日本结缕草总RNA提取和mRNA分离方法的优化  

The Optimization of Total RNA Extraction and mRNA Isolation from Zoysiagrass(Zoysia japonica)

作　　者：代小梅[1] 程晓霞[1] 曾会明[1] 韩烈保[1,2] 

机构地区：[1]北京林业大学草坪研究所,北京100083 [2]长江大学园艺园林学院,荆州434025

出　　处：《生物技术通报》2010年第12期132-136,共5页Biotechnology Bulletin

基　　金：国家"863"计划(2006AA10Z132);"十一五"国家科技支撑项目(2006BAD01A19;2006BAC18B04-1);北京市重点实验室和北京市重点学科专项

摘　　要：介绍了分别以日本结缕草根部、叶部、匍匐茎等为材料的总RNA提取的改良步骤,质量和浓度检测及其注意事项,并采用Promega公司的Poly ATract Systems Ⅲ(Z5300)试剂盒进行mRNA分离,尽管这一方法成熟可行,但仍存在洗脱体积较大,常需先沉淀浓缩后才能进行后续试验(如cDNA合成),在制备少量mRNA时回收率较低等问题。据此,对mRNA分离方法作了改良,介绍了mRNA分离的优化步骤,通过增加体系形成了一种更适合于日本结缕草mRNA分离的方法,从而为日本结缕草的分子生物学研究提供基础资料。结果证明所提取的总RNA和mRNA完整,质量较高,并应用到日本结缕草cD-NA文库的构建。By using the roots,leaves,or stolons of Zoysia japonica for the material,this paper described the improvement steps of the total RNA extraction,the detection of the quality and concentration of the total RNA,and the precautions during operations.Then the mRNA was separated by using the kit of Promega＇s PolyA Tract Systems Ⅲ（Z5300）.Although this approach is mature and feasible,there are still some problems such as larger elution which needs to be first precipitated and concentrated before follow-up experiments（such as cDNA synthesis）,low recovery rates in the preparation of a small amount of mRNA.Accordingly,this study made an improved method for the isolation of mRNA,and introduced the optimization steps for the separation of mRNA.By increasing the system,a more suitable method of the mRNA separation from Zoysia japonica was developed so that it provided the basis information for the molecular biology studies of Zoysia japonica.The results show that the total RNA and mRNA are of high quality and integrity and can be used for a cDNA library construction of Zoysia japonica.

关 键 词：日本结缕草 总RNA mRNA分离 

分 类 号：S688.4[农业科学—观赏园艺]