Using multiplex single-base extension typing to screen for mutants defective in RNA editing
Using multiplex single-base extension typing to screen for mutants defective in RNA editing
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DOI：
10.1038/nprot.2012.117
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发表时间：
2012-11-01
期刊：
影响因子：
14.8
通讯作者：
Brennicke, Axel
中科院分区：
文献类型：
--
作者：
Takenaka, Mizuki;Brennicke, Axel
RNA编辑是一种RNA成熟过程，改变特定RNARs中特定位置（编辑位点）的核苷酸;在植物细胞器中，最常见的核苷酸变化是从胞苷（C）到尿苷（U）。在怀疑影响RNA编辑的突变体中，必须分析所有已知的编辑位点。因此，为了筛选突变体群体，必须在每个编辑位点分析所有个体。我们描述了一种多重单核苷酸多态性（SNP）分型程序，以经济地筛选突变个体或群体在RNA或DNA中数百个核苷酸位置的差异。通过使用该方案，我们先前已经在随机突变的拟南芥群体中鉴定了RNA编辑缺陷的突变体。该程序需要2-3周来鉴定突变群体中的个体植物。在拟南芥中定位突变基因所需的时间在3至24个月之间。虽然这种方法是为了研究植物中的RNA编辑而开发的，但它也可以用于研究其他RNA修饰过程。它也可以用于研究其他生物体中的RNA编辑。
RNA editing is an RNA maturation process that changes the nucleotide present at particular positions (editing sites) in specific RNARs; in plant organelles, the most common nucleotide change is from cytidine (C) to uridine (U). In a mutant suspected of affecting RNA editing, all known editing sites have to be analyzed. Therefore, to screen a population of mutants, all individuals must be analyzed at every editing site. We describe a multiplex single-nucleotide polymorphism (SNP)-typing procedure to economically screen a mutant individual or population for differences at hundreds of nucleotide positions in RNA or DNA. By using this protocol, we have previously identified mutants defective in RNA editing in a randomly mutated population of Arabidopsis thaliana. The procedure requires 2-3 weeks to identify the individual plant in the mutant population. The time required to locate the mutated gene is between 3 and 24 months in Arabidopsis. Although this procedure has been developed to study RNA editing in plants, it could also be used to investigate other RNA modification processes. It could also be adapted to investigate RNA editing in other organisms.