Chapter 16

Abstract The number of clones that must be recovered will depend on a number of factors such as the type of selection (e.g. selection for homologous recombinants and against random integrants), number of colonies obtained after selection, method of screening for homologous recombinants (Southern vs PCR), knowledge about the targeting frequency for the locus of interest, etc. For obvious reasons, however, the more clones isolated the higher the probability of recovering a homologous recombinant. In situations in which we have targeted the same locus in multiple instances (e.g. the immunoglobulin heavy chain locus) we have generally observed a similar frequency of homologous recombination events and therefore know approximately how many colonies must be isolated to obtain a homologous recombinant. As a very general number, at least 250-300 colonies should be isolated for examination of a homologous recombination event.