Genomics

Dataset Information

43

Comparison of the transcriptomes of a Burkholeria pseudomallei bprR [bpss0688] mutant, a bprS [bpss0687] mutant and a bprRS double mutant with the wild-type B. pseudomallei K96243 parent strain.


ABSTRACT: Objectives: The Burkholderia pseudomallei bprR and bprS genes are predicted to encode the response regulator and sensor kinase components respectively of a two component signal transduction system. Inactivation of either single component of this two component signal transduction system led to attenuation of B. pseudomallei virulence and also reduced motility. However, inactivation of both components resulted in wild-tpe virulence and motility. Therefore, we used RNA-seq to compare the transcritptomes of the three mutant strains (bprR single, bprS single, bprRS double mutant) with the wild-type parent strain in order to identify the genes differenty regulated in the different mutants. Overall design: Methods: RNA sequencing was employed to determine the transcriptomes of three mutant strains (bprR single, bprS single, bprRS double mutant) with the wild-type parent strain B. pseudomallei K96243. B. pseudomallei parent strain K96243 and the directed bprS, bprR and bprRS mutants were grown overnight at 37 °C in LB with appropriate antibiotics (K96243 in gentamicin 64 µg/ml; mutants in gentamicin 64 µg/ml and tetracycline 25 µg/ml). Strains were subcultured 1/50 into fresh antibiotic-free medium and grown at 37 °C with shaking (200 rpm) to an OD600 of 0.5. The cells were harvested by centrifugation, and RNA purified using Trizol reagent (Gibco/BRL) as specified by the manufacturer. RNA was DNase treated using the TURBO DNA-free kit (Ambion), followed by processing with a QIAGEN RNeasy kit clean-up with on-column DNase digestion as per the manufacturer’s instruction. Double-stranded cDNA synthesis and high-throughput RNA-seq were performed as described previously (Henry R, Crane B, Powell D, Deveson Lucas D, et al: The transcriptomic response of Acinetobacter baumannii to colistin and doripenem alone and in combination in an in vitro pharmacokinetics/pharmacodynamics model. J Antimicrob Chemother. 2015; 70:1303-1313). For each strain three biological replicates were sequenced.

INSTRUMENT(S): Illumina MiSeq (Burkholderia pseudomallei K96243)

SUBMITTER: David Richard Powell  

PROVIDER: GSE77970 | GEO | 2016-04-27

SECONDARY ACCESSION(S): PRJNA312225

REPOSITORIES: GEO

Dataset's files

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GSE77970_counts.csv.gz Csv
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Publications

Perturbation of the two-component signal transduction system, BprRS, results in attenuated virulence and motility defects in Burkholderia pseudomallei.

Lazar Adler Natalie R NR   Allwood Elizabeth M EM   Deveson Lucas Deanna D   Harrison Paul P   Watts Stephen S   Dimitropoulos Alexandra A   Treerat Puthayalai P   Alwis Priyangi P   Devenish Rodney J RJ   Prescott Mark M   Govan Brenda B   Adler Ben B   Harper Marina M   Boyce John D JD  

BMC genomics 20160504


<h4>Background</h4>Burkholderia pseudomallei is the causative agent of melioidosis, a severe invasive disease of humans and animals. Initial screening of a B. pseudomallei signature-tagged mutagenesis library identified an attenuated mutant with a transposon insertion in a gene encoding the sensor component of an uncharacterised two-component signal transduction system (TCSTS), which we designated BprRS.<h4>Results</h4>Single gene inactivation of either the response regulator gene (bprR) or the  ...[more]

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