摘要
为寻找高效快速提取植物总RNA的检测方法,以白肋烟叶片为材料,用Trizol试剂盒提取总RNA。依据黄瓜花叶病毒和马铃薯X病毒的外壳蛋白保守序列设计特异性引物,进行RT-PCR检测,从感病叶片中分别扩增出326bp、392bp的目的片断,而对照叶片中无此扩增带。将PCR产物连接pGEM-T载体上,转化大肠杆菌DH5α,得到了含有目的片断的重组子。序列分析表明,与Gene Bank中报道的这两种病毒核苷酸序列进行比较,同源性均达到96%以上。从而证明Trizol试剂盒可以在1 h内提取到纯度高、含量高、完整性好的总RNA,快速、简便、可靠。
Total RNA from the leaves of N. tabacum cv. White burly was extracted by Trizol Reagent. Using the method, it was obtained the total RNA with the high purity, high quality and integrality in 1 h. Special primers of cucumber mosaic virus and potato X virus were designed respectively. The expected sizes of 326bp and 392bp were amplified by RT-PCR separately from the infected sample, but not from the control sample. The PCR products were ligated to pGEM-T Vector, sequencing validated their identities to the consensus sequence in the GenBank. The homology both achieve above 96%. That method is suitable for detecting RNA viruses by RT-PCR rapidly, simply, and reliably.
出处
《北京农学院学报》
2006年第4期68-70,共3页
Journal of Beijing University of Agriculture
基金
北京市教委会资助项目(KM200610020007)
北京市自然基金资助项目(5043026)
北京市市管高校人才强教计划资助项目
