农杆菌介导的大麦Mlo反义基因转化小麦获得抗白粉病后代

Transgenic Wheat Progenies Resistant to Powdery Mildew Generated by Agrobacterium Transformation of Barley Mlo Anti-Sense Gene

从大麦‘斯特林’幼叶总RNA中分离Mlo基因cDNA完整编码区,反向连接到植物双元载体(pBI-121.2)35S启动子下游,通过农杆菌介导的苗端转化法获得两种小麦基因型(‘烟优2801’和‘烟优361’)的转基因小麦。T0代405株中有55株PCR检测阳性,平均转化率达到13.58%,T0和T1基因组DNA Southern杂交可以证明大麦Mlo基因片段已整合到小麦基因组中并可传递到后代。两种基因型的转基因小麦T0和T1植株在温室及大田中均表现出对白粉病抗性的提高。农杆菌介导的苗端转化法可以简单、快速、高效地获得转基因株系;排除体细胞变异对转基因植株的影响;克服基因型对农杆菌转化的限制,是小麦遗传转化的一种实用方法。

Complete coding region sequence of Mlo cDNA from barley cultivar ‘Stirling＇ was cloned and inserted into the BamHI site of binary vector pBI121.2,driven by 35S promotor in reverse direction.Two wheat （Triticum aestivum L.） genotypes ‘Yanyou2801＇ and ‘Yanyou361＇ were transformed by Agrobacterium tumefaciens with shoot apical meristem transformation method.A total of 55 T0 transgenic plants were obtained,and the average transformation frequency was as high as 13.58%,Southern blot proved Mlo gene was integrated into T0 wheat genome and inherited to T1 progenies.The T0 and T1 transgenic plants of both genotypes showed higher resistance to powdery mildew than the controls in both green house and field.The shoot apical meristem transformation with A.tumefaciens is a simple,dependable and efficient method to generate transgenic wheat.It can overcome the difficult of the restriction to transformation of different wheat genotypes,and can get rid of the somoclonal variations due to tissue culture of immature embryos and calli.