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

被引：17

作者：

Boo, Sook Yee ^{[1
]}

Tan, Sheau Wei ^{[1
]}

Alitheen, Noorjahan Banu ^{[2
]}

Ho, Chai Ling ^{[2
]}

Omar, Abdul Rahman ^{[1
]}

Yeap, Swee Keong ^{[1
,3
]}

机构：

[1] Univ Putra Malaysia, Inst Biosci, Lab Vaccines & Immunotherapeut, Serdang 43400, Selangor, Malaysia

[2] Univ Putra Malaysia, Fac Biotechnol & Biomol Sci, Serdang 43400, Selangor, Malaysia

[3] Xiamen Univ Malaysia, China ASEAN Coll Marine Sci, Sepang 43900, Selangor, Malaysia

来源：

SCIENTIFIC REPORTS | 2020年 / 10卷 / 01期

关键词：

POLYMERASE-CHAIN-REACTION; HOUSEKEEPING GENES; INTERNAL CONTROL; HOST-DEFENSE; PCR ANALYSIS; EXPRESSION; NORMALIZATION; SELECTION; INTERLEUKIN-22; RNA;

D O I：

10.1038/s41598-020-65474-3

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

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.

引用

页数：9

共 39 条

[1] 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 [J].

Andersen, CL ;

Jensen, JL ;

Orntoft, TF .

CANCER RESEARCH, 2004, 64 (15) :5245-5250

[2] IL-22 mediates mucosal host defense against Gram-negative bacterial pneumonia [J].

Aujla, Shean J. ;

Chan, Yvonne R. ;

Zheng, Mingquan ;

Fei, Mingjian ;

Askew, David J. ;

Pociask, Derek A. ;

Reinhart, Todd A. ;

McAllister, Florencia ;

Edeal, Jennifer ;

Gaus, Kristi ;

Husain, Shahid ;

Kreindler, James L. ;

Dubin, Patricia J. ;

Pilewski, Joseph M. ;

Myerburg, Mike M. ;

Mason, Carol A. ;

Iwakura, Yoichiro ;

Kolls, Jay K. .

NATURE MEDICINE, 2008, 14 (03) :275-281

[3] Investigating reference genes for quantitative real-time PCR analysis across four chicken tissues [J].

Bages, S. ;

Estany, J. ;

Tor, M. ;

Pena, R. N. .

GENE, 2015, 561 (01) :82-87

[4] Utility of the housekeeping genes 18S rRNA, β-actin and glyceraldehyde-3-phosphate-dehydrogenase for normalization in real-time quantitative reverse transcriptase-polymerase chain reaction analysis of gene expression in human T lymphocytes [J].

Bas, A ;

Forsberg, G ;

Hammarström, S ;

Hammarström, ML .

SCANDINAVIAN JOURNAL OF IMMUNOLOGY, 2004, 59 (06) :566-573

[5] Comparison of quantitative real-time polymerase chain reaction with NanoString® methodology using adipose and liver tissues from rats fed seaweed [J].

Bentley-Hewitt, Kerry L. ;

Hedderley, Duncan I. ;

Monro, John ;

Martell, Sheridan ;

Smith, Hannah ;

Mishra, Suman .

NEW BIOTECHNOLOGY, 2016, 33 (03) :380-386

[6] Identification of stable reference genes for quantitative PCR in cells derived from chicken lymphoid organs [J].

Borowska, D. ;

Rothwell, L. ;

Bailey, R. A. ;

Watson, K. ;

Kaiser, P. .

VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY, 2016, 170 :20-24

[7] Highly multiplexed quantitative PCR-based platform for evaluation of chicken immune responses [J].

Borowska, Dominika ;

Kuo, Richard ;

Bailey, Richard A. ;

Watson, Kellie A. ;

Kaiser, Pete ;

Vervelde, Lonneke ;

Stevens, Mark P. .

PLOS ONE, 2019, 14 (12)

[8] Selection of reference genes for mRNA quantification in Trypanosoma brucei [J].

Brenndoerfer, Martin ;

Boshart, Michael .

MOLECULAR AND BIOCHEMICAL PARASITOLOGY, 2010, 172 (01) :52-55

[9] Inflammation: friend or foe for animal production? [J].

Broom, Leon J. ;

Kogut, Michael H. .

POULTRY SCIENCE, 2018, 97 (02) :510-514

[10] The role of interleukin-22 in hepatitis C virus infection [J].

Dambacher, Julia ;

Beigel, Florian ;

Zitzmann, Kathrin ;

Heeg, Malte H. J. ;

Goeke, Burkhard ;

Diepolder, Helmut M. ;

Auernhammer, Christoph J. ;

Brand, Stephan .

CYTOKINE, 2008, 41 (03) :209-216

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