定点整合抗虫基因到油菜叶绿体基因组并获得转基因植株  

Site-specific Integration of Insect-resistant Gene into Chloroplast Genome of Oilseed Rape and Acquisition of Transgenic Plants

作　　者：侯丙凯[1] 胡赞民[1] 党本元[1] 石锐[1] 陈正华[1] 

机构地区：[1]中国科学院遗传研究所,北京100101

出　　处：《植物生理与分子生物学学报》2002年第3期187-192,共6页Journal Of Plant Physiology and Molecular Biology

基　　金：国家自然科学基金资助 (No .39970 6 2 4)

摘　　要：以基因枪法进行了油菜叶绿体基因组的定点转化。载体 pNRAB携带抗壮观霉素的筛选标记基因aadA和抗虫基因cry1Aa10 ,基因的两侧被添加了可用于同源重组的叶绿体DNA序列。基因枪轰击过的油菜子叶柄经植株再生和壮观霉素筛选 ,获得了 36株抗性植株。PCR检测和Southern杂交显示 ,其中 4株的叶绿体基因组已被转化 ,外源基因已被定点整合进叶绿体基因组的rps7和ndhB基因之间。用转基因植株的叶片饲喂二龄小菜蛾 ,1周后幼虫死亡率达33%～ 47% ,存活幼虫的生长明显减慢 。Site-specific transformation of oilseed rape chloroplast genome was achieved by particle bombardment. The plasmid pNRAB carries a spectinomycin-resistant gene (aadA) from bacteria and an insect-resistant gene (cry1Aa10) from Bacillus thuringiensis(Bt) between two DNA fragments homologous with oilseed rape chloroplast DNA. Biolistic delivery of vector DNA, followed by spectinomycin selection, yielded 36 spectinomycin-resistant plants in 1 000 bombarded cotyledon petiels. PCR and Southern blot analysis confirmed that, 4 out 36 putative transformants, the aadA and cry1Aa10 genes had been specifically integrated into the site between rps7 and ndhB genes of chloroplast genomes (Figs.1,2). One week after the second instar larvae of Plutella xylostera were fed with the leaves of transplastomic oilseed rape, 33% to 47% mortalities were observed and the surviving larvae had significantly lower weight than the control. While the transgenic oilseed rapes were less damaged (Fig.3).

关 键 词：定点整合 抗虫基因 油菜 叶绿体基因组 转基因植株 

分 类 号：Q943.2[生物学—植物学]