Development And Identification Of Novel Transgenic Soybean Lines With High Isoflavone Content

Soybean not only a strict self-pollinated crop,but also an important economic crop in China,which contains rich protein and unsaturated fatty acids have an important nutritional value.The improvement of living standards put forward the higher requirements on the quality of soybeans.Soybean isoflavone is secondary metabolites of flavonoid metabolic pathways that play an important role in anti-aging,improving menopausal symptoms,preventing osteoporosis and cancer,reducing cardiovascular diseases,it has been widely used in medical,human health and short supply on the market.Breeding of Isoflavone content varieties is an effective way to ameliorate the current situation.In this study,soybean non-emasculation breeding method was used on the basis of traditional soybean hybrid method,the survival rate of hybrid pods and pseudo hybrid were compared,the results show that the new pollination method without emasculation have a higher survival rate of hybrid pod,and then the pseudo hybrid generation rate had no significant difference.In the same condition,female without emasculation hybrid approach was more scientific,reasonable and convenient,for large-scale promotion the method is more suitable.Using the High performance liquid chromatography to determine the soybean isoflavones grain content in the T4、T5、T6 generation which transformed Isomerase chalcone(CHS)gene、Isomerase chalcone and MYB12B2 transcription factor.The results show that the high transgenic generations have a significantly higher grain isoflavones concentration than its transgenic receptor and genetic stable.In order to further improve the isoflavone content of transgenic soybean,the IFS(Isoflavone synthase)overexpression plants crossbreed with Ji5,the overexpression plants of MYB12B2 transcription factor crossbreed with positive plants of IFS 、 CHI,respectively and F1 plants was got.Combing with agronomic traits,PCR detection technique,Basta leaf smear and Bar gene test strips to detect F1 plants.The results showed that the corresponding transgenic hybrid materials were obtained.The content of isoflavones in IFS,CHI overexpressing plants and F2 hybrids was detected by high performance liquid chromatgraphy(HPLC).The results showed that the contents of total isoflavones could be enhanced in the hybrids of IFS,CHI overexpression plant with Jilu5.The significance analysis demonstrated that the CHI transgenic plants have a better improved to the total isoflavone contents of soybean Ji5 than IFS overexpression plants.The content of total Isoflavones in the F2 hybrids of T(MYB12B2)x T(IFS)or T(CHI)was measured.The research proved that the total Isoflavones content significantly higher(P <0.05),and F1 hybrids of(MYB12B2x IFS)have a higher rate than T(MYB12B2)x T(CHI).