Protocol

Purification of Polysomes

  1. James A. Lake3
  1. 1Department of Biology and Center for the Study of Biological Complexity, Virginia Commonwealth University, Richmond, Virginia 23284;
  2. 2Primary Pharmacology Group, Pfizer Global Research and Development, Groton, Connecticut 06340;
  3. 3Department of Molecular, Cell and Developmental Biology and Department of Human Genetics, University of California, Los Angeles, California 90095

    Abstract

    Isolated polysomes (also known as translating ribosomes or polyribosomes) are mRNA–ribosome complexes that are frequently used for the in vitro study of the regulation of protein synthesis. Here we describe a protocol for the isolation of prokaryotic and plant polysomes by sucrose gradient sedimentation. The protocol allows for the separation of multiple ribosomes attached to mRNA from run-off ribosome monomers. In addition to collecting the polysome pellet, if the sucrose gradients used in this protocol are fractionated, it is possible to isolate and quantify the polysomes, the ribosome monomers, and the ribosomal subunits.

    Footnotes

    • ↵4 Correspondence: mcrivera{at}vcu.edu

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    1. doi:10.1101/pdb.prot081364 Cold Spring Harb Protoc 2015: pdb.prot081364- © 2015 Cold Spring Harbor Laboratory Press
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