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Blood, 15 June 2004, Vol. 103, No. 12, pp. 4511-4513.
Prepublished online as a Blood First Edition Paper on February 26, 2004; DOI 10.1182/blood-2003-07-2397.
Submitted July 16, 2003
Accepted February 16, 2004
Gene silencing by lentivirus-mediated delivery of siRNA in human CD34+ cells
Tibor Schomber, Christian P Kalberer, Aleksandra Wodnar-Filipowicz, and Radek C Skoda*
Department of Research, Experimental Hematology, Basel University Hospital, Basel, Switzerland
* Corresponding author; email: radek.skoda{at}unibas.ch.
To derive an efficient system for gene silencing in human hematopoietic stem cells we modified a lentiviral vector for small interfering RNA (siRNA) delivery. For this purpose, an H1-promotor driven siRNA expression cassette was introduced into a lentiviral vector and the p53 mRNA was chosen as a target for siRNA mediated gene silencing. Using the recombinant lentivirus we infected human cord blood derived CD34+ cells and obtained a transfection efficiency of up to 50%, as determined by expression of enhanced green fluorescent protein (EGFP). In EGFP positive LTC-IC and CFU-C derived cells we observed a reduction of p53 mRNA of up to 95%. Importantly, this reduction remained stable during several weeks of cell culture. Furthermore, p53 gene silencing resulted in decreased p21 mRNA levels and reduced the sensitivity of CD34+ cells towards the cytotoxic drug etoposide. Thus, lentiviral delivery of siRNA can allow for efficient and stable gene silencing in human HSC and will be very valuable for further gene function studies.
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