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中国农学通报

中国农学通报 ›› 2009, Vol. 25 ›› Issue (23): 78-81.

所属专题： 烟草种植与生产

• 生物技术科学 • 上一篇    下一篇

拟南芥和烟草幼嫩种子RNA不同提取方法的比较

谢传胜 宋国琦 王兴军 夏晗 赵传志 毕玉平   

  • 收稿日期:2009-07-10 修回日期:2009-08-07 出版日期:2009-12-05 发布日期:2009-12-05
  • 基金资助:
    山东省农业科学院博士科研启动基金;山东省农业科学院青年基金

RNA extraction from Arabidopsis thaliana and Nicotiana tobaccum immature seeds using different methods

  • Received:2009-07-10 Revised:2009-08-07 Online:2009-12-05 Published:2009-12-05

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摘要： 采用4种方法分别对拟南芥和烟草幼嫩种子的总RNA进行了提取和分析。结果表明，其中3种方法所提取的总RNA均能满足一般的下游操作。Trizol法提取的RNA质量低。CTAB-异丙醇法提取的总RNA产量最高，并具有提取步骤少，操作时间短，价格便宜等优点，可用于大量样品提取；百泰克试剂盒提取时间短，可在常温下操作；CTAB-LiCl法提取的总RNA基本无基因组DNA的污染，但是不利于小分子RNA的沉淀。经紫外分光光度计检测，3种方法提取的总RNAOD260/OD280均在1.80~1.97，表明蛋白质及其它有机溶剂的污染很少，OD260/OD230在2.03~2.37，表明盐类小分子的污染也较少；电泳检测28S rRNA和18S rRNA条带清晰，28S rRNA的亮度基本为18S rRNA的2倍，所提取的总RNA质量较高。将提取的总RNA 用于反转录，可获得高质量的cDNA，ds cDNA清晰的分布在100 bp~3000 bp之间。

关键词: 绿萍, 绿萍, 消长动态, 水稻产量, 水稻品质

Abstract: Total RNA was extracted from immature seed of Arabidopsis thaliana and Nicotiana tobaccum using four different methods . Trizol reagent resulted in low quantity and poor quality of RNA. The other three methods could isolate RNA from immature seed with quality that match the downstream experiments. Among these methods, CTAB-Isopropyl alcohol method could get the highest yield of total RNA, with the advantages of time saving and low cost. Bioteke kit is also a convenient method to isolate immature seed RNA under room temperature. CTAB-LiCl method could get higher quality total RNA without genome DNA contamination. However, LiCl couldn’t precipitate small molecule RNA well. The quality of total RNA isolated using the above three methods was analyzed with UV spectrophotometer and agarose gel electrophoresis. The OD260/OD280 ratio was 1.80-1.97 and OD260/OD230 ratio was 2.03-2.37. 28S rRNA and 18S rRNA were clearly detected in the RNA electrophoresis. Total RNA isolated by these methods were reverse transcribed and high quality cDNAs were obtained.