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Blood, Vol. 94 No. 9 (November 1), 1999:
pp. 3178-3184
Transgenic Analysis of a 100-kb Human  -Globin Cluster-Containing
DNA Fragment Propagated as a Bacterial Artificial Chromosome
Richard M. Kaufman,
Christine T.N. Pham, and
Timothy J. Ley
From the Division of Bone Marrow Transplantation and Stem Cell
Biology, Departments of Internal Medicine and Genetics, Washington
University School of Medicine, St Louis, MO.
To date, the normal transcriptional regulation of the human
 -globin gene cluster has been recapitulated most accurately in transgenic mice that carry large yeast artificial chromosome (YAC) or
ligated cosmid constructs. However, these large transgenes still
exhibit variegated expression levels, perhaps because they tend to
rearrange upon integration, or because the cloning vectors remain
attached to the globin inserts. To try to circumvent these potential
problems, we investigated the transgenic properties of a 100-kb DNA
fragment containing the entire human -globin cluster propagated in a
bacterial artificial chromosome (BAC). We created 9 independent mouse
lines, each carrying 1 to 6 copies of the human -globin cluster
without the attached BAC vector. Five of the lines carry unrearranged
copies of the cluster. Reverse-transcriptase polymerase chain reaction
(RT-PCR) analysis of adult F1 mice showed that 2 lines
express human globin at levels approximately equivalent to the
endogenous mouse -major genes. One line expresses no human globin, while the remaining 6 lines show intermediate expression levels. Complete   -globin gene switching occurs, but is
slightly delayed with respect to the endogenous mouse
embryonicadult switch. Since these data are similar to what
has been obtained using globin YACs or ligated cosmids, we conclude
that (1) globin transgenes propagated in BACs are no less likely to
rearrange than their cosmid or YAC counterparts, and (2) the retention
of YAC vector sequences in a transgene probably has no significant
impact on globin expression when using constructs of this size.
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