L. Graf et B. Torokstorb, IDENTIFICATION OF A NOVEL DNA-SEQUENCE DIFFERENTIALLY EXPRESSED BETWEEN NORMAL HUMAN CD34(-CELLS()CD38(HI) AND CD34(+)CD38(LO) MARROW), Blood, 86(2), 1995, pp. 548-556
We have applied the recently developed differential display method to
extend the molecular characterization of the less mature CD34(+)CD38(l
o) bone marrow progenitors in comparison with the CD34(+)CD38(hi) cell
s to better understand their functional differences. Immunomagnetic en
richment of CD34(+) cells followed by flow cytometry was used to isola
te CD34(+)CD38(lo) and CD34(+)CD38(hi) cells from human organ donor bo
ne marrow, A limited set of the poly A+ RNA sequences present in these
two cell populations was amplified by a combination of reverse transc
ription with an anchored oligo dT-based primer and polymerase chain re
action with the same oligo dT primer and arbitrary decamers. A radioac
tive tracer allowed these sequences to be displayed as a series of ban
ds on a denaturing polyacrylamide gel, Eight bands were chosen that ap
peared in multiple displays to represent gene sequences differentially
expressed between CD34(+)CD38(hi) and CD34(+)CD38(lo) cells. Comparis
on of the sequences with public DNA sequence databases available ident
ified one sequence as myeloperoxidase. Two other clones matched sequen
ce fragments of unknown function, whereas the remaining five are novel
sequences not present in existing databases, The relative level of ex
pression of all of the sequences was tested by an independent reverse
transcriptase-polymerase chain reaction with sequence-specific oligonu
cleotide primers. The lower level of myeloperoxidase mRNA in CD34(+)CD
38(lo) cells was confirmed, as was the higher expression of the novel
sequence 345. Sequence 345 expression is highest in CD34(+)CD38(-) cel
ls and decreases with increased CD38 expression, It is expressed in ne
gligible amounts in hematopoietic cell lines and other sources of huma
n tissue, suggesting it may have a functional role in normal hematopoi
esis. (C) 1995 by The American Society of Hematology.