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. 2010 Aug 9;5(8):e12088. doi: 10.1371/journal.pone.0012088

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Dumitriu et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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(A) Northern blot of RNA from liver at embryonic day 14.5 (E14.5), and marrow and spleen at 6, 10, and 20 weeks. RNA was detected with Sox6 coding-exon-2 and beta-globin (Hbb-b1) probes. 18S rRNA stained with ethidium bromide is shown as loading control. The decrease in Sox6 and Hbb-b1 RNA level in 10-week marrow probably reflects sample variation. (B) Northern blot of RNA from marrow and spleen of 10-week-old Sox6^fl/fl (C) and Sox6^fl/flErGFPCre (M) mice. The Sox6 probe was the coding exon 2, which is deleted in mutants. (C) Blood parameters and spleen index (percentage of spleen/body weight) in similar mice as in B. Averages with standard deviation are shown for 6 mice. Red, abnormal parameters. WBC, white blood cells; MCHC, mean cell hemoglobin concentration; RDW, red cell distribution width. (D) Erythroid colony formation assay using Sox6^fl/fl (C) and Sox6^fl/flErGFPCre (M) adult spleen. CFU-Es were counted after 3 days and mature BFU-Es after 3, 4, and 7 days. Columns, average with standard deviation obtained for 3 mice. **p<0.01. (E) FACS using CD71 and TER119. Left, representative profiles. CD71⁻/TER119⁻, non- and early erythroid cells; CD71⁺/TER119⁻, late proerythroblasts; CD71⁺/TER119⁺, erythroblasts; CD71⁻/TER119⁺, RBCs. The percentage of cells in each population is indicated. Right, ratios of erythroblasts (EB)/late proerythroblasts. The data in panels (E–H) are averages with standard deviation for 3 mice. **p<0.01. (F) FACS using CD44 and TER119. Left, representative profiles. Right, ratios of proerythroblasts (CD44^high/TER119⁻)/non-erythroid (NE) and erythroid precursors (EP; CD44^−/med/TER119⁻); and erythroblasts (CD44^high/TER119⁻)/non-erythroid and erythroid precursors. *p<0.05, **p<0.01. (G) FACS of erythroblasts (TER119⁺/CD71⁺). Left, representative TO/FSC profiles. Right, ratios of FSC^high/TO^high uncondensed versus FSC^low/TO^low condensed erythroblasts. **p<0.01. (H) FACS of blood. Left, representative TO/CD71 profiles. Right, percentages of atypical reticulocytes (CD71⁺/TO⁻). **p<0.01. (I) Decay profiles of biotin-labeled RBCs. Each value is average with standard deviation for 6 mice. Arrows, RBC half-lives.