Imperial College London

DrVladimirPelicic

Faculty of MedicineDepartment of Medicine

Reader in Molecular Bacteriology & Infection
 
 
 
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Contact

 

+44 (0)20 7594 2080v.pelicic Website

 
 
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Location

 

3.20Flowers buildingSouth Kensington Campus

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Summary

 

Publications

Citation

BibTex format

@article{Gurung:2017:nar/gkw1177,
author = {Gurung, I and Berry, J-L and Hall, AMJ and Pelicic, V},
doi = {nar/gkw1177},
journal = {Nucleic Acids Res},
title = {Cloning-independent markerless gene editing in Streptococcus sanguinis: novel insights in type IV pilus biology.},
url = {http://dx.doi.org/10.1093/nar/gkw1177},
volume = {45},
year = {2017}
}

RIS format (EndNote, RefMan)

TY  - JOUR
AB - Streptococcus sanguinis, a naturally competent opportunistic human pathogen, is a Gram-positive workhorse for genomics. It has recently emerged as a model for the study of type IV pili (Tfp)-exceptionally widespread and important prokaryotic filaments. To enhance genetic manipulation of Streptococcus sanguinis, we have developed a cloning-independent methodology, which uses a counterselectable marker and allows sophisticated markerless gene editing in situ. We illustrate the utility of this methodology by answering several questions regarding Tfp biology by (i) deleting single or mutiple genes, (ii) altering specific bases in genes of interest, and (iii) engineering genes to encode proteins with appended affinity tags. We show that (i) the last six genes in the pil locus harbouring all the genes dedicated to Tfp biology play no role in piliation or Tfp-mediated motility, (ii) two highly conserved Asp residues are crucial for enzymatic activity of the prepilin peptidase PilD and (iii) that pilin subunits with a C-terminally appended hexa-histidine (6His) tag are still assembled into functional Tfp. The methodology for genetic manipulation we describe here should be broadly applicable.
AU - Gurung,I
AU - Berry,J-L
AU - Hall,AMJ
AU - Pelicic,V
DO - nar/gkw1177
PY - 2017///
TI - Cloning-independent markerless gene editing in Streptococcus sanguinis: novel insights in type IV pilus biology.
T2 - Nucleic Acids Res
UR - http://dx.doi.org/10.1093/nar/gkw1177
UR - https://www.ncbi.nlm.nih.gov/pubmed/27903891
UR - http://hdl.handle.net/10044/1/42506
VL - 45
ER -