Part C: Generation of Human Gene Reporters Using Bacterial Artificial Chromosome Recombineering

The ability to insert a gene of interest into a specific chromosomal location has been facilitated by the use of “Bacterial Artificial Chromosome (BAC) recombineering.” In this scheme, homologous recombination occurs in bacteria between the gene of interest (flanked by appropriate regions of homology) and a bacterial artificial chromosome encompassing the target region. Successful recombination is then confirmed by PCR, and the recombineered BAC is subsequently purified and introduced into eukaryotic cells. This chapter describes the application of this technique to human embryonic stem cells (hES cells).