Protocol

Human Pluripotent Stem Cells

Volume 767 of the series Methods in Molecular Biology pp 333-341

Date:

Nucleofection of Human Embryonic Stem Cells

  • Helen FongAffiliated withDepartment of Biological Chemistry, Sue and Bill Gross Stem Cell Research Center, School of Medicine, University of California, Irvine
  • , K. A. Hohenstein ElliottAffiliated withPeter Donovan Lab, Human Genetics and Molecular Biology, McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine
  • , Leslie F. LockAffiliated withDepartments of Biological Chemistry and Developmental and Cell Biology, Sue and Bill Gross Stem Cell Research Center, School of Biological Sciences, University of California, Irvine
  • , Peter J. DonovanAffiliated withDepartments of Biological Chemistry and Developmental and Cell Biology, Sue and Bill Gross Stem Cell Research Center, School of Biological Sciences, University of California, Irvine Email author 

* Final gross prices may vary according to local VAT.

Get Access

Abstract

The ability to realize the full potential of human pluripotent stem cells (hPSCs) as tools for ­understanding human development and advancing the field of regenerative medicine is dependent on efficient methods to genetically manipulate these cells. There are several methods for introducing foreign DNA into cells such as electroporation, lipid-based transfection technology, and viral transduction. We describe here a method to transfect human embryonic stem cells (hESCs) using nucleofection technology. This unique method uses the Nucleofector II Device that combines the use of a cell type-specific Nucleofector Solution and preprogrammed electrical parameters to efficiently deliver DNA into the cell nucleus. The use of this technology allows high-efficiency transfer of nucleic acids into hESCs enabling both transient and stable manipulation of gene expression in these cells.

Key words

human embryonic stem cells pluripotent stem cells nucleofection transfection transgene expression RNA interference neurotrophins