Retrovirus-mediated Cited2 expression rescues the defective hematopoietic colony-forming activity in Lin⁻ c-Kit⁺ Cited2^−/− fetal liver cells. MSCV-Cited2-IRES-GFP plasmid– or MSCV-IRES-GFP control plasmid–mediated retrovirus transduction was performed on Cited2^−/− and Cited2^+/+ 13.5 dpc fetal liver cells. Briefly, after coculture with retrovirus producer cells, GFP⁺Lin⁻c-Kit⁺ fetal liver cells were sorted for GFP expression followed by analysis of Cited2 mRNA expression via real-time PCR. Meanwhile, 500 cells of the analyzed cell population were plated in triplicate in methylcellulose-based medium (M3434; StemCell Technologies) for colony-forming unit (CFU) assay. Colonies were counted 12 days after plating. (A) Cited2 was expressed in GFP⁺Lin⁻c-Kit⁺ Cited2^−/− fetal liver after transduction with Cited2-expressing retrovirus. (B) Retrovirus-mediated Cited2 expression in GFP⁺Lin⁻c-Kit⁺ Cited2^−/− fetal liver cells at 13.5 dpc significantly increased the frequency of BFU-Es, CFU-GMs, and CFU-GEMMs (n = 3) compared with the vector control (n = 3; *P < .01; average ± SD). (Cited2^+/+ fetal liver cells transduced with control virus [■]; Cited2^−/− fetal liver cells transduced with control virus [■]; Cited2^−/− fetal liver cells transduced with Cited2-expressing virus [□].)