Vectors with restriction site banks. V. pJRD215, a wide-host-range cosmid vector with multiple cloning sites

The construction of a new wide-host-range, restriction-site bank, cosmid-cloning vehicle (pJRD215) is described. The wide-host-range properties and the ability to be transferred by conjugation, extend genetic engineering to those Gram-negative species that cannot be transformed. The vector permits the cloning of genes from Gram-negative bacteria using a complementation screening procedure in a mutant host. This procedure is simplified by the possibility of construction of a cosmid gene bank so that only a few hundred clones need to be screened. Subsequent subcloning of the gene of interest is facilitated by the presence of at least 23 unique cloning sites.