Study On Molecular Strategies For Salt-tolerant Soybean Breeding

Soybean is an important oil crop plant. High soil salinity and the eelworm limit severely soybean growth, development and productivity. Therefore, developing new cultivars with enhanced salt stress and eelworm tolerance would undoubtedly have an enormous impact on global soybean production. The object pf this study was utilizing the high effective Agrobacterium-mediated transformation system of soybean embryo tip to introduce three methytransferase genes related to anion scavenging from different salt-tolerant plants into soybean,in order to explore a new route for plant gene engineering, and at the same time to provide new information for soybean genetic breeding.The study has explored the sterilization method, the optimization ofexplant, the optimization of medium and rhizogenesis method, in order to establish an high effective regenesis system of soybean embryo tip;we investigate the important factors such as filtration press, the concentration of Agrobacterium, dip-dying time and the employment of Agrobacterium inhibitor on the course of soybean genetic transformation;we introduce the methytransferase genes related to anion scavenging seperately from Arabidopsis thaliana, Thellungiella salsuginea and Suaeda salsa into soybean by Agrobacterium-mediated transformation system,and the transformation plants were tested by PCR,RT-PCR,Southern blotting analysis.We have established the high effective regenesis systemof soybean embryotip:the mature soybean seeds were soaped for 24h, then were dealed with 1.2% NaClO and 0.1% HgCl2 (1:1 mixed) for 10min. The embryo tips were collected and cultured on MS medium supplemented with 0.08 mg/L TDZ for 30d, then explants with multiple mud tissues were transferred to MS medium supplemented with 2.0 mg/LKT + 0.2 mg/L NAA for 20d. The induced seedings long than 5cm were transferred onto root-inducing medium (1/2MS+NAA 0.5mg/L) for 30d. Finally,they were transplanted into soil when the roots reached 3-4cm;We also opmized the transformation system of soybean embryo tip:the concentration of kanamycin was 90mg/L,the concentration of Agrobacterium wasOD₆₀₀=0.6,the dip-dying time was 12h, ampicillin can act as Agrobacterium inhibitor;After screening AtMCT,ThMCT and SsMCT transformation plants were78,67,82seperately,the results of PCR,RT-PCR,Southern blotting analysis on transfoemation plants were positive.This is the first time that we induce and differentiate soybean embryo tip with TDZ. the study establish high effective regenesis system of soybean embryo tip,the differentiation rate of multiple bud is higher than 92.6%;optimize the Agrobacterium-mediated transformation system of soybean embryo tip,the transformation rate is higher than 15%;the first time to heterologus express the methytransferase genes related to anion scavenging seperately from Arabidopsis thaliana, Thellungiella salsuginea and Suaeda salsa into soybean,The results of molecular analysis indicate that heterologus genes had been integrated into the transgenic plant genomes and expressed.