JARID2 depletion enhances human HSC expansion in vitro and in vivo. (A) RT-qPCR analysis of JARID2 expression in human hematopoietic progenitor cells. Results are normalized to housekeeping gene HPRT1. Data represent mean ± SD; n = 3. (B) JARID2 knockdown confirmed by RT-qPCR and western blot after 3 days of in vitro culture. Results for qPCR are normalized to housekeeping gene HPRT1. Data represent mean ± SD; n = 5 to 9. (C) Dose-dependent enrichment of CD34⁺ cells after 3 weeks in culture within the JARID2-depleted GFP⁺ population compared with GFP^– and shRNA-Scramble controls. Data represent mean + SD; n = 3. (D) Relative enrichment of CD34⁺ cells from fetal liver, cord blood, and adult BM after 14 and 21 days in culture. Data represent mean + SD; n = 3. *P < .05, **P < .01, ***P < .001, and ****P < .0001 were calculated by using a two-way ANOVA with multiple comparisons in GraphPad Prism5. (E-F) Graphs showing the percentage of human cell chimerism (huCD45⁺) in the peripheral blood and BM of mice transplanted with shRNA-JARID2.1, shRNA-JARID2.2, or shRNA-Scramble transduced GFP⁺ cells in (E) primary and (F) secondary recipients. Primary animals used for secondary transplant have square data points.