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Efficient Gene Knockdowns in Human Embryonic Stem Cells Using Lentiviral-Based RNAi

  • Asmin Tulpule
  • George Q. Daley
Part of the Methods in Molecular Biology book series (MIMB, volume 482)

Abstract

Human embryonic stem cells (hESCs) represent a powerful platform to study human development and its dysfunction in human disease. However, certain biological properties have hampered the application of standard gain of function and loss of function tools to these cells. For example, while traditional gene knockouts by homologous recombination (HR) have been reported, the low cloning efficiency of hESCs has made HR a lengthy and laborious undertaking. An alternative method of achieving loss of function is the use of small interfering RNAs (siRNAs) that can be introduced either as pre-synthesized duplexed oligonucleotides or via lentiviral vector. The use of a lentiviral vector to deliver siRNAs has proven to be a rapid and specific way to achieve highly efficient and persistent gene knockdowns in hESCs. In this chapter, we will summarize the key requirements for the successful application of lentiviral RNAi in hESCs.

Key words

RNAi RNA-interference shRNA siRNA human embryonic stem cell hESC lentivirus 

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Asmin Tulpule
    • 1
  • George Q. Daley
    • 1
  1. 1.Harvard Medical School, Karp Family Research Building, Division of Hematology/Oncology, Children’s Hospital BostonBostonUSA

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