Protocol

Preparation of Cell-Free Splicing Extracts from Saccharomyces cerevisiae

  1. Manuel Ares Jr.
Adapted from RNA: A Laboratory Manual, by Donald C. Rio, Manuel Ares Jr, Gregory J. Hannon, and Timothy W. Nilsen. CSHL Press, Cold Spring Harbor, NY, USA, 2011.

Abstract

Much of our understanding of the mechanism of splicing comes from the analysis of cell extracts able to carry out splicing complex formation and splicing reactions in vitro using exogenously added synthetic model pre-mRNA transcripts. This protocol describes the preparation of whole-cell extracts from the budding yeast Saccharomyces cerevisiae. These extracts can be used to dissect the biochemical steps of the splicing reaction and to determine the macromolecules, cofactors, and substrate features necessary for successful splicing.

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  1. doi:10.1101/pdb.prot078113 Cold Spring Harb Protoc 2013: pdb.prot078113- © 2013 Cold Spring Harbor Laboratory Press
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