杏叶片与果实总RNA提取方法研究

以杏叶片及幼果为材料,分别从操作耗时、RNA质量和产量等方面比较了CTAB法、改良CTAB法、Trizol法、SDS-苯酚法和改良SDS法5种不同的RNA提取方法。结果表明:5种方法均能从幼嫩叶片中提取到总RNA;经电泳检测,28S rRNA和18S rRNA两条主带清晰,A260/A280均大于1.8;其中以Trizol法提取效果最佳,改良CTAB法次之,产量分别达到了839.25μg/g和395.21μg/g。在果实RNA提取过程中,改良SDS法以及改良CTAB法能有效减少果实RNA的提取中酚类物质和多糖等杂质的影响,获得质量较高的RNA,二者产量分别为352.65μg/g和361.95μg/g。每种方法提取的叶片和果实的RNA经反转录后形成cDNA,并进行RT-PCR扩增,成功扩出看家基因泛素蛋白UBQ片段,说明了RNA质量能够满足文库构建、基因克隆等研究需要。

Study on Methods for RNA Extraction from Apricot Leaf and Fruit

The efficiency and the effect of five RNA isolation methods,including CTAB,modified CTAB,Trizol,SDS and modified SDS,were studied in the RNA extraction from apricot leaves and fruits.Of these five methods,the method of Trizol was found to be the best one,and the modified CTAB was better for the leaf RNA isolation.The RNA yields from apricot leaf were 839.25 μg/g and 395.21 μg/g,respectively.The modified CTAB and SDS methods were preferential ones for the isolation of higher quantity and better quality RNA from apricot fruit,the yield were 361.95 μg/g and 352.65 μg/g.All the RNA samples could be successfully used to amplify a specific cDNA fragment of UBQ,a housekeeping gene in plant,which suggests qualification of the RNA samples in apricot cDNA library construction or gene cloning.