Open AccessUnmarked gene modification in Streptococcus mutans by a cotransformation strategy with a thermosensitive plasmid
Author(s) -
Indranil Biswas,
Laura Drake,
Sean R. Johnson,
Damon Thielen
Publication year - 2007
Publication title -
biotechniques/biotechniques
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.617
H-Index - 131
eISSN - 1940-9818
pISSN - 0736-6205
DOI - 10.2144/000112414
Subject(s) - plasmid , streptococcus mutans , biology , mutant , replicon , selectable marker , gene , genetics , allele , bacteria , microbiology and biotechnology
Unmarked gene modifications are desirable for various genetic analyses; however, they have been difficult to construct without selectable markers. We describe here a new genetic method for constructing unmarked mutants in Streptococcus mutans. The desired mutant allele is first constructed and introduced into the strain by cotransformation with pGhost4, a thermosensitive plasmid that replicates in several low G +C Gram positive bacteria. With this method, we have modified two different loci with high frequency by insertion or deletion in S. mutans. Because pGhost4 contains a broad host range thermosensitive replicon, this method can be applied to any transformable low G+C Gram positive bacteria, including oral streptococci.