A. A. Comparison of transient HSAKIN17 gene silencing induced by either siRNA duplexes or EBV-based siRNA vectors. Twenty-four hours after seeding, HeLa cells were transfected with either EBV vectors or siRNA. Three days later, cells were trypsinized, counted, and proteins were analyzed as described. 1, pBD631 (empty vector, reverse); 2, pBD665 (empty vector, direct); 3, pBD650 (control vector, reverse); 4, pBD632 (siK663 vector, reverse); 5, pBD674 (siK180 vector, reverse); 6, pBD676 (siK906 vector, reverse); 7, pBD678 (siK180 vector, direct); 8, pBD680 (siK906 vector, direct); 9, siRNA K180 duplex; 10, siRNA K906 duplex. B. Stability of HSAKIN17 gene silencing 3, 10, and 18 days after transfection of HeLa cells with EBV vectors. 1, pBD631; 2, pBD632; 3, pBD664 (siK663 vector, direct); 4, pBD665; 5, pBD650. C. Enhanced efficiency of gene silencing mediated by EBV-based siRNA vectors in comparison with integrative plasmids. Forty-eight hours after transfection, 150,000 RKO cells were seeded in a 6 cm diameter dish in the presence of hygromycin B in the medium. Immunocytochemical staining of HSAkin17 (immunoglobulin G K36; magnification, ×50). Cells were counterstained with 4′,6-diamidino-2-phenylindole. 1, pBD641 (empty vector, Hygro); 2, pBD642 (siK663 vector, Hygro); 3, pBD631; 4, pBD632. Denis S.F. Biard et al. Mol Cancer Res 2005;3:519-529 ©2005 by American Association for Cancer Research