Gene silencing using RNA interference in embryonic stem cells.

Published

Journal Article

Pluripotent embryonic stem (ES) cells are an important model system to examine gene expression and lineage segregation during differentiation. One powerful approach to target and inhibit gene expression, RNAi, has been applied to ES cells with the goal of teasing out the cascades of gene expression/repression that shape the early embryo. In this chapter, we describe the current understanding of the mechanisms of gene silencing by small hairpin RNAs, as well as controls and caveats to using this approach in ES cells. A consideration of synthetic vs plasmid-based RNAi vectors, design of targeting constructs, transfection of ES cells, and flow sorting of targeted cells is followed by methods for the analysis of phenotype and behavior of targeted cell populations using immunohistochemistry, reverse transcriptase polymerase chain reaction, Western blotting, and scanning electron microscopy.

Full Text

Duke Authors

Cited Authors

  • Velkey, JM; Slawny, NA; Gratsch, TE; O'Shea, KS

Published Date

  • 2006

Published In

Volume / Issue

  • 329 /

Start / End Page

  • 233 - 261

PubMed ID

  • 16845995

Pubmed Central ID

  • 16845995

International Standard Serial Number (ISSN)

  • 1064-3745

Digital Object Identifier (DOI)

  • 10.1385/1-59745-037-5:233

Language

  • eng

Conference Location

  • United States