Hematopoietic cells enriched for stem cells are tolerant of Cfp1 depletion.

<p>Mutant and control mice were induced with poly(I:C) injections at days 0, 2, and 4. Bone marrow cells were collected at day 8 following initiation of Cre induction and analyzed by flow cytometry for the indicated cell surface markers. (A) Scatter plot showing a representative experiment. (B–C) Summary for enumeration of LSK (Lin- Sca1+ Kit+), LT-HSC (Lin- Sca1+ Kit+ CD34- Flt3-), ST-HSC (Lin- Sca1+ Kit+ CD34+ Flt3+), and more primitive ST-HSC (Lin- Sca1+ Kit+ CD34+ Flt3-). (A) and (B) present the data in terms of percent of total LDBMCs, while (C) presents cell number per femur. (For B–C, N = 11 for Cre- controls and N = 16 for Cre+ mutants for each of three independent experiments.) (D) Total RNA was isolated from purified LSK cells derived from the bone marrow of mice of the indicated genotypes (day 8 after poly(I:C) injections on days 0, 2, and 4), and semi-quantitative RT-PCR was performed to determine the level of <i>Cxxc1</i> transcript. RNA from wild type or <i>Cxxc1-null</i> ES cells was used as positive and negative controls, respectively <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113745#pone.0113745-Carlone1" target="_blank">[9]</a>. The conditions for all PCR reactions were confirmed to be within the linear range (data not shown). (E) Mutant and control mice were induced with poly(I:C) injections on days 0, 2, and 4, and BrdU was injected on day 7. On day 8 following initiation of Cre induction, bone marrow was isolated and analyzed for the frequency of BrdU incorporation into LSK cells and CD34/Flt3 subpopulations. Asterisks denote statistically significant differences (P≤0.02). (F) LSK cells were recovered from the bone marrow of mutant or control mice 8 days following Cre induction and examined for apoptosis by Annexin V and PI staining.