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Biology

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Molecular Biology

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Analyzing Gene Expression and Function

Ribosome Profiling for Translation Snapshot

Description

Ribosome profiling is a deep sequencing method that gives a snapshot of active translation in a cell. It sequences the mRNAs protected by ribosomes. This shows which messages are being read to make proteins at a specific moment.

The method has several uses in biology. It can track translation in ...

Transcript

确定基因组中被转录并翻译成蛋白质的区域,是理解基因组及其表达的关键步骤。

这一目标通过一种称为核糖体谱分析(ribosome profiling)的技术实现,该技术也被称为核糖体测序(ribo-seq)。该方法可定位核糖体在mRNA上的位置,并鉴定出正在被主动翻译成蛋白质的mRNA分子。

为了分离RNA,首先需要裂解细胞以获取其中的分子。随后用RNase处理裂解液,这些酶会降解未被核糖体覆盖的mRNA,仅留下受保护的mRNA片段。

核糖体保护的片段随后通过蔗糖梯度离心与未受保护的、被切割的片段分离。

随后,核糖体从 mRNA 片段上被移除,再通过逆转录酶进行 RT-PCR,将 RNA 转化为 DNA。

接下来,对 D...

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