一种适于cDNA-AFLP分析的茶树叶片总RNA提取方法

A Suitable Method of Total RNA Extraction from Tea Leaves for cDNA-AFLP Analysis

茶树富含多糖、多酚,从茶树组织中提取高质量的RNA较为困难,cDNA-AFLP技术对RNA质量要求较高,因而从茶树叶片中提取高质量的RNA是cDNA-AFLP试验正常进行的重要保障。该试验采用改进的CTAB法从茶树叶片中提取总RNA,结果表明：琼脂糖凝胶电泳显示28S、18S、5S 3条带清晰,完整性好,产率高;A260/A280值为2.07,范围为1.8~2.1,DNA和蛋白质污染较少;经逆转录、双链cDNA合成和cDNA-AFLP检测,证实提取的RNA能满足cDNA-AFLP技术对RNA质量和数量的要求。该方法为应用cDNA-AFLP技术顺利开展茶树分子生物学方面的研究奠定了基础,处理方式简单且成本较低。

Tea plant is rich in polyphenols,polysaccharides,it＇s difficult to extract high quality of RNA from tea organization,cDNA-AFLP technology requires high quality RNA,so extraction of high quality RNA from tea leaves is the important safeguard of cDNA-AFLP experimental.This paper adopted the improved CTAB extraction method of total RNA from tea leaves.Three bands of 28S,18S and 5S displayed clarity and good integrity by agarose gel electrophoresis,and the total RNA with high yield.The ratio of A260 and A280 of between 1.8 and 2.1,was 2.07,a spot of DNA and protein was polluted.It confirmed that the quality and quantity of the extraction of RNA could satisfy the requirements of reverse transcription and cDNA-AFLP on RNA after reverse transcription,double strand cDNA synthesis and cDNA-AFLP testing.The method laid a foundation for smoothly developing tea plant molecular biology research on the application of cDNA-AFLP technology,it is a simple and inexpensive method.