Insertion element IS102 resides in plasmid pSC101

Abstract

In vivo recombination was found to occur between plasmid pHS1, a temperature-sensitive replication mutant of pSC101 carrying tetracycline resistance, and plasmid ColE1 after selection for tetracycline resistance at the restrictive temperature, 42 degrees C. Extensive analysis of the physical structures of three of these recombinant plasmids, using restriction endonucleases and the electron microscope heteroduplex method, revealed that the plasmid pHS1 was integrated into different sites on ColE1. The recombinant plasmids contained a duplication of a unique 1-kilobase (kb) sequence of pHS1 in a direct orientation at the junctions between the two parental plasmid sequences. This was confirmed by comparing the nucleotide sequence of the recombinants and their parental plasmids. Nucleotide sequence analysis further revealed that nine nucleotides at the site of recombination of ColE1 were duplicated at the junction of each of the 1-kb sequences. The formation of recombinants was independent of RecA function. Based on our previous finding that a plasmid containing a deoxyribonucleic acid insertion (IS) element can recombine with a second plasmid to generate a duplication of the IS element, we conclude that the 1-kb sequence is an insertion sequence, which we named IS102. For convenience, we have also denoted the IS102 sequence as eta theta to assign the orientation of the sequence. Eighteen nucleotides at one end (eta end) were found to be repeated in an inverted orientation at the other end (theta end) of IS102. The nucleotide sequence of the eta end of the sequence was found to be identical to the sequence at the ends of the transposon Tn903, which is responsible for transposition of the kanamycin resistance gene.