AMPLIFICATION OF BACTERIOPHAGE-MU DNA BY ROLLING CIRCLE DNA-REPLICATION IN-VITRO

When mini-Mu DNA was allowed to transpose and replicate in vitro over a prolonged period, the products consisted not only of simple inserts and cointegrates but also high molecular weight DNA many times the uni t length of mini-Mu. This high molecular weight product contained pred ominantly full-length mini-Mu DNA and relatively little non-Mu DNA (th e vector harboring the mini-Mu element and the target for transpositio n in the reaction system). It arose from rolling circle DNA replicatio n of templates created by intramolecular strand transfer, which is cat alyzed by Mu transposition proteins. A donor substrate, which is a sup ercoiled plasmid bearing a mini-Mu element, gave rise to large amounts of the high molecular weight product provided that the vector segment outside the mini-Mu element was 2 kilobase pairs or more. When a dono r substrate had a vector segment of only 600 base pairs, the mini-Mu e lement first had to transpose to a larger circular target before givin g rise to the high molecular weight product. These results suggest a m echanism by which Mu DNA can be amplified for lytic development withou t transposing multiple times. By establishing a circular template, mul tiple copies of Mu can be processively generated from a single initiat ion event.