Efficient generation of hybrid human-mouse hemoglobin in mice expressing the μLCR-Aγ transgene.

(A) Isoelectric focusing of hemolysates from E15 embryos revealed hybrid human-mouse hemoglobins. Presence (+) or absence (−) of the μLCR-Aγ transgene and the EKLF genotype (± or −/− ) is indicated above each lane. Six hemoglobin (Hb) bands were identifiable, labeled 1 to 6, according to migration from anode to cathode. Hemoglobins 1 and 3 were detectable only in mice that harbored the μLCR-Aγ transgene. They were equally prevalent in EKLF^−/− (lane 4) and^± embryos (lanes 1 and 3). Bands 4 and 5 represent murine β-major and β-minor hemoglobin, respectively; each was markedly and selectively reduced in EKLF^−/− blood. The direction of the anode and cathode is indicated. (B, C) Electrospray mass spectroscopy on gel-purified bands 1 and 3. Hemoglobin band 1 (B) contained 2 proteins of 16 009 and 16 146 kd, which correspond to the predicted molecular weights of human ^Aγ-globin and murine ζ-globin, respectively. Hemoglobin band 3 (C) contained proteins whose molecular masses were consistent with murine α globins, α 1 (Mr 14 981.0), α 5 (Mr 14 995.0), and human Aγ globin (Mr 16 009.3). (D) Reverse-phase HPLC separation of globins expressed by animal 3 in A (genotype EKLF^+/+ Aγ⁺). Peaks annotated with dots represent artefacts of sample storage (single dot, mixed disulfides of murine β-major and β-minor with either cysteine of glutathione; double dot, disulfide-linked murine β-globin dimers).