Development of a high-titer retrovirus producer cell line capable of gene transfer into rhesus monkey hematopoietic stem cells.
Retroviral-mediated gene transfer into primitive hematopoietic cells has been difficult to achieve in large-animal models. We have developed an amphotropic producer clone that generates greater than 10(10) recombinant retroviral particles (colony-forming units) per ml of culture medium. Autologous rhesus monkey bone-marrow cells were cocultured with either high (2 x 10(10) colony-forming units/ml) or low (5 x 10(6) colony-forming units/ml) titer producer clones for 4-6 days and reinfused into sublethally irradiated animals. The proviral genome was detected in blood and bone-marrow cells from all three animals reconstituted with cells cocultured with the high-titer producer cells. In contrast, three animals reconstituted with bone marrow cocultured with the low-titer producer clone exhibited no evidence of gene transfer.
Bodine
D M
DM
Clinical Hematology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892.
McDonagh
K T
KT
Brandt
S J
SJ
Ney
P A
PA
Agricola
B
B
Byrne
E
E
Nienhuis
A W
AW
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002478:Cells, Cultured; D046508:Culture Techniques / Q000379:methods; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification; D006367:HeLa Cells / Q000166:cytology; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D008253:Macaca mulatta; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D012190:Retroviridae / Q000235:genetics* / Q000254:growth & development; D014162:Transfection
D004279:DNA, Viral; D015345:Oligonucleotide Probes; D012367:RNA, Viral