Chen Hao, Alonso Jose M, Stepanova Anna N
Department of Plant and Microbial Biology, Program in Genetics, North Carolina State University, Raleigh, NC, USA.
Methods Mol Biol. 2022;2494:61-98. doi: 10.1007/978-1-0716-2297-1_6.
Protein production from mRNA is one of the fundamental molecular processes in a cell. Accurate genome-wide information on the levels of translation and ribosome distribution on mRNA can be gathered by carrying out ribosome footprinting, aka Ribo-seq. Herein, we present a detailed protocol describing the construction of parallel Ribo-seq and RNA-seq libraries from Arabidopsis seedlings treated with the plant hormone auxin. The improved protocol for ribosome footprint library generation can be easily adapted to analyzing the effects on translation of genetic perturbations and various abiotic and biotic factors to shed the much-needed light on translational regulation in plants.
从mRNA产生蛋白质是细胞中的基本分子过程之一。通过进行核糖体足迹分析（又名Ribo-seq），可以收集全基因组范围内关于mRNA上翻译水平和核糖体分布的准确信息。在此，我们提供了一份详细的方案，描述了从用植物激素生长素处理的拟南芥幼苗构建平行Ribo-seq和RNA-seq文库的过程。改进后的核糖体足迹文库生成方案可以很容易地用于分析基因扰动以及各种非生物和生物因素对翻译的影响，从而为植物中的翻译调控提供急需的见解。
