高质量的小麦种子总RNA的快速提取方法

A Method for Rapid Isolation of High-Quality Total RNA from Wheat Seeds

提取高质量的RNA是从基因表达水平上研究小麦种子发育的必要条件。现有提取方法难以快速得到高纯度的小麦种子总RNA。本试验将冷酚法和Trizol一步法相结合,在5h左右就可得到高质量的总RNA。通过琼脂糖凝胶电泳、紫外分光光度计检测,提取的总RNA具有清晰的28SrRNA、18SrRNA条带,OD260/DO280比值在1.90～2.00之间。将提取的总RNA用于反转录,可获得高质量的cDNA,在cDNA-AFLP分析上可得到清晰的条带。说明这种方法获得的总RNA纯度和完整度非常高,完全满足分子生物学研究的要求。

Isolation of high-quality RNA is a prerequisite to study the development of wheat seeds at the gene expression level. Rapid isolation of high-quality total RNA from wheat seeds is difficult with the extraction methods reported previously. In the present study, cold phenolic method and Trizol single-step method were combined for RNA extraction, and high-quality total RNA was obtained in about five hours. The quality of total RNA was analyzed with agarose gel electrophoresis and UV spectrophotometer. Clear bands of 28S rRNA and 18S rRNA were shown in the RNA electrophoresis, and the value of OD260/OD280 was 1.90 to 2.00. Using the total RNA isolated by this method for reverse transcription, high quality of cDNA could be obtained, and clear bands were detected in subsequent cDNA-AFLP analysis. These results demonstrated that the purity and integrality Of total RNA isolated with the new method were significantly satisfactory for the demands of molecular biological experiments.