Evaluation of reference genes for reverse transcription quantitative PCR analyses of fish-pathogenic Francisella strains exposed to different growth conditions

BMC Research Notes

Samuel Duodu¹  Duncan John Colquhoun¹  Hanne Cecilie Winther-Larsen²  Espen Brudal³ 

[1] Section for Bacteriology, Norwegian Veterinary Institute, PO Box 750, 0106, Sentrum, Oslo, Norway;School of Pharmacy, University of Oslo, PO Box 1068, Blindern, Oslo 0316, Norway;Laboratory for Microbial Dynamics (LaMDa), University of Oslo, PO Box 1068, Blindern, Oslo 0316, Norway

关键词: Francisellosis;    Fish;    Francisella noatunensis;    RT-qPCR;    Gene expression;   

Others  :  1143371
DOI  :  10.1186/1756-0500-6-76

 received in 2012-12-21, accepted in 2013-02-27,  发布年份 2013

【 摘 要 】

Background

Reverse transcription quantitative PCR has become a powerful technique to monitor mRNA transcription in response to different environmental conditions in many bacterial species. However, correct evaluation of data requires accurate and reliable use of reference genes whose transcription does not change during the course of the experiment. In the present study exposure to different growth conditions was used to validate the transcription stability of eight reference gene candidates in three strains from two subspecies of Francisella noatunensis, a pathogen causing disease in both warm and cold water fish species.

Results

Relative transcription levels for genes encoding DNA gyrase (gyrA), RNA polymerase beta subunit (rpoB), DNA polymerase I (polA), cell division protein (ftsZ), outer membrane protein (fopA), riboflavin biosynthesis protein (ribC), 16S ribosomal RNA (16S rRNA) and DNA helicases (uvrD) were quantified under exponential, stationary and iron-restricted growth conditions. The suitability of selected reference genes for reliable interpretation of gene expression data was tested using the virulence-associated intracellular growth locus subunit C (iglC) gene.

Conclusion

Although the transcription stability of the reference genes was slightly different in the three strains studied, fopA, ftsZ and polA proved to be the most stable and suitable for normalization of gene transcription in Francisella noatunensis ssp.

【 授权许可】

   
2013 Brudal et al; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

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