A cosmid vector that facilitates restriction enzyme mapping

Abstract

We describe the construction and use of a cosmid vector, loric, which is derived from the phage lambda origin of replication and appears to be more stable than ColE1-derived cosmids. Loric recombinants can be efficiently packaged in vivo to yield 100-300 micrograms of DNA per liter that is linear and has single-stranded cos ends. We call such molecules "phosmids." Phosmid restriction maps can be rapidly generated by labeling either the left or right cos site by annealing on a 32P-labeled oligonucleotide complementary to either cos-L or cos-R. Partial restriction enzyme digestion, agarose gel electrophoresis, and autoradiography are used to size restriction fragments of increasing length, all of which terminate at the labeled cos site. The procedures have been tested by isolating and mapping a region of the H-2 locus of mouse chromosome 17.