Figure 2
Figure 2. Transgenic hϵ-globin mRNA accumulates to high levels in vivo in intact mouse erythroid progenitors. (A) Structures of transgenes encoding hϵ- and hβ-globin mRNAs. μβLCR-hϵ and μβLCR-hβ contain the full-length transcribed regions of the hϵ-globin gene (gray) and hβ-globin gene (black), each identically flanked by DNA containing the hβ-globin gene promoter and 3′-flanking region/enhancer. Both constructs are linked to a micro–β-locus control region (μβLCR).22 (B) Analysis of hϵ-globin mRNA stability in a representative mouse. Total mRNA was recovered from the bone marrow (B) and peripheral reticulocytes (R) of a representative hϵ mouse and was subsequently subjected to RPA using [32P]-labeled hϵ-globin and internal control mα-globin probes. The specificities of the mα-globin and hϵ-globin antisense mRNA probes were demonstrated by parallel assay of reticulocyte RNA from a nontransgenic control mouse. (C) Transgenic hϵ-globin mRNA is highly stable in terminally differentiating mouse erythroid cells. The average stability of hϵ-globin mRNA in animals from each of 4 independent transgenic lines was determined as described in panel B. The average across all 4 lines (0.87 ± 0.52) is indicated by a gray line. Error bars represent 1 SD. The stability of hϵ-globin mRNA is defined as (hϵ/mα)P/(hϵ/mα)B, where P and B indicate peripheral blood and bone marrow, respectively. The previously reported average stability of hβ-globin mRNA derived from 5 transgenic lines (▨; 1.47 ± 0.49) is summarized for comparison.21

Transgenic hϵ-globin mRNA accumulates to high levels in vivo in intact mouse erythroid progenitors. (A) Structures of transgenes encoding hϵ- and hβ-globin mRNAs. μβLCR-hϵ and μβLCR-hβ contain the full-length transcribed regions of the hϵ-globin gene (gray) and hβ-globin gene (black), each identically flanked by DNA containing the hβ-globin gene promoter and 3′-flanking region/enhancer. Both constructs are linked to a micro–β-locus control region (μβLCR).22  (B) Analysis of hϵ-globin mRNA stability in a representative mouse. Total mRNA was recovered from the bone marrow (B) and peripheral reticulocytes (R) of a representative hϵ mouse and was subsequently subjected to RPA using [32P]-labeled hϵ-globin and internal control mα-globin probes. The specificities of the mα-globin and hϵ-globin antisense mRNA probes were demonstrated by parallel assay of reticulocyte RNA from a nontransgenic control mouse. (C) Transgenic hϵ-globin mRNA is highly stable in terminally differentiating mouse erythroid cells. The average stability of hϵ-globin mRNA in animals from each of 4 independent transgenic lines was determined as described in panel B. The average across all 4 lines (0.87 ± 0.52) is indicated by a gray line. Error bars represent 1 SD. The stability of hϵ-globin mRNA is defined as (hϵ/mα)P/(hϵ/mα)B, where P and B indicate peripheral blood and bone marrow, respectively. The previously reported average stability of hβ-globin mRNA derived from 5 transgenic lines (▨; 1.47 ± 0.49) is summarized for comparison.21 

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