CLONING, EXPRESSION, AND PARTIAL-PURIFICATION OF REP78 - AN ADENOASSOCIATED VIRUS-REPLICATION PROTEIN

Using the vaccinia virus/T7-RNA polymerase transient protein expressio n system, the AAV Rep78 protein was expressed in mammalian cells. Rep7 8 protein was found localized primarily to the nucleus of cells. Maxim al steady-state protein levels were reached as early as 12 hr postinfe ction, with no discernable increase at later time points. The Rep78 pr otein has been partially purified from nuclear extracts of the express ion system. We have successfully used the cloned, purified Rep78 prote in to complement an uninfected HeLa cell extract in an in vitro AAV DN A replication assay. Rep78-containing fractions are sufficient to make an uninfected HeLa cell extract competent for AAV DNA replication. (C ) 1994 Academic Press, Inc.