Damaj Mona B, Beremand Phillip D, Buenrostro-Nava Marco T, Riedel Beth, Molina Joe J, Kumpatla Siva P, Thomas Terry L, Mirkov T Erik
Department of Plant Pathology and Microbiology, Texas AgriLife Research, Texas A&M System, Weslaco, TX 78596, USA.
Int J Plant Genomics. 2009;2009:765367. doi: 10.1155/2009/765367. Epub 2010 Jan 27.
High-throughput functional genomic procedures depend on the quality of the RNA used. Copurifying molecules can negatively impact the functionality of some plant RNA preparations employed in these procedures. We present a simplified, rapid, and scalable SDS/phenol-based method that provides the high-quantity and -quality RNA required by the newly emerging biotechnology applications. The method is applied to isolating RNA from tissues of two biotechnologically important crop plants, sugarcane and citrus, which provide a challenge due to the presence of fiber, polysaccharides, or secondary metabolites. The RNA isolated by this method is suitable for several downstream applications including northern blot hybridization, microarray analysis, and quantitative RT-PCR. This method has been used in a diverse range of projects ranging from screening plant lines overexpressing mammalian genes to analyzing plant responses to viral infection and defense signaling molecules.
高通量功能基因组学方法依赖于所用RNA的质量。共纯化分子可能会对这些方法中使用的一些植物RNA制剂的功能产生负面影响。我们提出了一种基于SDS/苯酚的简化、快速且可扩展的方法，该方法可提供新兴生物技术应用所需的高质量和高产量RNA。该方法用于从两种具有重要生物技术意义的作物甘蔗和柑橘的组织中分离RNA，由于存在纤维、多糖或次生代谢物，这给分离带来了挑战。通过该方法分离的RNA适用于多种下游应用，包括Northern杂交、微阵列分析和定量RT-PCR。该方法已用于一系列不同的项目，从筛选过表达哺乳动物基因的植物品系到分析植物对病毒感染和防御信号分子的反应。
