农杆菌介导的RSVIRBSDV双抗基因共转化水稻研究.pdfVIP

摘 要 水稻是我国最重要的粮食作物之一，其产量和品质对于我国的粮食生产和粮食安 全都至关重要。水稻条纹叶枯病（RSV）和黑条矮缩病（RBSDV）为水稻主要的病 毒病害，严重影响水稻的生产，因此培育抗病水稻新品种是水稻育种的重要目标。另 外，由于对转基因产品安全性的争议日趋激烈，研究开发去除标记基因的转化系统势 在必行，利用共转化系统去除标记基因具有简便快捷的特点。 97 本研究以晚粳 成熟胚作为起始材料，采用农杆菌双载体共转化的方法，将编码 RSV和RBSDV的双抗CP基因的干涉载体与编码潮霉素抗性基因（hpt）的表达载体转 入水稻，获得如下主要结果： 1 97 32 90.3% ．皖粳 成熟胚在 ℃光照条件下诱导愈伤的出愈率高达 ，且愈伤生长较 快； 2．选用次生愈伤组织更适合农杆菌侵染，且在铺有滤纸的共培养上23℃黑暗培 48h 养 ，可大大降低农杆菌的污染机率，并可提高抗性愈伤率。 3．抗性愈伤经50mg/L的潮霉素筛选培养基筛选3周后，转入分化培养，共获得 690株水稻再生植株，PCR检测结果显示，其中85株为双阳性转化植株。 4 Southern CP ．部分转化植株经 杂交检测，表明外源 基因已成功整合到水稻基因 组中。 关键词：水稻；共转化；农杆菌介导；标记基因；Southern杂交 Abstract Abstract AAbbssttrraacctt Rice(Oryza sativa) is the staple food that feeds most of the population in China. Its yield and quality is crucial for both the food production and food security. Rice stripe virus (RSV) and rice black steaked dwarf virus (RBSDV) are main viral diseases which severely affect rice production. Thus, breeding novel rice varieties with disease resistance becomes the key goal of rice breeding.Toward the daily increasing controversy on the safety of transgenic product, development of a transformation system capable of removing off the marker gene becomes an urgently required issue. Cotransformation system has the advantages of simple operation procedure and conveniently acquiring of marker-free plants. Employing mature embryos as starting material, we transformed fused CPgene of RSV and RBSDV (abbreviated as RSV/RBSDV gene) into Japonica rice Wanjing97 via Agrobacterium-mediated cotransformation. Themain results are asfollows: 1. The callus emerging rate could b