Development of an integrative,lacZ transcriptional-fusion plasmid vector for Streptococcus mutans and its use to isolate expressed genes

A new integrational vector, pFP1, containing a promoterless lacZ gene has been constructed for use with Streptococcus mutans. The vector can be grown in Escherichia coli, but cannot replicate in S. mutans. pFP1 can be transformed into S. mutans with selection for kanamycin resistance. It integrates into the chromosome when homologous DNA is present in the vector. pFP1 provides a way for cloning and identifying new genes of S. mutans. When a number of S. mutans transformants were tested on agar containing different sugars, some 19 distinct clones with sucrose- and/or glucose-responsive promoters were isolated. Sequence analysis indicated that the cloned DNA encoded all or part of 29 putative proteins with 52% to 100% similarity to known proteins. When assayed for β-galactosidase activity in BTR medium containing 2% sucrose, glucose or maltose, several genes showed some evidence of sugar regulation, including gtfBK, ftf, scrA, and most dramatically for sucrose regulation, fruA.