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a Harvard Stem Cell Institute and Harvard Medical School, Department of Biological Chemistry and Molecular Pharmacology and Division of Hematology/Oncology, Children’s Hospital, Boston, Massachusetts, USA;
b Washington University School of Medicine, St. Louis, Missouri, USA;
c Technion, Rambam Medical Center, Haifa, Israel
Key Words. Human embryonic stem cells • RNA interference • Transgene expression • Retroviral and lentiviral vectors • Oct4 • Nanog • Self-renewal
Correspondence: George Q. Daley, M.D., Ph.D., Harvard Stem Cell Institute and Harvard Medical School, Department of Biological Chemistry and Molecular Pharmacology and Division of Hematology/Oncology, Children’s Hospital, Boston, Massachusetts 02115, USA. Telephone: 617-919-2013; Fax: 617-730-0222; e-mail: george.daley{at}childrens.harvard.edu
RNA interference methodology suppresses gene expression, thus mimicking loss-of-function mutation and enabling in vitro and in vivo gene function analysis. In this study, we used retroviral and lentiviral vectors to deliver small interfering RNAs and report high-efficiency silencing of a green fluorescent protein (GFP) trans gene and the stem cell–specific transcription factors Oct4/POU5F1 and Nanog in human embryonic stem cells. Gene knockdown of Oct4 and Nanog promotes differentiation, thereby demonstrating a role for these factors in human embryonic stem cell self-renewal.
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