Fig. 5.
Fig. 5. Mice transplanted with either −/− control cells or a low frequency of −/− rFancc cells had progenitors that were resistant to multiple inhibitory cytokines. / (A) Semiquantitative PCR. HEL cells with a single provirus copy were used as the standard DNA for dilution, and untransduced HEL cells were used as the negative control. DNA extracted from the peripheral blood of mouse 6 was amplified with primers specific for proviral DNA. Approximately 1%-2% of peripheral blood cells from mouse 6 contained proviral DNA. (B) Inhibitory cytokine progenitor assay. Sorted CD45.2+ cells from mice 3, 5, and 6 were cultured in methylcellulose progenitor assays at 5 × 104 cells/mL with either IFN-γ (10 ng/mL), TNF-α (10 ng/mL), or MIP-1α (50 ng/mL). Each condition was plated in triplicate and scored on day 7 of culture. The percent maximal colony formation was determined by dividing the number of colonies scored with each inhibitory cytokine by untreated control cultures. Low-density cells fromFancc−/− and WT mice were used as controls. Baseline progenitor numbers scored in the untreated groups were 84 forFancc−/− , 82 forFancc+/+, 195 for mouse 3, 106 for mouse 5, and 89 for mouse 6. Error bars represent SEM.

Mice transplanted with either −/− control cells or a low frequency of −/− rFancc cells had progenitors that were resistant to multiple inhibitory cytokines.

(A) Semiquantitative PCR. HEL cells with a single provirus copy were used as the standard DNA for dilution, and untransduced HEL cells were used as the negative control. DNA extracted from the peripheral blood of mouse 6 was amplified with primers specific for proviral DNA. Approximately 1%-2% of peripheral blood cells from mouse 6 contained proviral DNA. (B) Inhibitory cytokine progenitor assay. Sorted CD45.2+ cells from mice 3, 5, and 6 were cultured in methylcellulose progenitor assays at 5 × 104 cells/mL with either IFN-γ (10 ng/mL), TNF-α (10 ng/mL), or MIP-1α (50 ng/mL). Each condition was plated in triplicate and scored on day 7 of culture. The percent maximal colony formation was determined by dividing the number of colonies scored with each inhibitory cytokine by untreated control cultures. Low-density cells fromFancc−/− and WT mice were used as controls. Baseline progenitor numbers scored in the untreated groups were 84 forFancc−/− , 82 forFancc+/+, 195 for mouse 3, 106 for mouse 5, and 89 for mouse 6. Error bars represent SEM.

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