提取高质量茶树总RNA的方法研究

由于茶树富含多糖、多酚,从茶苗组织中尤其是从茶苗的根中提取高质量的RNA较为困难,试用了多种方法都未能获得高质量的RNA.为此,作者借鉴多年生的草本植物RNA提取方法-CTAB法,并在此基础上进行了改进,首次提取茶苗嫩根中的总RNA.电泳检测显示,该方法提取的总RNA 28S和18S条带清晰且28S较18S条带亮,紫外光谱分析显示A260/A280的比值为1.93.用于RT-PCR反应,成功克隆了492 bp的谷氨酰胺合成酶基因片段.说明用该方法提取的RNA纯度和完整性好.试用该方法从营养成分丰富的茶籽胚中提取总RNA,效果同样很好.

Method for high-quality total RNA isolation from tea plant [Camellia sinensis (L.) O.Kuntze]

Extraction of high-quality RNA from tea plant tissue especially from roots is difficult due to high levels of phenols and polysaccharides.We experienced many published protocols for RNA isolation from the tender roots of tea plant but failed to yield high-quality RNA for analysis.Therefore,we developed a modified CTAB method.The results indicated there are two clear bands for 28S and 18S rRNA,and A260/A280 absorbance ratio is 1.93.One gene fragment about 492 bp that encodes glutamine synthetase was successfully amplified by RT-PCR,suggesting the purity and integrity of isolated RNA.The method of CTAB is also suitable for extraction total RNA from embryo of tea seed.