Determining the role of specific signaling molecules during lymphocyte development in vivo: instant transgenesis.
A common method of determining the role of specific signaling molecules during lymphocyte development is to generate a transgenic mouse. This procedure, while informative, is time consuming, expensive and ultimately does not guarantee a defined answer. Here we present a protocol in which the in vivo effects of a gene of interest on both B and T lymphocyte development may be determined simultaneously in a relatively short time period. This is achieved by introducing a defined gene, such as a wild-type or mutated signaling molecule, into a lymphoid progenitor population by retroviral infection. The retrovirus generates a bicistronic message encoding the gene of interest and GFP, thus enabling identification of retrovirally transduced cells in subsequent lymphocyte lineages. The cells are then introduced into mice deficient for recombinase activating gene 1 (Rag-/- mice), thus allowing the development of donor-derived B and T lymphocytes in vivo. Using this technique, results can be obtained within 3-8 weeks.
Nakagawa
Rinako
R
Division of Cancer Sciences and Molecular Pathology, Section of Experimental Haematology, Royal Infirmary, 10 Alexandra Parade, University of Glasgow, Scotland G31 2ER, UK.
Mason
Susan M
SM
Michie
Alison M
AM
D000818:Animals; D019070:Cell Lineage; D018014:Gene Transfer Techniques; D016654:Genes, RAG-1 / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins; D047908:Intracellular Signaling Peptides and Proteins / Q000032:analysis / Q000378:metabolism*; D008214:Lymphocytes / Q000166:cytology* / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D012190:Retroviridae
D047908:Intracellular Signaling Peptides and Proteins; D049452:Green Fluorescent Proteins