NOVEL VECTORS FOR IMPROVING CLONING AND EXPRESSION IN LOW COPY NUMBER PLASMIDS.

摘要

Methods that facilitates 1) cloning of large inserts into BAC plasmids, 2) isolation of large amounts of BAC DNA (by increasing plasmid copy number), and 3) increasing heterologous expression from BAC plasmid inserts (by increasing plasmid copy number and/or introducing promoters into the insert). A vector for increasing the copy number of plasmids, comprising a transposable element containing a moderate or high copy number origin of replication capable of in vitro transposition into a target plasmid is provided. The target plasmid is a single or low copy plasmid, e.g. a BAC vector, that is useful for cloning large pieces of DNA. The transposon plasmid may contain any moderate or high copy origin of replication that is compatible with a bacterial host such as E. coli. Thus, an exemplary ori is the colE1 ori from pBR322.