摘要
用修改了的异硫氰酸胍法从两种森林昆虫Clostera an-astomosis和Saperda populnea的不同组织中快速提取总RNA。电泳检测和cDNA合成分析结果表明,RNA未降解。经紫外分光光度计检测,A260/A280在1.8到2.0之间,表明RNA的纯度很高。用RT-PCR合成的双链cDNA的长度大于2kb,表明mRNA的完整性。用PCR的方法克隆了C.anasto-mosisβ肌动蛋白和几丁质酶的基因片段,表明RNA可用于其它分子操作。使用这个方法,在4个小时内可至少从8个样品中提取RNA并进行电泳分析。这些结果表明用这个修改后的一步法提取昆虫总RNA是省时,省钱和有效的。图3参10。
A modified guanidinium isothiocyanate method was used to extract total RNA from two forest insect species Clostera anastomosis and Saperda populnea. The integrity of RNA was demonstrated by the methods of gel electrophoresis and cDNA analysis. Typical A260/ A280 absorbance ratio of the total RNA was in range of 1.8 to 2.0. The size of double strand cDNAs obtained by RT-PCR was more than 2 kb, which indicated that intact mRNA was obtained. The fragments of β-actin and chitinase gene from the RNA of C. anastomosis were obtained by RT-PCR, which indicated that the RNA could be used for other molecular operation. By this procedure, RNAs could be extracted and analyzed by electrophoresis from at least 8 samples within 4 hours. These results showed that this method was time- and cost-saving and effective.
基金
This study was supported by Key Project of Harbin City (2005AA6CN074)
