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. 1991 Nov 1;88(21):9760–9764. doi: 10.1073/pnas.88.21.9760

Expression of a swine class II gene in murine bone marrow hematopoietic cells by retroviral-mediated gene transfer.

G E Shafer 1, D W Emery 1, K Gustafsson 1, S Germana 1, W F Anderson 1, D H Sachs 1, C LeGuern 1
PMCID: PMC52800  PMID: 1946400

Abstract

As a first step in assessing the efficacy of a gene transfer approach to the induction of transplantation tolerance in our miniature swine model, double-copy retroviral vectors engineered to express a drug-resistance marker (neomycin) and a swine class II DRB cDNA were constructed. Infectious particles containing these vectors were produced at a titer of greater than 1 x 10(6) G418-resistant colony-forming units/ml using both ecotropic and amphotropic packaging cell lines. Flow cytometric analysis of DRA-transfected murine fibroblasts subsequently transduced with virus-containing supernatants demonstrated that the transferred sequences were sufficient to produce DR surface expression. Cocultivation of murine bone marrow with high-titer producer lines leads to the transduction of 40% of granulocyte/macrophage colony-forming units (CFU-GM) as determined by the frequency of colony formation under G418 selection. After nearly 5 weeks in long-term bone marrow culture, virus-exposed marrow still contained G418-resistant CFU-GM at a frequency of 25%. In addition, virtually all of the transduced and selected colonies contained DRB-specific transcripts. These results suggest that a significant proportion of very primitive myelopoietic precursor cells can be transduced with the DRB recombinant vector and that vector sequences are expressed in the differentiated progeny of these cells.

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