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Construction of T. thermophilus rpsL mutants by gene replacement. The P90R Strd strain was transformed with plasmid pUC18 containing the rpsL gene with the desired mutation (here, K42T). Plasmid pUC18 does not replicate in T. thermophilus; therefore, replacement of chromosomal rpsL with the plasmid-borne rpsL gene occurs via homologous recombination with selection on antibiotic-free medium.
