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| 逆转录环介导等温扩增技术检测烟草环斑病毒的研究 | |
| 引用本文： | 张吉红,陈先锋,孙辉. 逆转录环介导等温扩增技术检测烟草环斑病毒的研究[J]. 植物病理学报, 2014, 44(6): 629-633 |
| 作者姓名： | 张吉红 陈先锋 孙辉 |
| 作者单位： | 宁波出入境检验检疫局,宁波 315012; 宁波市农业环境与农产品质量监督管理总站,宁波 315012 |
| 基金项目： | 宁波市科研项目(2011C50080);国家质检总局项目(2013IK279) |
| 摘 要： | 本研究采用环介导等温扩增技术(Loop-mediated isothermal amplification, LAMP),建立了一种快速、简便的烟草环斑病毒检测方法。根据TRSV外壳蛋白编码基因上的8个位点,共设计了6条引物,通过RT-LAMP扩增得到特征性的梯度条带。特异性试验表明,引物对TRSV的检测具有良好的特异性;灵敏度试验显示RT-LAMP比普通RT-PCR高10倍。通过反应温度和时间的优化,该方法只需在水浴锅中60℃等温扩增60 min,整个检测周期约1.5 h,结果采用SYBR green I染色显示,易于观察和判定。 |
| 关 键 词： | 烟草环斑病毒 环介导等温扩增 检测 |
| 收稿时间： | 2013-06-21 |
Research on detection of Tobacco ringspot virus by RT-LAMP |
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| ZHANG Ji-hong,CHEN Xian-feng,SUN Hui. Research on detection of Tobacco ringspot virus by RT-LAMP[J]. Acta Phytopathologica Sinica, 2014, 44(6): 629-633 | |
| Authors: | ZHANG Ji-hong CHEN Xian-feng SUN Hui |
| Affiliation: | Ningbo Entry-Exit Inspection and Quarantine Burean,Ningbo 315012,China;Ningbo Central Station of Agricultural Environment and Agricultural Products Quality Supervision management, Ningbo 315012,China |
| Abstract: | The study established a rapid and simple Tobacco ringspot virus (TRSV )detection method using Loop-mediated isothermal amplification (LAMP). According to the eight sites on TRSV coat protein coding genes, six primers were designed and characteristic ladder-like bands were obtained by RT-LAMP. Specificity tests showed that the primers were specific for detection of TRSV and sensitivity test showed that RT-LAMP was 10 times higher than RT-PCR. By the optimizing of temperature and time condition, the method just need the pot with a water bath 60℃ isothermal amplification for 60 min, the entire testing cycle is about 1.5 h, and it is very convenient to observe the results by using SYBR green I. |
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