适用于基因全长克隆的灰毡毛忍冬不同器官总RNA提取方法筛选  

Screening of Total RNA Extraction Methods For Full-length Gene Cloning from Different Organs in Lonicera macranthoides Hand.-Mazz.

作　　者：刘湘丹[1] 徐玉琴[1] 王珊[1] 童巧珍[1] 周日宝[1] 

机构地区：[1]湖南中医药大学药学院,湖南长沙410208

出　　处：《湖南中医药大学学报》2015年第12期60-63,共4页Journal of Hunan University of Chinese Medicine

基　　金：国家自然科学基金(81203007);湖南省自然科学基金(13JJ9010);湖南省教育厅科技创新平台(14K071);湖南省"中药学"重点学科建设项目资助(湘教通[2011]76号);国家中医药管理局"药用植物学"重点学科资助(国中医药发[2009]30号);中药学湖南省重点学科开放基金(zy201403);湖南中医药大学青苗计划

摘　　要：目的获取灰毡毛忍冬不同器官总RNA的最佳提取方法。方法以灰毡毛忍冬叶、茎、花蕾为材料,分别采用SDS法、Trizol法、改良CTAB法、多糖多酚试剂盒法提取总RNA,比较分析实验结果。结果 SDS法不适用于三种器官样品总RNA的提取;Trizol法仅适用于叶总RNA的提取;改良CTAB法对茎总RNA提取效果最佳;而多糖多酚试剂盒法提取的花蕾总RNA质量高、完整性较好,且通过RT-PCR扩增能获得特异性条带。结论 Trizol法最适于灰毡毛忍冬叶总RNA提取;改良CTAB法最适于茎总RNA提取;多糖多酚试剂盒法最适于花蕾总RNA提取;3种方法提取出来的总RNA均能满足灰毡毛忍冬中相关基因全长克隆及时空表达分析的要求。Objective To screen of total RNA extraction methods from different organs in Lonicera Macranthoides Hand.-Mazz（L. Macranthoides）. Methods The total RNA of leaf, stem and flower buds from L. macranthoides, respectively, were isolated by SDS, Trizol, improved CTAB and polysaccharide polyphenols kit methods, and the results were compared. Results SDS was not suitable for this experiment; Trizol only could isolate total RNA from its leaf. Improved CTAB was the best way to isolate total RNA from its stem; Polysaccharide polyphenols kit method could isolate total RNA from its flower buds, and obtained specific bands by RT-PCR. Conclusion Trizol can be used to isolate the total RNA from leaf of L. macranthoides.Improved CTAB was the best method to isolate the total RNA from its stem; Polysaccharide polyphenols kit method could be used to isolate the total RNA from its flower buds. The total RNA by the three methods could meet the request of full-length gene cloning and temporal and spatial expression analysis.

关 键 词：灰毡毛忍冬 基因全长克隆 总RNA 不同器官 提取方法 

分 类 号：R284.1[医药卫生—中药学]