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Title: Reference gene selection for the shell gland of laying hens in response to time-points of eggshell formation and nicarbazin
Contributor(s): Samiullah, Samiullah (author); Roberts, Julie R  (author); Wu, Shubiao  (author)orcid 
Publication Date: 2017
Open Access: Yes
DOI: 10.1371/journal.pone.0180432Open Access Link
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Abstract: Ten reference genes were investigated for normalization of gene expression data in the shell gland of laying hens. Analyses performed with geNorm revealed that hypoxanthine phosphoribosyltransferase 1 (HPRT1) and hydroxymethylbilane synthase (HMBS) were the two most stable reference genes in response to post-oviposition time alone (POT) or with nicarbazin treatment (POT+N) of laying hens. NormFinder analyses showed that the two most stable reference genes in response to POT and POT+N were 18S ribosomal RNA (18S rRNA), ribosomal protein L4 (RPL4) and HMBS, RPL4, respectively. BestKeeper analyses showed that 18S rRNA, RPL4 and HPRT1, HMBS were the two most stable reference genes for POT, and POT+N, respectively. Of the ten reference genes, all except B2M showed geNorm M <0.5, suggesting that they were stably expressed in the shell gland tissue. Consensus from these three programs suggested HPRT1 and HMBS could be used as the two most stable reference genes in the present study. Expression analyses of four candidate target genes with the two most and the two least stable genes showed that a combination of stable reference genes leads to more discriminable quantification of expression levels of target genes, while the least stable genes failed to do so. Therefore, HMBS and HPRT1 are recommended as the two most stable reference genes for the normalization of gene expression data at different stages of eggshell formation in brown-egg laying hens. Available statistical programs for reference gene ranking should include more robust analysis capability to analyse the gene expression data generated from factorial design experiments.
Publication Type: Journal Article
Source of Publication: PLoS One, 12(7), p. 1-20
Publisher: Public Library of Science
Place of Publication: United States of America
ISSN: 1932-6203
Fields of Research (FoR) 2008: 070702 Veterinary Anatomy and Physiology
070206 Animal Reproduction
060405 Gene Expression (incl. Microarray and other genome-wide approaches)
Fields of Research (FoR) 2020: 300902 Veterinary anatomy and physiology
300305 Animal reproduction and breeding
310505 Gene expression (incl. microarray and other genome-wide approaches)
Socio-Economic Objective (SEO) 2008: 970107 Expanding Knowledge in the Agricultural and Veterinary Sciences
830309 Poultry
Socio-Economic Objective (SEO) 2020: 280101 Expanding knowledge in the agricultural, food and veterinary sciences
100411 Poultry
Peer Reviewed: Yes
HERDC Category Description: C1 Refereed Article in a Scholarly Journal
Appears in Collections:Journal Article
School of Environmental and Rural Science

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