Animal Science, School of Environmental and Rural Science, University of New England, Armidale, New South Wales, 2351, Australia.
School of Animal and Veterinary Sciences, The University of Adelaide, Roseworthy, South Australia, 5371, Australia.
Sci Rep. 2017 Oct 27;7(1):14271. doi: 10.1038/s41598-017-14693-2.
Ten reference genes were investigated for normalisation of candidate target gene expression data in the shell gland and spleen of laying hens challenged with two strains of infectious bronchitis virus (IBV). Data were analysed with geNorm, NormFinder and BestKeeper, and a comprehensive ranking (geomean) was calculated. In the combined data set of IBV challenged shell gland samples, the comprehensive ranking showed TATA-box binding protein (TBP) and tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein zeta (YWHAZ) as the two most stable, and succinate dehydrogenase complex flavoprotein subunit A (SDHA) and albumin (ALB) as the two least stable reference genes. In the spleen, and in the combined data set of the shell gland and spleen, the two most stable and the two least stable reference genes were TBP and YWHAZ, and ribosomal protein L4 (RPL4) and ALB, respectively. Different ranking has been due to different algorithms. Validation studies showed that the use of the two most stable reference genes produced accurate and more robust gene expression data. The two most and least stable reference genes obtained in the study, were further used for candidate target gene expression data normalisation of the shell gland and spleen under an IBV infection model.
研究了 10 个参考基因，以规范感染两种传染性支气管炎病毒（IBV）的蛋鸡泄殖腔和脾脏中候选靶基因表达数据。使用 geNorm、NormFinder 和 BestKeeper 对数据进行了分析，并计算了综合排名（几何平均值）。在 IBV 挑战的泄殖腔样本的综合数据集，综合排名显示 TATA 框结合蛋白（TBP）和酪氨酸 3-单加氧酶/色氨酸 5-单加氧酶激活蛋白 zeta（YWHAZ）是最稳定的两个，琥珀酸脱氢酶复合物黄素蛋白亚单位 A（SDHA）和白蛋白（ALB）是最不稳定的两个。在脾脏和泄殖腔和脾脏的综合数据集，最稳定的两个和最不稳定的两个参考基因分别为 TBP 和 YWHAZ，核糖体蛋白 L4（RPL4）和 ALB。不同的排名是由于不同的算法。验证研究表明，使用两个最稳定的参考基因可以产生准确且更稳健的基因表达数据。在研究中获得的两个最稳定和最不稳定的参考基因，进一步用于 IBV 感染模型下泄殖腔和脾脏候选靶基因表达数据的正常化。
