Assessing messenger RNA decapping in cellular extracts.

Naomi Bergman, Joseph Milone, Elizabeth J. Bates, Mateusz Opyrchal, Vivian Bellofatto, Jeffrey Wilusz

Research output: Contribution to journalArticle

2 Scopus citations

Abstract

Removal of the 5' cap from a messenger RNA (mRNA) is an integral part of all mRNA decay pathways and can be a highly regulated event. Assays designed to assess decapping in vitro need to effectively resolve four products of mRNA decay: 7meGpppG produced by 3'-5' shortening of the transcript by the exosome, 7meGMP produced by the scavenger decapping enzyme DcpS acting on the product of exosomal decay, 7meGDP produced by the Dcp1/2 decapping enzyme, and free phosphate, which can be generated by phosphatases in the extract acting upon either of the two products of decapping noted above. We have outlined both thin-layer chromatography and acrylamide-gel based approaches that can be used to assess decapping activities.

Original languageEnglish
Pages (from-to)181-192
Number of pages12
JournalMethods in molecular biology (Clifton, N.J.)
Volume257
DOIs
StatePublished - 2004
Externally publishedYes

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