Development of a versatile procedure based on natural transformation for marker-free targeted genetic modification in Streptococcus thermophilus.

摘要

A versatile natural transformation protocol was established in order to facilitate marker-less targeted genetic modification in 18 out of the 19 Streptococcus thermophilus strains tested. Its efficiency enables the use of in vitro-amplified mutagenesis fragments to perform large genetic deletion or insertion. Depending on the phenotype linked to the mutation, marker-less mutants can either be selected in two steps i.e. resistance marker insertion and excision using an adapted Cre-loxP system, or in one step using a powerful positive screening procedure as illustrated here for histidine prototrophy.