Selective amplification of Brucella melitensis mRNA from a mixed host:pathogen total RNA

Rossetti, Carlos A.; Galindo, Cristi L.; Garner, Harold R.; Adams, L. Garry

Selective amplification of Brucella melitensis mRNA from a mixed host:pathogen total RNA

dc.contributor.author	Rossetti, Carlos A.	en
dc.contributor.author	Galindo, Cristi L.	en
dc.contributor.author	Garner, Harold R.	en
dc.contributor.author	Adams, L. Garry	en
dc.date.accessioned	2012-08-24T11:16:10Z	en
dc.date.available	2012-08-24T11:16:10Z	en
dc.date.issued	2010-09-28	en
dc.date.updated	2012-08-24T11:16:10Z	en
dc.description.abstract	Background Brucellosis is a worldwide anthropozoonotic disease caused by an in vivo intracellular pathogen belonging to genus Brucella. The characterization of brucelae transcriptome's during host-pathogen interaction has been limited due to the difficulty of obtaining an adequate quantity of good quality eukaryotic RNA-free pathogen RNA for downstream applications. Findings Here, we describe a combined protocol to prepare RNA from intracellular B. melitensis in a quantity and quality suitable for pathogen gene expression analysis. Initially, B. melitensis total RNA was enriched from a host:pathogen mixed RNA sample by reducing the eukaryotic RNA..Then, to increase the Brucella RNA concentration and simultaneously minimize the contaminated host RNA in the mixed sample, a specific primer set designed to anneal to all B. melitensis ORF allows the selective linear amplification of sense-strand prokaryotic transcripts in a previously enriched RNA sample. Conclusion The novelty of the method we present here allows analysis of the gene expression profile of B. melitensis when limited amounts of pathogen RNA are present, and is potentially applicable to both in vivo and in vitro models of infection, even at early infection time points.	en
dc.description.version	Published version	en
dc.format.mimetype	application/pdf	en
dc.identifier.citation	BMC Research Notes. 2010 Sep 28;3(1):244	en
dc.identifier.doi	https://doi.org/10.1186/1756-0500-3-244	en
dc.identifier.uri	http://hdl.handle.net/10919/18824	en
dc.language.iso	en	en
dc.rights	Creative Commons Attribution 4.0 International	en
dc.rights.holder	Carlos A Rossetti et al.; licensee BioMed Central Ltd.	en
dc.rights.uri	http://creativecommons.org/licenses/by/4.0/	en
dc.title	Selective amplification of Brucella melitensis mRNA from a mixed host:pathogen total RNA	en
dc.title.serial	BMC Research Notes	en
dc.type	Article - Refereed	en
dc.type.dcmitype	Text	en