Gentamicin resistance gene residing in Serretia marsescens R plasmid, pIS199G2 has been cloned after it was transformed to E. coli. Two plasmids, pSY1 and pSW1 were obtained by ligating partial digests of pIS199G2 with Sau3A1 into pUC9. Plamid pSY1 contains 1.5kb insert and DNA sequence analysis showed that -35 region of gentamicin resistance gene has been provided by inverted repeat of Tn3. Plasmid pSW1 was found to be formed by self ligation of Sau3A fragments, containing the Ori site of R plasmid and an ampicillin resistance gene. In spite of its lower copy number, however, pSW1 showed much stronger resistance to gentamicin than pSY1. In order to explain these discrepancies between these plasmids, DNA sequence of pSW1 was determined in this study. This study revealed that 1386 bp of Tn3 including bla gene was situated at 5' of gentamicin resistance gene, while only 70 bp of that is found in pSY1. Based on the DNA sequence comparison and a modified MIC test of variable recombinant plasmids, it was concluded that the bla gene and 3' region of Tn3 sequence in pSW1 increase the expression of the gentamicin resistant gene.
