Retrovirus cDNA expression library screening for oncogenes

This chapter describes a retrovirus-based cDNA expression system and its successful application to the identification of novel oncogenes. The system described here permits the stable transfer and expression of large numbers of cDNA clones into equivalent numbers of recipient cells. This allows for the efficient screening of complex cDNA libraries, and facilitates the identification of transforming sequences that are present at low frequency within the cDNA population. In these systems, cDNA expression libraries are constructed in retroviral plasmids, and then converted into libraries of infectious retroviral particles. Four major advantages are obtained through the use of retroviral library transfer: (1) the ability to screen large numbers of cDNA clones on an equivalent number of recipient cells, (2) the relatively high levels of expression obtained with retrovirally transferred cDNA, (3) the potential to use cell lines that have been inaccessible to expression cloning because of low transfection efficiencies, and (4) the development of highly efficient recovery mechanisms for the proviral inserts. The chapter concludes with a discussion on cloning polymerase chain reaction products.