A selective lambda phage cloning vector with automatic excision of the insert in a plasmid

Abstract

A bacteriophage lambda cloning vehicle has been constructed for the generation of cDNA libraries. The vector has the following properties. (1) It has a unique BamHI site engineered into the lambda gam gene. Segments of DNA can be cloned into this site and clones with an insert can be selected by their ability to grow on an Escherichia coli host lysogenic for phage P2 (Spi- phenotype). (2) When the recombinant phage infects a Cre-producing E. coli strain, a site-specific recombination event results in the excision of a plasmid replicon with the cloned insert. (3) Single-stranded DNAs can be recovered by growing helper M13 phages on bacteria harboring such plasmids. The vector, lambda MGU2, has been used to construct a nematode (Caenorhabditis elegans) cDNA library.