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Prepublished online as a Blood First Edition Paper on August 15, 2002; DOI 10.1182/blood-2002-02-0355.
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Blood, 15 December 2002, Vol. 100, No. 13, pp. 4410-4419
HEMATOPOIESIS
Transgenic targeting with regulatory elements of the human
CD34 gene
Hanna S. Radomska,
David A. Gonzalez,
Yutaka Okuno,
Hiromi Iwasaki,
Andras Nagy,
Koichi Akashi,
Daniel G. Tenen, and
Claudia S. Huettner
From the Harvard Institutes of Medicine, Harvard
Medical School, and the Department of Cancer Immunology and AIDS, Dana
Farber Cancer Institute, Boston, MA; and The Samuel Lunenfeld Research
Institute, Mount Sinai Hospital, Toronto, Canada.
The human CD34 gene is expressed on early progenitor
and stem cells in the bone marrow. Here we report the isolation of the human CD34 locus from a human P1 artificial chromosome (PAC) library and the characterization and evaluation of this genomic fragment for
expression of reporter genes in stable cell lines and transgenic mice.
We show that a 160-kb fragment spanning 110 kb of the 5' flanking
region and 26 kb of the 3' flanking region of the
CD34 gene directs expression of the human CD34
gene in the bone marrow of transgenic mice. The expression of human
CD34 transgenic RNA in tissues was found to be similar to that of the
endogenous murine CD34 gene. Colony-forming cell assays
showed that bone marrow cells staining positive for human CD34 consist
of early progenitor cells in which expression of CD34 decreased with
cell maturation. In order to test the construct for its ability to
express heterologous genes in vivo, we used homologous recombination in
bacteria to insert the tetracycline-responsive transactivator
protein tTA. Analysis of transgenic human CD34-tTA mice by cross
breeding with a strain carrying Cre recombinase under control of a
tetracycline-responsive element demonstrated induction of Cre
expression in mice in a pattern consistent with the expression of the
human CD34 transgene.
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