抗纹枯病、根腐病的转SN1基因小麦的获得与鉴定

SN1是源于马铃薯的一种抗菌肽, 可以抑制多种植物病原菌的生长。小麦纹枯病(主要病原菌为禾谷丝核菌Rhizoctonia cerealis)和根腐病(主要病原菌为平脐蠕孢菌Bipolaris sorokiniana)是小麦的主要土传真菌病害。本研究利用基因工程技术构建了SN1基因的单子叶植物表达载体pA25-SN1, 它受玉米泛素(ubiquitin)启动子的控制;采用基因枪法将pA25-SN1转化小麦推广品种扬麦18幼胚愈伤组织4 000块, 获得203株再生植株, 通过PCR检测出阳性植株55株, 转化率为1.38%。对转SN1基因小麦T₀~T₂代植株, 进行外源基因的PCR、Southern blot、RT-PCR、荧光定量RT-PCR(Q-RT-PCR)分析和小麦纹枯病菌与根腐病菌接种及其抗病性鉴定。结果表明, 转入的SN1基因已经整合到转基因小麦的基因组中, 能够在转基因小麦中遗传、转录与表达。SN1基因的表达提高了转基因植株对小麦纹枯病和根腐病的抗性, 其抗病性可以遗传。

Development and Characterization of SN1 Transgenic Wheat Plants with Enhanced Resistance to Rhizoctonia cerealis and Bipolaris sorokiniana

SN1 is an anti-microbial peptide in potato,and can inhibit the growth of important phytopathogens.In this study,the transformation vector of SN1 gene expressing highly in monocot plants,pA25-SN1,was correctly constructed,in which SN1 gene was driven by maize ubiquitin promoter.Four thousand embryo calli of Yangmai 18 were bombarded by the particle containing pA25-SN1.Among the regenerated 203 plants,55 positive transgenic individuals were identified by PCR assay in the T0 generation with a transformation frequency of 1.38%.The transgenic wheat plants from T0 to T2 generations were subjected to PCR,Southern blot,RT-PCR,and Q-RT-PCR assays.The disease responses of these positive transgenic plants were also evaluated by inoculating with Rhizoctonia cerealis and Bipolaris sorokiniana.The results showed that SN1 gene was integrated into these transgenic lines,inherited from T0 to T2,and expressed in the wheat background.The transgenic wheat plants expressing SN1 displayed an enhanced resistance to R.cerealis and B.sorokiniana compared with the untransformed Yangmai 18,and the enhanced resistance was inheritable.