IS30, a new insertion sequence of Escherichia coli K12.

Abstract

Three independent spontaneous mutations of prophage P1 affecting the ability of the phage to reproduce vegetatively are due to the insertion of a mobile genetic element, called IS30. The same sequence is also carried in the R plasmid NR 1-Basel, but not in the parental plasmid NR 1. Southern hybridisation study indicates that the Escherichia coli K 12 chromosome carries several copies of IS30 as a normal resident. IS30 is 1.2 kb long and contains unique restriction cleavage sites for BglII, ClaI, HindIII, NciI and HincII, and it is cleaved twice by the enzymes HpaII and TaqI. The ends of IS30 are formed by 26 bp long inverted repeats with 3 bases mismatched. Upon transposition IS30 generates a duplication of only 2 bp of the target. The following observations suggest a pronounced specificity in target selection by IS30. In transposition to the phage P1 genome a single integration site was used three times independently, and in both orientations. A short region of sequence homology has been identified between the P1 and NR 1-Basel insertion sites. IS30 has mediated cointegration as well as deletion. The entire IS30 sequences were duplicated in the cointegrates between a pBR322 derivative containing IS30 and the genome of phage P1-15, and several loci on the P1-15 genome served as fusion sites, some of which were used more than once.