Center for Cancer Research, Massachusetts General Hospital, Charlestown, MA, USA.
Department of Medicine, Harvard Medical School, Boston, MA, USA.
Methods Mol Biol. 2021;2252:3-26. doi: 10.1007/978-1-0716-1150-0_1.
Modern DNA sequencing technologies have allowed for the sequencing of tens of thousands of bacterial genomes. While this explosion of information has brought about new insights into the diversity of the prokaryotic world, much less is known of the identity of proteins encoded within these genomes, as well as their rates of production. The advent of ribosome profiling, or the deep sequencing of ribosome-protected footprints, has recently enabled the systematic evaluation of every protein-coding region in a given experimental condition, the rates of protein production for each gene, and the variability in translation rates across each message. Here, I provide an update to the bacterial ribosome profiling approach, with a particular emphasis on a simplified strategy to reduce cloning time.
现代 DNA 测序技术已经可以对成千上万的细菌基因组进行测序。虽然这一信息的爆炸式增长为我们了解原核生物世界的多样性带来了新的视角，但对于这些基因组中编码的蛋白质的身份以及它们的产生速度，我们了解得还很少。核糖体图谱分析（或核糖体保护足迹的深度测序）的出现，最近使我们能够系统地评估给定实验条件下每个蛋白质编码区域的情况，包括每个基因的蛋白质产生率以及每个信息的翻译率变化。在这里，我将提供细菌核糖体图谱分析方法的更新，特别强调一种简化的策略，以减少克隆时间。
