Identification of Reference Genes in Chicken Intraepithelial Lymphocyte Natural Killer Cells Infected with Very-virulent Infectious Bursal Disease Virus

Abstract Due to the limitations in the range of antibodies recognising avian viruses, quantitative real-time PCR (RT-qPCR) is still the most widely used method to evaluate the expression of immunologically related genes in avian viruses. The objective of this study was to identify suitable reference...

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Autores principales:	Sook Yee Boo, Sheau Wei Tan, Noorjahan Banu Alitheen, Chai Ling Ho, Abdul Rahman Omar, Swee Keong Yeap
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2020
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/7f4faba3fa3945edb1a53a82187f0e95
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spelling	oai:doaj.org-article:7f4faba3fa3945edb1a53a82187f0e952021-12-02T14:59:15ZIdentification of Reference Genes in Chicken Intraepithelial Lymphocyte Natural Killer Cells Infected with Very-virulent Infectious Bursal Disease Virus10.1038/s41598-020-65474-32045-2322https://doaj.org/article/7f4faba3fa3945edb1a53a82187f0e952020-05-01T00:00:00Zhttps://doi.org/10.1038/s41598-020-65474-3https://doaj.org/toc/2045-2322Abstract Due to the limitations in the range of antibodies recognising avian viruses, quantitative real-time PCR (RT-qPCR) is still the most widely used method to evaluate the expression of immunologically related genes in avian viruses. The objective of this study was to identify suitable reference genes for mRNA expression analysis in chicken intraepithelial lymphocyte natural killer (IEL-NK) cells after infection with very-virulent infectious bursal disease virus (vvIBDV). Fifteen potential reference genes were selected based on the references available. The coefficient of variation percentage (CV%) and average count of these 15 genes were determined by NanoString technology for control and infected samples. The M and V values for shortlisted reference genes (ACTB, GAPDH, HMBS, HPRT1, SDHA, TUBB1 and YWHAZ) were calculated using geNorm and NormFinder. GAPDH, YWHAZ and HMBS were the most stably expressed genes. The expression levels of three innate immune response related target genes, CASP8, IL22 and TLR3, agreed in the NanoString and RNA sequencing (RNA-Seq) results using one or two reference genes for normalisation (not HMBS). In conclusion, GAPDH and YWHAZ could be used as reference genes for the normalisation of chicken IEL-NK cell gene responses to infection with vvIBDV.Sook Yee BooSheau Wei TanNoorjahan Banu AlitheenChai Ling HoAbdul Rahman OmarSwee Keong YeapNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 10, Iss 1, Pp 1-9 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Sook Yee Boo Sheau Wei Tan Noorjahan Banu Alitheen Chai Ling Ho Abdul Rahman Omar Swee Keong Yeap Identification of Reference Genes in Chicken Intraepithelial Lymphocyte Natural Killer Cells Infected with Very-virulent Infectious Bursal Disease Virus
description	Abstract Due to the limitations in the range of antibodies recognising avian viruses, quantitative real-time PCR (RT-qPCR) is still the most widely used method to evaluate the expression of immunologically related genes in avian viruses. The objective of this study was to identify suitable reference genes for mRNA expression analysis in chicken intraepithelial lymphocyte natural killer (IEL-NK) cells after infection with very-virulent infectious bursal disease virus (vvIBDV). Fifteen potential reference genes were selected based on the references available. The coefficient of variation percentage (CV%) and average count of these 15 genes were determined by NanoString technology for control and infected samples. The M and V values for shortlisted reference genes (ACTB, GAPDH, HMBS, HPRT1, SDHA, TUBB1 and YWHAZ) were calculated using geNorm and NormFinder. GAPDH, YWHAZ and HMBS were the most stably expressed genes. The expression levels of three innate immune response related target genes, CASP8, IL22 and TLR3, agreed in the NanoString and RNA sequencing (RNA-Seq) results using one or two reference genes for normalisation (not HMBS). In conclusion, GAPDH and YWHAZ could be used as reference genes for the normalisation of chicken IEL-NK cell gene responses to infection with vvIBDV.
format	article
author	Sook Yee Boo Sheau Wei Tan Noorjahan Banu Alitheen Chai Ling Ho Abdul Rahman Omar Swee Keong Yeap
author_facet	Sook Yee Boo Sheau Wei Tan Noorjahan Banu Alitheen Chai Ling Ho Abdul Rahman Omar Swee Keong Yeap
author_sort	Sook Yee Boo
title	Identification of Reference Genes in Chicken Intraepithelial Lymphocyte Natural Killer Cells Infected with Very-virulent Infectious Bursal Disease Virus
title_short	Identification of Reference Genes in Chicken Intraepithelial Lymphocyte Natural Killer Cells Infected with Very-virulent Infectious Bursal Disease Virus
title_full	Identification of Reference Genes in Chicken Intraepithelial Lymphocyte Natural Killer Cells Infected with Very-virulent Infectious Bursal Disease Virus
title_fullStr	Identification of Reference Genes in Chicken Intraepithelial Lymphocyte Natural Killer Cells Infected with Very-virulent Infectious Bursal Disease Virus
title_full_unstemmed	Identification of Reference Genes in Chicken Intraepithelial Lymphocyte Natural Killer Cells Infected with Very-virulent Infectious Bursal Disease Virus
title_sort	identification of reference genes in chicken intraepithelial lymphocyte natural killer cells infected with very-virulent infectious bursal disease virus
publisher	Nature Portfolio
publishDate	2020
url	https://doaj.org/article/7f4faba3fa3945edb1a53a82187f0e95
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Identification of Reference Genes in Chicken Intraepithelial Lymphocyte Natural Killer Cells Infected with Very-virulent Infectious Bursal Disease Virus

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