国家自然科学基金项目(项目批准号:30960158),内蒙古自然科学基金项目(项目编号:K32227)和国家转基因专项(任务编号:2009zx08009-019B)
蒙古沙冬青[Ammopiptanthus mongolicus (Maxim.) Cheng f.] 是分布于我国西北荒漠区的常绿旱生阔叶灌木,因其富含多糖和多酚等次生代谢物质,用常规RNA提取方法难以从中获得高质量总RNA。本研究通过在热酚法的RNA提取液中加入高浓度的KAc和β-巯基乙醇,从该植物的不同样品中提取到高质量总RNA,并用筛选到的合适试剂盒分离到高纯度的mRNA。所得到的总RNA和mRNA已被成功应用于基因克隆和SMART全长cDNA文库的构建。
Ammopiptanthus mongolicus (Maxim.) is an evergreen broad-leaf shrub in the northwest desert of China. Because of the high content in polysaccharides, polyphenols and other secondary metabolites, it is difficult to extract high-quality total RNA from the plant with conventional methods. In this research, by adding high concentration of potassium acetate and β-mercaptoethanol into the extraction buffer of Hot Phenol Method, we successfully extracted high-quality total RNA from different samples of the plant. We also obtained high-purity mRNA with suitable mRNA isolation kit. Both the total RNA and mRNA have been successfully applied to gene cloning and SMART (Switching Mechanism At 5’end of RNA Transcript) cDNA library construction.
林清芳.蒙古沙冬青总RNA提取与mRNA分离方法的研究[J].植物遗传资源学报,2011,12(3):460-463.
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