大豆子叶节遗传转化体系优化及抗逆基因AtNHX5的转化研究  

Study on Optimization of Soybean Cotyledonary Node Genetic Transformation System and the Transformation of Resistance Gene At NHX5

作　　者：杨权[1] 王月月[1] 刘炎光[1] 蒋春志[2] 张孟臣[2] 张洪霞[3] 张洁[1] 王冬梅[1] 

机构地区：[1]河北农业大学生命科学学院,河北保定071001 [2]河北省农林科学院粮油作物研究所,河北石家庄050000 [3]中国科学院上海植物生理研究所,上海200032

出　　处：《大豆科学》2015年第2期205-211,共7页Soybean Science

基　　金：转基因生物新品种培育重大专项(2011ZX08004-002-002;2014ZX0800402B-001)

摘　　要：以河北省优良大豆品种冀豆15、五星2号和NF-58的子叶节为受体材料,利用农杆菌介导法进行遗传转化,探讨了影响农杆菌侵染后子叶节不定芽诱导的因素。结果表明:以萌发6 d、4℃处理24 h的子叶节为外植体,农杆菌侵染后经超声波处理30 s、共培养基中添加20 mg·L-1硝酸银,能够提高子叶节丛生芽诱导率,转化植株经草铵膦筛选以及PCR检测,T0代转化率达0.97%。利用该体系对大豆品种五星2号进行At NHX5基因的遗传转化,获得3株T0代RT-PCR检测阳性植株,且2-1号T1代阳性植株检测具有一定耐盐性,初步证明获得了转At NHX5基因的大豆新材料。In this study,soybean cotyledonary node of Jidou 15, Wuxing 2 and NF- 58 were used as materials. An efficient Agrobacterium-mediated gene transformation system based on the examinations of several factors influencing transfomlation effi- ciency was developed for soybean.the results indicated that the optimum transformation conditions were as follow： seeds ger- minated 6 days, 4℃ low temperature treatment, 20 mg· L-L anti-oxidant silver nitrate during co-cultivation, ultrasonic treat- ment for infected explants 30 s. In the above condition, PCR-positive rate was 0. 97%. Using the optimized system an stress- resistant gene AtNHX5 was transformed. Using the optimized system, the gene was transformed into soybean Wuxing 2. After detected the transgenic plants resistant to glufosinate by PCR, the transformation efficiency was 0. 23%. The expression of At- NHX5 was assessed by RT-PCR analysis. One positive plant of T1 generation was obtained by detection of PCR, which has salt- tolerance. It preliminarily demonstrated that the target gene was integrated into the soybean genome.

关 键 词：大豆 子叶节 At NHX5基因 遗传转化 除草剂 

分 类 号：S565.1[农业科学—作物学]