Production of avian retroviruses and tissue-specific somatic retroviral gene transfer in vivo using the RCAS/TVA system

Spatiotemporal retroviral gene transfer into specific somatic mammalian cells using the avian RCAS(replication-competent avian sarcoma-leukosis virus long terminal repeat with splice acceptor)/tumor virus A(TVA) system is a versatile tool for performing lineage tracing and gene function analysis in vivo. RCASretroviruses carrying the subgroup Aenvelope transduce only genetically engineered mammalian cells that express the cognate avian retroviral receptor TVA. The RCAS/TVAgene delivery system has been successfully used in various different mouse TVA-expression models. This protocol contains a detailed description of the production of high-titer RCASretroviruses in chicken fibroblasts and the transduction of proliferating TVA-positive somatic mammalian cells in vivo. By taking advantage of the combination of the RCAS/TVAwith the ‘universal’ Cre/loxP system, the protocol can be used in nearly every proliferating cell type in vivo. The protocol takes 4 weeks from transfection of chicken fibroblasts, which act as the host cells for viral production, to the transduction of TVA-transgenic mice.