RESUMO
OBJECTIVE: This study was aimed at sequence analysis and function annotation of plasmid pKKma carrying a mariner transposon. METHODS: Primers were designed based on the partial known sequence and used for directly sequencing plasmid pKKma. Transposon mutagenesis libraries were constructed to analyze the mutagenesis efficiency of plasmid pKKma. RESULTS: pKKma comprises 6879 bp with 7 open reading frames (ORFs). Among them, ORF6 encodes a mariner transposase of 348 amino acids (aa), a C9 variant of Himar1 type transposase. Two inverted terminal repeats (ITRs) are identified and of 27 bp each. ORF7 encodes gentamycin resistance gene aacC1, locating between two ITRs. Transposable sequence alignment with other mariner transposons shows that the coverage is 2.0% -47.7% and the homology is 3.2% to 99.7%. The result indicates pKKma is significantly different from the other vectors with mariner transposon. The transposition efficiency is also analyzed. It's (3.1 x 10(-4)) - (4.8 x 10(-4)) for S. marcascens and (1.3 x 10(-3)) - (1.7 x 10(-3)) for C. freundii, respectively. CONCLUSION: pKKma carries a new mariner transposon and could be used to study the role of genes by constructing transposon libraries in bacteria.
