Earticle

초록

영어

Potential utility of 14 candidate housekeeping genes as normalization reference for RT-qPCR analysis with developmental samples (fertilized eggs to late veliger larvae) in Pacific abalone Haliotis discus hannai was evaluated using four different statistical algorithms (geNorm, NormFinder, BestKeeper and comparative ΔCT method). Different algorithms identified different genes as the best candidates, and geometric mean-based final ranking from the most to the least stable expression was as follow: RPL5, RPL4, RPS18, RPL8, RPL7, UBE2, RPL7A, GAPDH, RPL36, PPIB, EF1A, ACTB and B-TU. The findings were further validated via relative quantification of metallothionein (MT) transcripts using the stable and unstable reference genes, and expression levels of MT were greatly influenced according to the choice of reference genes. In overall, our data suggest that RPL5 and RPS18, either singly or in combination, are appropriate for normalizing gene expression in developmental samples of this abalone species, whereas ACTB, B-TU and EF1A are less stable and not recommended. In addition, our findings propose that standard deviations in geometric ranking as well as geometric mean itself should also be taken into account for the final selection of reference gene(s). This study could be a useful basis to facilitate the generation of accurate and reliable RT-qPCR data with developmental samples in this abalone species.

목차

ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
Abalone specimens and biological sampling
Candidate reference genes
Nucleic acid preparation and qPCR
Statistical analysis using different software programs
Validation of expression levels based on choice of different reference genes
RESULTS
Fertilization and hatching success
Cq profiles of reference genes
Time course expression pattern of candidate references during development
geNorm-based analysis of expression stability
Expression stability based on BestKeeper, NormFinder and comparative ΔCT
Geometric mean-based overall ranking
Influence of reference gene choice on relative expression of target gene
DISCUSSION
CONFLICTS OF INTEREST
ACKNOWLEDGEMENTS
AUTHOR'S AFFILIATION, POSITION AND ORCID NO.
REFERENCES

키워드

저자

• Sang Yoon Lee [ Department of Marine Bio-Materials and Aquaculture, Pukyong National University, Busan 48513, Korea ]
• Choul-Ji Park [ Genetics and Breeding Research Center, National Institute of Fisheries Science, Geoje 53334, Korea ]
• Yoon Kwon Nam [ Department of Marine Bio-Materials and Aquaculture, Pukyong National University, Busan 48513, Korea ] Corresponding Author

참고문헌

발행기관

간행물

관심저널 등록

• 간행물명

  Journal of Animal Reproduction and Biotechnology

• 간기

  계간

• pISSN

  2671-4639

• eISSN

  2671-4663

• 수록기간

  2019~2025

• 십진분류

  KDC 527 DDC 636