Reverse genetic studies of homologous DNA recombination using the chicken B–lymphocyte line, DT40

Abstract

DT40 is an avian leucosis virus–transformed chicken B–lymphocyte line which exhibits high ratios of targeted to random integration of transfected DNA constructs. This efficient targeted integration may be related to the ongoing diversification of the variable segment of the immunoglobulin gene through homologous DNA recombination–controlled gene conversion. DT40s are a convenient model system for making gene–targeted mutants. Another advantage is the relative tractability of these cells, which makes it possible to disrupt multiple genes in a single cell and to generate conditionally gene–targeted mutants including temperature–sensitive mutants. There are strong phenotypic similarities between murine and DT40 mutants of various genes involved in DNA recombination. These similarities confirm that the DT40 cell line is a reasonable model for the analysis of vertebrate DNA recombination, despite obvious concerns associated with the use of a transformed cell line, which may have certain cell–line–specific characteristics. Here we describe our studies of homologous DNA recombination in vertebrate somatic cells using reverse genetics in DT40 cells.