Abstract
The recent development of high-throughput technologies based on RNA sequencing has allowed a better description of the role of post-transcriptional regulation in gene expression. In particular, the development of degradome approaches based on the capture of 5′monophosphate decay intermediates allows the discovery of a new decay pathway called co-translational mRNA decay. Thanks to these approaches, ribosome dynamics could now be revealed by analysis of 5′P reads accumulation. However, library preparation could be difficult to set-up for non-specialists. Here, we present a fast and efficient 5′P degradome library preparation for Arabidopsis samples. Our protocol was designed without commercial kit and gel purification and can be easily done in one working day. We demonstrated the robustness and the reproducibility of our protocol. Finally, we present the bioinformatic reads-outs necessary to assess library quality control.
Highlights
Turnover of messenger RNAs is a crucial and dynamic mean to control and alter gene expression to answer developmental and environmental cues in eukaryotes
The development of RNA degradome approaches extends our understanding of the role of messenger RNAs (mRNAs) turnover in proper gene expression
Different degradome approaches were developed but all of them are based on the capture of 5 monophosphate decay intermediates
Summary
Turnover of messenger RNAs (mRNAs) is a crucial and dynamic mean to control and alter gene expression to answer developmental and environmental cues in eukaryotes. Degradome approach and polysome RNA sequencing, the authors demonstrated that degradome data can be used to assess translation efficiency [8]. We propose a fast and efficient 5 P degradome library preparation thought the improvement of GMUCT approach [7].
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