Short interfering (si) RNAs have now been shown to inhibit gene expression in several species, including mammals (Elbashir et al.: Nature 411: 494–498, 2001; Fire et al.: Nature 391: 806–811, 1998). RNA inhibition in primary cells such as stem cells would facilitate rapid gene discovery in a postgenome era. While retroviruses can deliver siRNA expression cassettes for stable expression (Barton and Medzhitov: Proc Natl Acad Sci USA 99: 14943–14945, 2002; Paddison et al.: Proc Natl Acad Sci USA 99: 1443–1448, 2002; Rubinson et al.: Nat Genet 33: 401–406, 2003), an efficient method for direct transfer of siRNA to stem cells is still lacking. Here, we established electroporation to deliver siRNA to hematopoietic progenitors. On average, at least 80% of cells take up the RNA, and these display nearly 100% knockout of marker gene expression at both the RNA and protein level. Moreover, knockdown of the hematopoietic regulator, CD45, results in 3-fold more hematopoietic colonies in a progenitor assay. These results demonstrate that transient transfection of siRNA to primary cells can have substantial functional consequences. This technology may be applicable to a variety of primary cell types. genesis 36: 203–208, 2003.© 2003 Wiley-Liss, Inc.