Selective footprinting of 40S and 80S ribosome subpopulations (Sel-TCP-seq) to study translation and its control

Result code in IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388971%3A_____%2F22%3A00562220" target="_blank" >RIV/61388971:_____/22:00562220 - isvavai.cz</a>

Result on the web

<a href="https://www.nature.com/articles/s41596-022-00708-4" target="_blank" >https://www.nature.com/articles/s41596-022-00708-4</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1038/s41596-022-00708-4" target="_blank" >10.1038/s41596-022-00708-4</a>

Result language

angličtina

Original language name

Selective footprinting of 40S and 80S ribosome subpopulations (Sel-TCP-seq) to study translation and its control

Original language description

This extension of the TCP-seq protocol describes procedures for selective profiling of 40S and 80S ribosome subpopulations bound by a factor of interest, permitting detailed studies of the different stages of translation in mammalian and yeast cells.nMultiple aspects of mRNA translation are subject to regulation. Here we present a ribosome footprinting protocol to determine the location and composition of 40S and 80S ribosome complexes on endogenous mRNAs transcriptome-wide in vivo in yeast and mammalian cells. We present an extension of the translation complex profiling (TCP-seq) protocol, originally developed in yeast, by including an immunoprecipitation step to assay the location of both 40S and 80S ribosome complexes containing proteins of interest. This yields information on where along mRNAs the ribosome-bound protein of interest joins the ribosome to act, and where it leaves again, thereby monitoring the sequential steps of translation and the roles of various translation factors therein. Rapid fixation of live cells ensures the integrity of all translation complexes bound to mRNA at native positions. Two procedures are described, differing mainly in the fixation conditions and the library preparation. Depending on the research question, either procedure offers advantages. Execution of a Sel-TCP-seq experiment takes 5-10 working days, and initial data analysis can be completed within 2 days.

Czech name

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Czech description

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Type

J_imp - Article in a specialist periodical, which is included in the Web of Science database

CEP classification

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OECD FORD branch

10606 - Microbiology

Project

Result was created during the realization of more than one project. More information in the Projects tab.

Continuities

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Publication year

2022

Confidentiality

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Name of the periodical

Nature Protocols

ISSN

1754-2189

e-ISSN

1750-2799

Volume of the periodical

17

Issue of the periodical within the volume

10

Country of publishing house

GB - UNITED KINGDOM

Number of pages

49

Pages from-to

2139-2187

UT code for WoS article

000828917600001

EID of the result in the Scopus database

2-s2.0-85134645475