An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either <it>Piscirickettsia salmonis </it>or IPNV
<p>Abstract</p> <p>Background</p> <p>Due to the limited number of species specific antibodies against fish proteins, differential gene expression analyses are vital for the study of host immune responses. Quantitative real-time reverse transcription PCR (qRT-PCR) is one...
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| Format: | Article |
| Language: | English |
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BMC
2010-04-01
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| Series: | BMC Research Notes |
| Online Access: | http://www.biomedcentral.com/1756-0500/3/101 |
| _version_ | 1818063745571618816 |
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| author | Bols Niels C Marshall Sergio H Peña Andrea A |
| author_facet | Bols Niels C Marshall Sergio H Peña Andrea A |
| author_sort | Bols Niels C |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p>Due to the limited number of species specific antibodies against fish proteins, differential gene expression analyses are vital for the study of host immune responses. Quantitative real-time reverse transcription PCR (qRT-PCR) is one of the most powerful tools for this purpose. Nevertheless, the accuracy of the method will depend on the careful selection of genes whose expression are stable and can be used as internal controls for a particular experimental setting.</p> <p>Findings</p> <p>The expression stability of five commonly used housekeeping genes [beta-actin (<it>ACTB</it>), elongation factor 1-alpha (<it>EF1A</it>), ubiquitin (<it>UBQ</it>), glyceraldehyd-3-phosphate dehydrogenase (<it>GAPDH</it>) and tubulin alpha (<it>TUBA</it>)] were monitored in salmonid cell lines CHSE-214 and RTS11 after infection with two of the most fastidious fish pathogens, the facultative bacterium <it>Piscirickettsia salmonis </it>and the aquabirnavirus IPNV (Infectious Pancreatic Necrosis Virus). After geNorm analysis, <it>UBQ </it>and <it>EF1A </it>appeared as the most stable, although <it>EF1A </it>was slightly upregulated at late stages of <it>P. salmonis </it>infection in RTS11. <it>ACTB </it>instead, showed a good performance in each case, being always considered within the three most stable genes of the panel. In contrast, infection-dependent differential regulation of <it>GAPDH </it>and <it>TUBA </it>was also demonstrated.</p> <p>Conclusion</p> <p>Based on the data presented here with the cell culture models CHSE-214 and RTS11, we suggest the initial choice of <it>UBQ</it>, <it>ACTB </it>and <it>EF1A </it>as reference genes in qRT-PCR assays for studying the effect of <it>P. salmonis </it>and IPNV on the host immune response.</p> |
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| institution | Directory Open Access Journal |
| issn | 1756-0500 |
| language | English |
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| publishDate | 2010-04-01 |
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| spelling | doaj.art-b196438d746f4ac8b23d2aa14c0b4cb22022-12-22T01:45:06ZengBMCBMC Research Notes1756-05002010-04-013110110.1186/1756-0500-3-101An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either <it>Piscirickettsia salmonis </it>or IPNVBols Niels CMarshall Sergio HPeña Andrea A<p>Abstract</p> <p>Background</p> <p>Due to the limited number of species specific antibodies against fish proteins, differential gene expression analyses are vital for the study of host immune responses. Quantitative real-time reverse transcription PCR (qRT-PCR) is one of the most powerful tools for this purpose. Nevertheless, the accuracy of the method will depend on the careful selection of genes whose expression are stable and can be used as internal controls for a particular experimental setting.</p> <p>Findings</p> <p>The expression stability of five commonly used housekeeping genes [beta-actin (<it>ACTB</it>), elongation factor 1-alpha (<it>EF1A</it>), ubiquitin (<it>UBQ</it>), glyceraldehyd-3-phosphate dehydrogenase (<it>GAPDH</it>) and tubulin alpha (<it>TUBA</it>)] were monitored in salmonid cell lines CHSE-214 and RTS11 after infection with two of the most fastidious fish pathogens, the facultative bacterium <it>Piscirickettsia salmonis </it>and the aquabirnavirus IPNV (Infectious Pancreatic Necrosis Virus). After geNorm analysis, <it>UBQ </it>and <it>EF1A </it>appeared as the most stable, although <it>EF1A </it>was slightly upregulated at late stages of <it>P. salmonis </it>infection in RTS11. <it>ACTB </it>instead, showed a good performance in each case, being always considered within the three most stable genes of the panel. In contrast, infection-dependent differential regulation of <it>GAPDH </it>and <it>TUBA </it>was also demonstrated.</p> <p>Conclusion</p> <p>Based on the data presented here with the cell culture models CHSE-214 and RTS11, we suggest the initial choice of <it>UBQ</it>, <it>ACTB </it>and <it>EF1A </it>as reference genes in qRT-PCR assays for studying the effect of <it>P. salmonis </it>and IPNV on the host immune response.</p>http://www.biomedcentral.com/1756-0500/3/101 |
| spellingShingle | Bols Niels C Marshall Sergio H Peña Andrea A An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either <it>Piscirickettsia salmonis </it>or IPNV BMC Research Notes |
| title | An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either <it>Piscirickettsia salmonis </it>or IPNV |
| title_full | An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either <it>Piscirickettsia salmonis </it>or IPNV |
| title_fullStr | An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either <it>Piscirickettsia salmonis </it>or IPNV |
| title_full_unstemmed | An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either <it>Piscirickettsia salmonis </it>or IPNV |
| title_short | An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either <it>Piscirickettsia salmonis </it>or IPNV |
| title_sort | evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either it piscirickettsia salmonis it or ipnv |
| url | http://www.biomedcentral.com/1756-0500/3/101 |
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