| 云南栘［木衣］叶片总RNA提取方法的比较与改进 |
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| 引用本文： | 慈晓彤,,石辰,,王大玮,,沈莲文,,何承忠,,蔡年辉,,段安安,.云南栘［木衣］叶片总RNA提取方法的比较与改进[J].西北林学院学报,2020,35(3):95-99. |
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| 作者姓名： | 慈晓彤 石辰 王大玮 沈莲文 何承忠 蔡年辉 段安安 |
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| 作者单位： | （1.西南林业大学,云南省高校林木遗传改良与繁育重点实验室,云南 昆明 650224; 2.西南林业大学,西南山地森林保育与利用省部共建教育部重点实验室,云南 昆明 650224) |
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| 摘 要： | 高质量RNA的提取是开展云南栘［木衣］分子生物学研究的前提。采用OMEGA (Plant RNA Kit 50)试剂盒、TIANGEN(TRNzol-A+Reagent)试剂盒、CTAB法、Trizol法、SDS法及其改良法6种方法提取云南栘［木衣］叶片的RNA,并用琼脂糖凝胶电泳、核酸蛋白检测仪及RT-PCR技术比较了6种不同方法提取的总RNA样品的质量。结果表明，2种试剂盒法和trizol法不适合云南栘［木衣］RNA提取，无法得到28S、18SrRNA条带，CTAB法和SDS法虽能得到RNA的3条带，但拖带现象严重，A260/A230的比值较低，对SDS法进行改进，提高β-巯基乙醇的量，在缓冲液中增加PVP，使用HiBind RNA Mini柱，有效地抑制多酚多糖的污染，得到了较高质量、可用于RT-PCR扩增的云南栘［木衣］的RNA。对其开展分子生物学研究提供技术支撑。
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| 关 键 词： | 云南栘［木衣］ 叶片 RNA 比较与改进 |
Comparison and Improvement of Total RNA Extraction Methods from Docynia delavayi Leaves |
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| CI Xiao-tong,' target="_blank" rel="external">,SHI Chen,' target="_blank" rel="external">,WANG Da-wei,' target="_blank" rel="external">,SHEN Lian-wen,' target="_blank" rel="external">,HE Cheng-zhong,' target="_blank" rel="external">,CAI Nian-hui,' target="_blank" rel="external">,DUAN An-an,' target="_blank" rel="external">. Comparison and Improvement of Total RNA Extraction Methods from Docynia delavayi Leaves[J].Journal of Northwest Forestry University,2020,35(3):95-99. |
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| Authors: | CI Xiao-tong " target="_blank">' target="_blank" rel="external"> SHI Chen " target="_blank">' target="_blank" rel="external"> WANG Da-wei " target="_blank">' target="_blank" rel="external"> SHEN Lian-wen " target="_blank">' target="_blank" rel="external"> HE Cheng-zhong " target="_blank">' target="_blank" rel="external"> CAI Nian-hui " target="_blank">' target="_blank" rel="external"> DUAN An-an " target="_blank">' target="_blank" rel="external"> |
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| Institution: | (1.Key Laboratory for Forest Genetic and Tree Improvement&Propagation in Universities of Yunnan Province,Southwest Forestry University,Kunming 650224,Yunnan,China; 2.Key Laboratory for Forest Resource Conservation and Utilization in the Southwest Mountains of China,Southwest Forestry University,Kunming 650224,Yunnan,China) |
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| Abstract: | RNA extraction is a prerequisite for molecular biology research of Docynia delavayi,a precious tree species occurring in southwestern China.In this study,OMEGA (Plant RNA Kit 50) kit,TIANGEN (TRNzol-A+Reagent) kit,and the methods of CTAB,Trizol,SDS,and improved SDS were used to extract total RNA from D.delavayi leaves.The amounts of total RNA extracted by different methods were compared by gel electrophoresis,nucleic acid protein meter and RT-PCR.The results showed that the two kits and trizol method were not suitable for RNA extraction because 28S and 18S rRNA bands could not be obtained.CTAB and SDS methods could obtain three bands of RNA,but the tailing phenomenon was serious， the ratio of A260/A230 was lower.SDS method could be improved by increasing the addition of β-mercaptoethanol,adding PVP in the buffer,and using HiBind RNA Mini column,which effectively inhibited the contamination of polyphenol polysaccharide,and a higher quality RNA to be used for RT-PCR amplification could be obtained.It laid the foundation for its molecular biology research. |
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| Keywords: | Docynia delavayiDocynia delavayi leaf RNA comparison and improvement |
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