LIANG Ling-ling,DENG Yu-xing,HUANG Qiong-lin,et al.Comparison of Total RNA Extraction Methods from Nervilia fordii (Hance) Schltr.Leaves[J].Northern Horticulture,2013,37(02):91-94.
青天葵叶片总RNA提取方法的比较研究
- Title:
- Comparison of Total RNA Extraction Methods from Nervilia fordii (Hance) Schltr.Leaves
- 文章编号:
- 1001-0009(2013)02-0091-04
- 分类号:
- S 567.23+9
- 文献标志码:
- A
- 摘要:
- 采用RNeasy Plant Mini Kit法、RNAiso Plus法、RNAiso for Polysaccharide-rich Plant Tissue法、CTAB法和SDS法，并结合4种抽提试剂组合，分别提取青天葵叶片总RNA。通过琼脂糖凝胶电泳、紫外检测及RT-PCR反应对提取效果进行了比较分析。结果表明：除RNAiso Plus法外，其它方法提取的总RNA均有一定程度的DNA污染。RNAiso Plus法组③(1/5体积KAC+4/5体积PCI)完整性最好，SDS法组④（1/5体积NaCl+4/5体积PCI）浓度最高，为372.4 ng/μL。5种方法共20个处理组获得的RNA反转录后均能扩增出18S rRNA基因片段。该研究建立起了适用于青天葵叶片的总RNA提取方法，为青天葵的基因克隆、表达及转录组分析奠定了技术基础。
- Abstract:
- Five different extraction methods(RNeasy Blant Miri kit,RNAiso Plus,RNAiso for Polysaccharide-rich Plant Tissue,CTAB and SDS)，combined with four different extraction solutions were used to extract total RNA from Nervilia fordii (Hance) Schltr.leaves.The effect and efficiency of extraction was evaluated by agarose gel electrophoresis，ultraviolet spectrophotometry and RT-PCR.The results showed that apart from RNAiso Plus method，total RNA resulted from the other methods all showed genomic DNA pollution.Among these methods，the total RNA resulted from RNAiso Plus method followed with extraction solution ③(1/5KAC+4/5PCI) was the most integrated，and the total RNA resulted from SDS method followed with extraction solution ④(1/5NaCl+4/5PCI) had the highest concentration (372.4 ng/μL).Total RNA obtained from twenty treatment groups have been used to successfully amplify 18s rRNA gene segment by RT-PCR.This study established a proper total RNA extraction method for Nervilia fordii (Hance) Schltr.leaves，and it provided technique basis for gene cloning，expression，and transcriptomic analysis of the plant.
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备注/Memo
第一作者简介：梁凌玲（1987-），女，在读硕士，研究方向为中药资源的可持续利用研究和开发。E-mail:linglingyyzz@126.com.