来自α-变形菌的转运RNA的电荷状态分析
Charging State Analysis of Transfer RNA from an α-proteobacterium.
作者信息
Yin Liang, Harwood Caroline S
机构信息
Department of Microbiology, Seattle, USA.
出版信息
Bio Protoc. 2020 Dec 5;10(23):e3834. doi: 10.21769/BioProtoc.3834.
Transfer RNA (tRNA) is an essential link between the genetic code and proteins. During the process of translation, tRNA is charged with its cognate amino acid and delivers it to the ribosome, thus serving as a substrate of protein synthesis. To analyze the charging state of a particular tRNA, total RNA is purified and analyzed on an acid-urea gel. Separated RNA is then transferred to a membrane and detected with a probe for the tRNA of interest. Here, we present an improved protocol to analyze the tRNA charging state in the α-proteobacterium . Compared to the classical method, the RNA isolation step is optimized to suit this organism. Additionally, a non-radioactive platform is used for electrophoresis and Northern blots. This significantly reduces the time and the effort required for this protocol.
转运RNA（tRNA）是遗传密码与蛋白质之间的重要纽带。在翻译过程中，tRNA被其同源氨基酸负载，并将其输送到核糖体，从而作为蛋白质合成的底物。为了分析特定tRNA的负载状态，需纯化总RNA并在酸性尿素凝胶上进行分析。然后将分离的RNA转移到膜上，并用针对感兴趣tRNA的探针进行检测。在此，我们提出了一种改进的方案来分析α-变形杆菌中的tRNA负载状态。与经典方法相比，RNA分离步骤进行了优化以适应该生物体。此外，非放射性平台用于电泳和Northern印迹。这显著减少了该方案所需的时间和工作量。
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