改良Trizol法从灌浆期小麦胚乳中提取高质量总RNA的研究

An Improved Trizol Method for Extracting High-quality Total RNA from Wheat Endosperm at Filling Stage

[目的]在传统的Trizol法基础上进行改良,探索一种适合于灌浆期小麦胚乳高质量总RNA提取的方法.[方法]用改良的Trizol法从花后10、15、20 d的小麦胚乳中提取总RNA,经核酸分析仪及电泳检测其质量,进行RT-PCR扩增小麦SBEⅡa基因片段并测序比对.[结果]OD260/OD280:1.95～1.98,OD260/OD230:2.0～2.25,产率:544.00～645.46 μg/g,表明提取的总RNA浓度和纯度均达到要求.测序及Blast比对表明成功克隆小麦SBEⅡa基因片段.[结论]改良后的Trizol法可以高效的从灌浆期小麦胚乳中提取高质量的总RNA.

【Objective】The main aim was to isolate high-quality total RNA from wheat endosperm at filling stage by using an improved Trizol method.【Method】Total RNA was extracted from wheat endosperm of 10,15,20 day post anthering using improved Trizol method.Ultraviolet speetrophotometer and agarose gel electrophoresis were used to detect its quality.SBEⅡa gene fragment was amplified by RT-PCR,then sequencing and blasting comparison.【Result】The results showed that the value of OD 260/OD 280 was between 1.95 and 1.98,OD 260/OD 230 between 2.0 and 2.25,and the product yield between 544.00 and 645.46μg/g.This indicated that the total RNA concentration and purity meet the experimental requirements.The sequencing and Blast results showed that wheat SBEⅡa gene fragment was cloned.【Conclusion】The improved Trizol can be efficiently used to extract high-quality total RNA from wheat endosperm.