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논문 기본 정보
- 자료유형
- 학술저널
- 저자정보
- 발행연도
- 2016.1
- 수록면
- 1 - 11 (11page)
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Background: The evaluation of suitable reference genes as normalization controls is a prerequisite requirement for launching quantitative reverse transcription-PCR (RT-qPCR)-based expression study. In order to select the stable reference genes in abalone Haliotis discus hannai tissues (gill and hepatopancreas) under heavy metal exposure conditions (Cu, Zn, and Cd), 12 potential candidate housekeeping genes were subjected to expression stability based on the comprehensive ranking while integrating four different statistical algorithms (geNorm, NormFinder, BestKeeper, and ΔCT method).
Results: Expression stability in the gill subset was determined as RPL7 > RPL8 > ACTB > RPL3 > PPIB > RPL7A > EF1A > RPL4 > GAPDH > RPL5 > UBE2 > B-TU. On the other hand, the ranking in the subset for hepatopancreas was RPL7 > RPL3 > RPL8 > ACTB > RPL4 > EF1A > RPL5 > RPL7A > B-TU > UBE2 > PPIB > GAPDH. The pairwise variation assessed by the geNorm program indicates that two reference genes could be sufficient for accurate normalization in both gill and hepatopancreas subsets. Overall, both gill and hepatopancreas subsets recommended ribosomal protein genes (particularly RPL7) as stable references, whereas traditional housekeepers such as β-tubulin (B-TU) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes were ranked as unstable genes. The validation of reference gene selection was confirmed with the quantitative assay of MT transcripts.
Conclusions: The present analysis showed the importance of validating reference genes with multiple algorithmic approaches to select genes that are truly stable. Our results indicate that expression stability of a given reference gene could not always have consensus across tissue types. The data from this study could be a good guide for the future design of RT-qPCR studies with respect to metal regulation/detoxification and other related physiologies in this abalone species.
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참고문헌 (41)
참고문헌 신청
Amiard JC / 2006 / Metallothioneins in aquatic invertebrates : their role in metal detoxification and their use as biomarkers / Aquat Toxicol 76:160~202
Andersen CL / 2004 / Normalization of real-time quantitative reverse transcription-PCR data : a model-based variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets / Cancer Res 64:5245~5250
Bustin SA / 2009 / The MIQE guidelines : minimum information for publication of quantitative real-time PCR experiments / Clin Chem 55:611~622
Cho YS / 2008 / Differential modulations of two glyceraldehyde 3-phosphate dehydrogenase mRNAs in response to bacterial and viral challenges in a marine teleost Oplegnathus fasciatus(Perciformes) / Fish Shellfish Immunol 25:472~476
조영선 / 2011 / Isoform-specific response of two GAPDH paralogs during bacterial challenge and metal exposure in mud loach (Misgurnus mizolepis; Cypriniformes) kidney and spleen / 한국어병학회지 24(3):269~278
Andersen CL / 2004 / Normalization of real-time quantitative reverse transcription-PCR data : a model-based variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets / Cancer Res 64:5245~5250
Bustin SA / 2009 / The MIQE guidelines : minimum information for publication of quantitative real-time PCR experiments / Clin Chem 55:611~622
Cho YS / 2008 / Differential modulations of two glyceraldehyde 3-phosphate dehydrogenase mRNAs in response to bacterial and viral challenges in a marine teleost Oplegnathus fasciatus(Perciformes) / Fish Shellfish Immunol 25:472~476
조영선 / 2011 / Isoform-specific response of two GAPDH paralogs during bacterial challenge and metal exposure in mud loach (Misgurnus mizolepis; Cypriniformes) kidney and spleen / 한국어병학회지 24(3):269~278
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