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Identification of a novel DNA sequence differentially expressed between
normal human CD34+CD38hi and CD34+CD38lo marrow cells
L Graf and B Torok-Storb
Clinical Research Division, Fred Hutchinson Cancer Research Center,
Seattle, WA 98104, USA.
We have applied the recently developed differential display method to
extend the molecular characterization of the less mature CD34+CD38lo bone
marrow progenitors in comparison with the CD34+CD38hi cells to better
understand their functional differences. Immunomagnetic enrichment of CD34+
cells followed by flow cytometry was used to isolate CD34+CD38lo and
CD34+CD38hi cells from human organ donor bone marrow. A limited set of the
poly A+ RNA sequences present in these two cell populations was amplified
by a combination of reverse transcription with an anchored oligo dT-based
primer and polymerase chain reaction with the same oligo dT primer and
arbitrary decamers. A radioactive tracer allowed these sequences to be
displayed as a series of bands on a denaturing polyacrylamide gel. Eight
bands were chosen that appeared in multiple displays to represent gene
sequences differentially expressed between CD34+CD38hi and CD34+CD38lo
cells. Comparison of the sequences with public DNA sequence databases
available identified one sequence as myeloperoxidase. Two other clones
matched sequence fragments of unknown function, whereas the remaining five
are novel sequences not present in existing databases. The relative level
of expression of all of the sequences was tested by an independent reverse
transcriptase-polymerase chain reaction with sequence-specific
oligonucleotide primers. The lower level of myeloperoxidase mRNA in
CD34+CD38lo cells was confirmed, as was the higher expression of the novel
sequence 345. Sequence 345 expression is highest in CD34+CD38- cells and
decreases with increased CD38 expression. It is expressed in negligible
amounts in hematopoietic cell lines and other sources of human tissue,
suggesting it may have a functional role in normal hematopoiesis.
Volume 86,
Issue 2,
pp. 548-556,
07/15/1995
Copyright © 1995 by The American Society of Hematology
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