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

Fontaine, Laetitia;Dandoy, Damien;Boutry, Céline;Delplace, Brigitte;Hols, Pascal;et.al.
(2010) Applied and Environmental Microbiology — Vol. 76, n° 23, p. 7870-7877 (2010)

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Authors
  • Fontaine, LaetitiaUCLouvain
    Author
  • Dandoy, DamienUCLouvain
    Author
  • Boutry, CélineUCLouvain
    Author
  • Delplace, BrigitteUCLouvain
    Author
  • de Frahan, Marie HenryUCLouvain
    Author
  • Hols, Pascalorcid-logoUCLouvain
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Abstract
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.
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Fontaine, L., Dandoy, D., Boutry, C., Delplace, B., de Frahan, M. H., Fremaux, C., Horvath, P., Boyaval, P., & Hols, P. (2010). Development of a versatile procedure based on natural transformation for marker-free targeted genetic modification in Streptococcus thermophilus. Applied and Environmental Microbiology, 76(23), 7870-7877. https://doi.org/10.1128/AEM.01671-10 (Original work published 2010)