不同沉淀方法处理胶乳总RNA对基因扩增效率的影响

doi:10.13560/j.cnki.biotech.bull.1985.2016.11.014

生物技术通报 ›› 2016, Vol. 32 ›› Issue (11): 124-129.doi: 10.13560/j.cnki.biotech.bull.1985.2016.11.014

不同沉淀方法处理胶乳总RNA对基因扩增效率的影响

吴春太¹, 黎瑜¹, 冯伟瑜², 曾日中¹

1. 中国热带农业科学院橡胶研究所农业部橡胶树生物学与遗传资源利用重点实验室海南省热带作物栽培生理学重点实验室,儋州 571737;
2. 海南大学农学院,海口 570228

收稿日期:2016-03-28 出版日期:2016-11-25 发布日期:2016-11-11
作者简介:吴春太,男,博士,副研究员,研究方向：橡胶树产胶特性研究,E-mail：chuntaiwu@163.com;黎瑜同为本文第一作者
基金资助:
国家自然科学基金资助项目（31270713）,橡胶研究所基本科研业务费资助项目（1630022012008）

Effects of the Latex Total RNA Precipitated with Different Methods on Amplification Efficiency

WU Chun-tai¹, LI Yu¹, FENG Wei-yu², ZENG Ri-zhong¹

1. Rubber Research Institute,Chinese Academy of Tropical Agriculture Sciences/Key Laboratory of Biology and Genetic Resources of Rubber Tree,Ministry of Agriculture,the People's Republic of China/Hainan Provincial Key Laboratory of Physiology for Tropical Crops,Danzhou 571737;
2. College of Agriculture,Hainan University,Haikou 570228

Received:2016-03-28 Published:2016-11-25 Online:2016-11-11

摘要/Abstract

摘要： 为探明沉淀方法对胶乳总RNA提取及PCR扩增效率的影响,并寻求较好的RNA沉淀方法,针对胶乳中蛋白、多糖、橡胶粒子含量高的特点,在本实验室改进的SDS法的基础上,分别用LiCl、NaCl+异丙醇、NaCl+NaAc+无水乙醇和NaAc+无水乙醇对橡胶树胶乳总RNA进行沉淀,并对所得RNA的完整性、纯度、含量和RT-PCR扩增效率进行比较。结果表明,4种方法均可获得完好的RNA,但其纯度、含量和PCR扩增效率随方法不同而有所差别,其中NaCl+NaAc+无水乙醇沉淀的橡胶树胶乳总RNA纯度、产率和18S、REF基因扩增效果均较高,故认为用NaCl+NaAc+无水乙醇沉淀胶乳总RNA是一种较为有效的方法。

关键词: 橡胶树胶乳, 总RNA提取, RNA沉淀

Abstract: This work is to find out the effects of different precipitation methods on the latex total RNA and amplification efficiency of PCR as well as to seek the better precipitation method. According to the characteristics that Herva brasiliensis latex was rich in proteins,polysaccharides and rubber particles,on the basis of SDS method modified in our laboratory,the total RNA in the latex of H. brasiliensis was precipitated respectively with LiCl,NaCl + isopropanol,NaCl + NaAc + absolute ethanol,and NaAc + absolute ethanol,and then the integrity of the RNA,purity,content,and RT-PCR amplification efficiency were compared. The results indicated that complete RNA was obtained by any of 4 methods. But the differences in purity and content of RNA and amplification efficiency of RT-PCR among 4 methods were also observed. In which the purity and yield of total RNA precipitated with NaCl + NaAc + absolute ethanol and 18S and REF gene amplification efficiency of latex tissues in rubber tree were higher,indicating that NaCl + NaAc + absolute ethanol was effective method to precipitate total RNA.

Key words: Hevea brasiliensis latex, total RNA extraction, RNA precipitation

吴春太, 黎瑜, 冯伟瑜, 曾日中. 不同沉淀方法处理胶乳总RNA对基因扩增效率的影响[J]. 生物技术通报, 2016, 32(11): 124-129.

WU Chun-tai, LI Yu, FENG Wei-yu, ZENG Ri-zhong. Effects of the Latex Total RNA Precipitated with Different Methods on Amplification Efficiency[J]. Biotechnology Bulletin, 2016, 32(11): 124-129.

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不同沉淀方法处理胶乳总RNA对基因扩增效率的影响

Effects of the Latex Total RNA Precipitated with Different Methods on Amplification Efficiency

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