Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically <named-content content-type="genus-species">Staphylococcus aureus</named-content> and <named-content content-type="genus-species">Staphylococcus epidermidis</named-content>

ABSTRACT The strong restriction barrier present in Staphylococcus aureus and Staphylococcus epidermidis has limited functional genomic analysis to a small subset of strains that are amenable to genetic manipulation. Recently, a conserved type IV restriction system termed SauUSI (which specifically r...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Ian R. Monk, Ishita M. Shah, Min Xu, Man-Wah Tan, Timothy J. Foster
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2012
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/7474971ef04644b1a717447b4bbf1a28
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:7474971ef04644b1a717447b4bbf1a28
record_format	dspace
spelling	oai:doaj.org-article:7474971ef04644b1a717447b4bbf1a282021-11-15T15:39:08ZTransforming the Untransformable: Application of Direct Transformation To Manipulate Genetically <named-content content-type="genus-species">Staphylococcus aureus</named-content> and <named-content content-type="genus-species">Staphylococcus epidermidis</named-content>10.1128/mBio.00277-112150-7511https://doaj.org/article/7474971ef04644b1a717447b4bbf1a282012-05-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00277-11https://doaj.org/toc/2150-7511ABSTRACT The strong restriction barrier present in Staphylococcus aureus and Staphylococcus epidermidis has limited functional genomic analysis to a small subset of strains that are amenable to genetic manipulation. Recently, a conserved type IV restriction system termed SauUSI (which specifically recognizes cytosine methylated DNA) was identified as the major barrier to transformation with foreign DNA. Here we have independently corroborated these findings in a widely used laboratory strain of S. aureus. Additionally, we have constructed a DNA cytosine methyltransferase mutant in the high-efficiency Escherichia coli cloning strain DH10B (called DC10B). Plasmids isolated from DC10B can be directly transformed into clinical isolates of S. aureus and S. epidermidis. We also show that the loss of restriction (both type I and IV) in an S. aureus USA300 strain does not have an impact on virulence. Circumventing the SauUSI restriction barrier, combined with an improved deletion and transformation protocol, has allowed the genetic manipulation of previously untransformable strains of these important opportunistic pathogens. IMPORTANCE Staphylococcal infections place a huge burden on the health care sector due both to their severity and also to the economic impact of treating the infections because of prolonged hospitalization. To improve the understanding of Staphylococcus aureus and Staphylococcus epidermidis infections, we have developed a series of improved techniques that allow the genetic manipulation of strains that were previously refractory to transformation. These developments will speed up the process of mutant construction and increase our understanding of these species as a whole, rather than just a small subset of strains that could previously be manipulated.Ian R. MonkIshita M. ShahMin XuMan-Wah TanTimothy J. FosterAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 3, Iss 2 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Ian R. Monk Ishita M. Shah Min Xu Man-Wah Tan Timothy J. Foster Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically <named-content content-type="genus-species">Staphylococcus aureus</named-content> and <named-content content-type="genus-species">Staphylococcus epidermidis</named-content>
description	ABSTRACT The strong restriction barrier present in Staphylococcus aureus and Staphylococcus epidermidis has limited functional genomic analysis to a small subset of strains that are amenable to genetic manipulation. Recently, a conserved type IV restriction system termed SauUSI (which specifically recognizes cytosine methylated DNA) was identified as the major barrier to transformation with foreign DNA. Here we have independently corroborated these findings in a widely used laboratory strain of S. aureus. Additionally, we have constructed a DNA cytosine methyltransferase mutant in the high-efficiency Escherichia coli cloning strain DH10B (called DC10B). Plasmids isolated from DC10B can be directly transformed into clinical isolates of S. aureus and S. epidermidis. We also show that the loss of restriction (both type I and IV) in an S. aureus USA300 strain does not have an impact on virulence. Circumventing the SauUSI restriction barrier, combined with an improved deletion and transformation protocol, has allowed the genetic manipulation of previously untransformable strains of these important opportunistic pathogens. IMPORTANCE Staphylococcal infections place a huge burden on the health care sector due both to their severity and also to the economic impact of treating the infections because of prolonged hospitalization. To improve the understanding of Staphylococcus aureus and Staphylococcus epidermidis infections, we have developed a series of improved techniques that allow the genetic manipulation of strains that were previously refractory to transformation. These developments will speed up the process of mutant construction and increase our understanding of these species as a whole, rather than just a small subset of strains that could previously be manipulated.
format	article
author	Ian R. Monk Ishita M. Shah Min Xu Man-Wah Tan Timothy J. Foster
author_facet	Ian R. Monk Ishita M. Shah Min Xu Man-Wah Tan Timothy J. Foster
author_sort	Ian R. Monk
title	Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically <named-content content-type="genus-species">Staphylococcus aureus</named-content> and <named-content content-type="genus-species">Staphylococcus epidermidis</named-content>
title_short	Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically <named-content content-type="genus-species">Staphylococcus aureus</named-content> and <named-content content-type="genus-species">Staphylococcus epidermidis</named-content>
title_full	Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically <named-content content-type="genus-species">Staphylococcus aureus</named-content> and <named-content content-type="genus-species">Staphylococcus epidermidis</named-content>
title_fullStr	Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically <named-content content-type="genus-species">Staphylococcus aureus</named-content> and <named-content content-type="genus-species">Staphylococcus epidermidis</named-content>
title_full_unstemmed	Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically <named-content content-type="genus-species">Staphylococcus aureus</named-content> and <named-content content-type="genus-species">Staphylococcus epidermidis</named-content>
title_sort	transforming the untransformable: application of direct transformation to manipulate genetically <named-content content-type="genus-species">staphylococcus aureus</named-content> and <named-content content-type="genus-species">staphylococcus epidermidis</named-content>
publisher	American Society for Microbiology
publishDate	2012
url	https://doaj.org/article/7474971ef04644b1a717447b4bbf1a28
work_keys_str_mv	AT ianrmonk transformingtheuntransformableapplicationofdirecttransformationtomanipulategeneticallynamedcontentcontenttypegenusspeciesstaphylococcusaureusnamedcontentandnamedcontentcontenttypegenusspeciesstaphylococcusepidermidisnamedcontent AT ishitamshah transformingtheuntransformableapplicationofdirecttransformationtomanipulategeneticallynamedcontentcontenttypegenusspeciesstaphylococcusaureusnamedcontentandnamedcontentcontenttypegenusspeciesstaphylococcusepidermidisnamedcontent AT minxu transformingtheuntransformableapplicationofdirecttransformationtomanipulategeneticallynamedcontentcontenttypegenusspeciesstaphylococcusaureusnamedcontentandnamedcontentcontenttypegenusspeciesstaphylococcusepidermidisnamedcontent AT manwahtan transformingtheuntransformableapplicationofdirecttransformationtomanipulategeneticallynamedcontentcontenttypegenusspeciesstaphylococcusaureusnamedcontentandnamedcontentcontenttypegenusspeciesstaphylococcusepidermidisnamedcontent AT timothyjfoster transformingtheuntransformableapplicationofdirecttransformationtomanipulategeneticallynamedcontentcontenttypegenusspeciesstaphylococcusaureusnamedcontentandnamedcontentcontenttypegenusspeciesstaphylococcusepidermidisnamedcontent
_version_	1718427771651227648

Transforming the Untransformable: Application of Direct Transformation To Manipulate Genetically <named-content content-type="genus-species">Staphylococcus aureus</named-content> and <named-content content-type="genus-species">Staphylococcus epidermidis</named-content>

Ejemplares similares