RNAi screen in human CB CD34 cells. (A) Overall design of the screening strategy. Large numbers of primary CB CD34⁺ cells are infected with the lentiviral short hairpin RNA (shRNA) library and subsequently passaged in long-term cultures (10 weeks) followed by colony-forming cell (CFC) assays to positively select for clones that have acquired enhanced self-renewal/proliferation ability. Potential hits are identified by sequence analysis of proviral inserts from the selected cells. (B) CFC levels after 10 weeks culture. Twelve pools of library-transduced cells were independently assayed. Control pools were transduced with shRNA against GFP in 2 pools, and 1 pool was left untransduced (mock). Pool 7 showed high levels of BFU-E growth indicated by *. (C) Distribution of proviral shRNAs among the screening pools that showed increased CFC levels. The graph shows relative abundance of shRNAs in each screening pool as an overlay on the colony numbers shown in panel B.