Abstract
Epitope tags are short peptide sequences that are particularly useful for the characterization of proteins against which no antibody has been developed. Influenza hemagglutinin (HA) tag is one of the most widely used epitope tags as several valuable monoclonal and polyclonal antibodies that can be used in various techniques are commercially available. Therefore, adding a HA tag to a protein of interest is quite helpful to get rapid and cost less information regarding its localization, its expression or its biological function. In this chapter, we describe a process, derived from the Datsenko and Wanner procedure, which allows the introduction of an internal 2HA tag sequence into a chromosomal gene of the bacterial pathogen Salmonella.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Ochman H, Groisman EA (1996) Distribution of pathogenicity islands in Salmonella spp. Infect Immun 64:5410–5412
Hensel M, Shea JE, Gleeson C et al (1995) Simultaneous identification of bacterial virulence genes by negative selection. Science 269:400–403
Figueira R, Holden DW (2012) Functions of the Salmonella pathogenicity island 2 (SPI-2) type III secretion system effectors. Microbiology 158:1147–1161. doi:10.1099/mic.0.058115-0
Stein MA, Leung KY, Zwick M et al (1996) Identification of a Salmonella virulence gene required for formation of filamentous structures containing lysosomal membrane glycoproteins within epithelial cells. Mol Microbiol 20:151–164. doi:10.1111/j.1365-2958.1996.tb02497.x
Brumell JH, Goosney DL, Finlay BB (2002) SifA, a type III secreted effector of Salmonella typhimurium, directs Salmonella-induced filament (Sif) formation along microtubules. Traffic 3:407–415
Zhang L, Hernan R, Brizzard B (2001) Multiple tandem epitope tagging for enhanced detection of protein expressed in mammalian cells. Mol Biotechnol 19:313–321. doi:10.1385/MB:19:3:313
Henry T, Couillault C, Rockenfeller P et al (2006) The Salmonella effector protein PipB2 is a linker for kinesin-1. Proc Natl Acad Sci U S A 103:13497–13502. doi:10.1073/pnas.0605443103
Freeman JA, Ohl ME, Miller SI (2003) The Salmonella enterica serovar typhimurium translocated effectors SseJ and SifB are targeted to the Salmonella-containing vacuole. Infect Immun 71:418–427
Schroeder N, Henry T, de Chastellier C et al (2010) The virulence protein SopD2 regulates membrane dynamics of Salmonella-containing vacuoles. PLoS Pathog 6:e1001002. doi:10.1371/journal.ppat.1001002
Datsenko KA, Wanner BL (2000) One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 97:6640–6645. doi:10.1073/pnas.120163297
Brown NF, Szeto J, Jiang X et al (2006) Mutational analysis of Salmonella translocated effector members SifA and SopD2 reveals domains implicated in translocation, subcellular localization and function. Microbiology 152:2323–2343. doi:10.1099/mic.0.28995-0
Acknowledgments
W.Z. was supported by the China Scholarship Council (CSC Grant). The authors thank Aude-Agnès Blanchoin for her excellent technical assistance.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Zhao, W., Méresse, S. (2015). A Method to Introduce an Internal Tag Sequence into a Salmonella Chromosomal Gene. In: Schatten, H., Eisenstark, A. (eds) Salmonella. Methods in Molecular Biology, vol 1225. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1625-2_5
Download citation
DOI: https://doi.org/10.1007/978-1-4939-1625-2_5
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-1624-5
Online ISBN: 978-1-4939-1625-2
eBook Packages: Springer Protocols