快速选育抗黄瓜花叶病毒的转基因烟草纯合品系

Fast Homozygosity of Transgenic Tobacco Line Resistant to Cucumber Mosaic Virus

以来源于烟草推广品种ＮＣ８９花药的单倍体苗为转化受体，同时导入了黄瓜花叶病毒（ＣＭＶ）的外壳蛋白（ＣＰ）基因和卫星ＲＮＡ（Ｓａｔ—ＲＮＡ）基因，在获得的共１１２株转化植株中，有３株在人工接种浓度高达２００μｇ／ｍｌ的ＣＭＶ时仍不发病。通过０．４％秋水仙素处理使之加倍并收获种子，而后连续两代在大田条件下鉴定其ＣＭＶ抗性及综合农艺性状，并与单独导入了ＣＭＶＣＰ基因或ＣＭＶＳａｔ—ＲＮＡ基因的转基因ＮＣ８９品系及未转化的对照ＮＣ８９进行了比较。结果表明，同时表达ＣＭＶ外壳蛋白和卫星ＲＮＡ的双单倍体转基因植株，在Ｒ—１代即表现出一致的可稳定遗传的ＣＭＶ抗性，抗性水平较单独表达ＣＰ或Ｓａｔ—ＲＮＡ的转基因ＮＣ８９提高１倍左右。

Anther -derived haploid plants of a commercial tobacco cultivar NC89 were transformed with a chimeric vector containing coat protein (CP) and satellite RNA (Sat RNA ) genes of cucumber mosaic virus (CMV ). 112 kanamycin resistant transformed haploid plants were obtained and tested for their resistance to CMV by inoculation with high concentration of CMV (200μg/ml). Three plants that showed symptomless 30 days after inoculation were then diploidized by colchicine,and homozygous dihaploid lines with high level of resistance to CMV were obtained. The three transgenic lines were further tested and compared with the transgenic lines expressing CMV CP or Sat-RNA gene only and the non -transformed NC89 plants under field conditions during the following two seasons. The results showed that the progenies of these dihaploid transgenic lines were homozygous after only one generation and were highly resistant to CMV under natrual field infection and mechanical inoculation conditions. The resistance was about twice that was given by the Sat -RNA or the CP gene alone in transformed plants.