Genetic Manipulation of Hematopoietic Stem Cells 
to determine if they are novel receptors involved 
in the regulation of hematopoiesis. 
To analyze better the behavior of stem cells and 
their progeny in culture and after transplant, we 
are testing a new method for introducing unique 
genetic identifiers into individual stem cells. 
This method allows identification of the stem 
cells by the polymerase chain reaction (PGR), 
which is especially suited for analysis of very 
small numbers of cells. A family of vectors carry- 
ing the bacterial gene neo have been constructed 
that are identical except for small variations in 
size. This size variation is conveniently distin- 
guished by PGR. When mixed together these vec- 
tors provide an array of potential markers that can 
integrate into each stem cell. Because each stem 
cell receives one to four vectors, the integration 
of a particular subset of the vectors provides a 
genetic fingerprint unique to each stem cell fam- 
ily. Preliminary results with these vectors indi- 
cate that we can mark the most primitive hemato- 
poietic precursors very efficiently. We plan to 
use this method to compare different culture 
conditions for stem cells. 
Retroviral vector genetic marking may also 
prove to be very informative in clinical bone 
marrow transplantation. GoUaborative studies 
with Albert Deisseroth (M.D. Anderson Hospital, 
Houston) investigated whether the techniques 
that have been used to optimize mouse bone 
marrow stem cell gene transfer could be used for 
human cells. These experiments demonstrate the 
importance of supporting cells in the culture 
(called stromal cells) for efficient gene transfer. 
The work was performed using an assay of human 
stem cells that depends on their persistence in 
culture. We hope that many valuable lessons will 
be learned from these studies that will contribute 
to the long-term goal of treating disease by the 
use of therapeutic vectors. 
Human bone marrow colony called an erythroid burst or BFU-E. The colony grows from a 
single cell to form 3-9 clusters of red blood cells. Measurement of gene transfer into such 
colonies has helped to improve gene therapy protocols. 
Research and photograph by Kateri Moore in the laboratory offohn Belmont. 
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