years it was caused widespread concern category ofscience and technology workers. In daily life, based on the research of the relevantsynthetase gene in the metabolic pathway is of great significance. Isoflavone synthesisapproach is a branch of plant flavonoids metabolic pathway. The amount of isoflavonesynthesis enzyme expression change or loss of expression function and enzymeinactivation will directly affect the flavonoids metabolic production. IFS2, CHS8andCHI is the key enzyme in soybean isoflavone synthesis pathway,which plays a keyrole of isoflavone synthesis.Soybean transformation is difficult due to low regeneration rate, the affectivefactors such as germination way and time, different ways of explant placemation,plant hormones during shoot induction and elongation. Therefore, this study of jilin47soybean of Agrobacterium-mediated cotyledon transformant in wereoptimizatied.The results showed jilin47cotyledon choose liquid nutrient solution13~14h. The measurement of GUS efficiency showed that liquid contaminationscratches can improve the adhesion of agrobacterium rate, thereby improving theefficiency of infection with high transformation rate.Through the way of explanstplacemention in the co-cultivation,we can found when putting up will improve shootinduction rate. We found the best combination in shoot elongation stage is50mg·L-1the Asn and Gln,5ug·L-1triacontanol can effectively promote the growth of shootelongation and speed. In the rooting stage, the3/8of the MSB medium is the mostsuitable concentration and0.5mg·L-1IBA was added to rooting status can be improved.According to the method of jilin47soybean was infected with Agrobacteriumtumefaciens, the IFS, CHS, CHI was transfered to jilin47, jilin35and Williams82.The of transgenic plants detection including PCR in the T1, T2generation plants,basta painting leaf in T2generation plants, the bar gene strip and Western blot proteindetection for resistantplants. The result showed that the target gene in soybeangenome.We were used HPLC to test the content of isoflavone of transgenic plants comefrom jilin47、jilin35、and Williams82in which isoflavone synthase relatedgenes(IFS2, CHS8and CHI Ⅱ) offspring plants. The results showed that theisoflavones content of three varieties of transgenic plants were increased. In the twovarieties jilin35and jilin47, the isoflavones with IFS gene increased most, CHS geneplayed the second place in increasing the amount of isoflavones, CHIⅡ increasedthe least.