不同提取方法在橡胶树总RNA提取中的应用研究

总RNA的纯度和完整性对橡胶树分子生物学实验至关重要。为探索更适合橡胶树总RNA的提取方法,本文以巴西橡胶树无性系热研7-33-97叶片为试材,利用凝胶电泳、紫外吸光值测定、RT-PCR检测等方法对CTAB-LiCl法和CTAB-NaAc法提取的总RNA进行比较。结果显示:CTAB-LiCl法提取RNA不仅耗时产率低,且RNA溶液中的Li+和Cl-影响反转录效率。CTAB-NaAc法提取叶片的总RNA经DNaseⅠ处理后凝胶电泳条带完整,产率高,无DNA污染;A260/A280比值为2.03,A260/A230比值为1.96;反转录的cDNA经PCR检测质量较好。另用CTAB-NaAc法对橡胶树的花、树皮、胚、根进行总RNA的提取,均能得到较高质量的总RNA。

Application of CTAB-NaAc Method to Extract Total RNA from Rubber Tree

The purity and integrity of total RNA are essential for molecular biology experiments of rubber tree.In this paper,to get a more appropriate method of extracting total RNA of rubber tree,we used the CTAB-LiCl method and the CTAB-NaAc method to extract total RNA from leaves of the brazil clones of Hevea brasiliensis(CATAS 7-33-97),and compared the two methods by gel electrophoresis,UV-value determination,RT-PCR and so on.The results showed that the yield of total RNA by CTAB-LiCl method extraction is low,and the Li+ and Cl-in RNA solution are side effects on RT efficiency.The total RNA extracted from rubber tree by CTAB-NaAc method after treated with DNaseⅠ,its gel electrophoresis with neat,complete,high production rates,no DNA contamination.A260/A280 ratio is 2.03,A260/A230 ratio is 1.96.Gel electrophoresis band of cDNA dispersion range,and the cDNA has a better performer in the subsequent PCR experiments.Using this method to extract total RNA from flowers,bark,embryo,roots and leaves of the rubber tree,all have been got higher-quality total RNA.