The empty vector control cell line had no effect on the luciferase expression, either transfected with the sensor construct or the mutated construct (C, second panel) Supporting Information 4: B and T cell development of miR-221-expressing

preB-I cells in vitro. Representative cell lines of Pax5-/- (A), wildtype preB-I (B) and miR-221 (C) transduced cell lines were cultured under conditions that allow T-lineage cell development in vitro. Flow cytometry profiles are shown for CD44/CD25 and CD4/CD8 of each cell line. Pax5-/- cells develop into T-lineage cells within 23 days. PreB-I cells transduced with miR-221 or -222 did not develop into T-lineage cells in vitro but remained CD19+ B cells (see Supporting Information 2B) Supporting Information 5: Phenotype of CD45.1+ donor-derived selleck compound cells in the CD45.2+ hosts. Flow cytometric analysis of the phenotypes of the CD45.1+ cells in BM (A), spleen (B) and the peritoneal cavity (C). FACS plots of one representative mouse in the presence of doxycycline are shown. The numbers in the flow cytometry profiles indicate the respective gate percentages. Supporting Information 6: Ex vivo maturation of transplanted cells. CD45.1+GFP+ BM cells and CD45.1+GFP- spleen cells from CD45.2 mice transplanted

with CD45.1+rtTA+tetO-miR-221+preB-I cells into mice, either fed for 4 weeks with doxycycline, or kept without, were cultured for 3 days in the presence of αCD40, PLX3397 IL4 and IL5 and 1 μg doxycycline/ml. On

day 3, the cells were this website harvested and stained for CD19, IgM and MHC-II on their surface and compared to wild type cells sorted from the BM as CD19+ IgM- from wild type C57BL/6 mice. Supporting Information 7: Termination of doxycycline-induced miR-221 expression terminates preB-cell retention in spleen and peritoneal cavity. From mice transplanted with miR-221-expressing cells (A) and subclone 32 (B) in the presence of doxycycline as described in Figure 4, kept for 4 weeks, the doxycycline-containing drinking water was replaced by normal water. Thereafter, mice were analyzed 2 (A, third panel) and 4 weeks later (A, fourth panel, B, third panel). Total cell numbers of CD45.1+ cells in the spleen and peritoneal cavity were calculated by live cell counting and trypan blue exclusion followed by flow cytometry. Each black dot represents one mouse. Each green dot represents one mouse, were CD45.1+GFP+ were detected after the doxycycline was removed for 4 weeks. Horizontal lines represent the median of calculated cells. Dashed lines denote the limits of FACS phenotype detection (see Materials and Methods). Supporting Information 7: Termination of doxycycline-induced miR-221 expression terminates preB-cell retention in spleen and peritoneal cavity. From mice transplanted with miR-221-expressing cells (A) and subclone 32 (B) in the presence of doxycycline as described in Figure 4, kept for 4 weeks, the doxycycline-containing drinking water was replaced by normal water.