Production of transgenic rats by lentiviral transduction of male germ-line stem cells

Franklin K Hamra, Joel Gatlin, Karen M. Chapman, Dana M. Grellhesl, J. Victor Garcia, Robert E Hammer, David L. Garbers

Research output: Contribution to journalArticlepeer-review

198 Scopus citations

Abstract

Primary cultures of rat spermatogenic cells that did not bind to collagen matrices were able to colonize and form mature spermatozoa when transferred to testes of recipient males. Up to 73% of the progeny from matings with recipient males were derived from the transferred spermatogenic cells. Subsequently, two populations of germ cells were obtained by selection on laminin matrices. Both populations expressed the spermatogenic cell marker, DAZL, but not the somatic cell marker, vimentin, The cells that bound to laminin represented ≈5% of the total population and were greatly enriched in ability to colonize a recipient testis, suggesting an enrichment in germ-line stem cells. The colonization potential was maintained for at least 7 days in culture. These cells were subsequently transduced with a lentiviral enhanced GFP reporter vector and then transferred to WT recipient males. After mating, 26 of 44 pups were derived from the cultured donor germ cells, and 13 pups carried the lentiviral transgene. Based on Southern analysis, the transgene was integrated at a different genetic locus in each animal and was transmitted to ≈50% of pups in the F2 generation. Thus, by using these procedures, ≈30% of pups in the F1 generation inherited and stably transmitted a lentiviral transgene that integrated at various genomic sites.

Original languageEnglish (US)
Pages (from-to)14931-14936
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume99
Issue number23
DOIs
StatePublished - Nov 12 2002

ASJC Scopus subject areas

  • General

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