Base substitutions in transposable element IS1 cause DNA duplication of variable length at the target site for plasmid co-integration

Abstract

We demonstrate that base substitutions in the IS1 sequence affect the length of the nucleotide sequence which is duplicated during IS1-mediated co-integration. IS1K, an IS1 variant present in the Escherichia coli chromosome, has seven base substitutions in its sequence as compared with that of IS1R derived from the plasmid R100. All substitutions are located in the internal region of IS1K. We have constructed plasmids containing IS1R, IS1K and hybrids between them: one contains four base substitutions causing an amino acid substitution in the insA gene and the other has three substitutions producing an amino acid substitution in the insB gene. We have isolated co-integrate plasmids formed by each IS1 and analysed nucleotide sequences of the target sites duplicated at the co-integration junctions. The results show that IS1K generates duplications of 8 or 14 bp as well as 9 bp, while IS1R exclusively generates the 9-bp duplications. Both hybrid IS1s also create 8- or 7-bp target duplications in addition to 9-bp duplications. These results indicate that the base substitutions in either insA or insB are sufficient for the occurrence of unusual target duplications, suggesting that both genes are involved in the target duplication.