Selection of reference genes for quantitative real-time PCR analysis in canine mammary tumors using the GeNorm algorithm
Selection of reference genes for quantitative real-time PCR analysis in canine mammary tumors using the GeNorm algorithm
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DOI：
10.1354/vp.43-6-934
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发表时间：
2006-11-01
影响因子：
2.4
通讯作者：
Sterner-Kock, A.
中科院分区：
文献类型：
--
作者：
Etschmann, B.;Wilcken, B.;Sterner-Kock, A.
11 个参考基因（18s 核糖体核糖核酸 [RNA]、28s 核糖体 RNA、泛素、β-肌动蛋白、甘油醛脱氢酶、ATP 合酶亚基 5B、羟甲基双烷合酶、次黄嘌呤磷酸核糖基转移酶、核糖体蛋白 L32、色氨酸 5-单加氧酶激活蛋白 (zeta使用定量逆转录聚合酶链反应（qRT-PCR）对犬乳腺肿瘤中的基因表达谱实验进行分析，作为基因表达谱实验的参考。在 22 个来自乳腺组织的组织学特征切除肿瘤样本和来自同一个体的 22 个非肿瘤性乳腺组织样本中测量了候选者的转录水平。结果用于使用 GeNorm 工具对候选参考基因进行排名。经确定，在犬乳腺组织样本中，次黄嘌呤磷酸核糖基转移酶、ATP 合酶亚基 5B、核糖体蛋白 L32 和泛素的组合可产生稳定的参考基因表达水平，而甘油醛脱氢酶或核糖体 RNA 的使用不适合对该组织类型中的 qRT-PCR 结果进行标准化。
Eleven reference genes (18s ribosomal ribonucleic acid [RNA], 28s ribosomal RNA, ubiquitin, beta-actin, glycerine aldehyde dehydrogenase, ATP-synthase subunit 5B, hydroxymethylbilane synthase, hypoxanthine-phosphoribosyl transferase, ribosomal protein L32, tryptophan 5-monooxygenase activation protein (zeta polypeptide), and TATA-Box binding protein) were analyzed in use as references for gene expression profiling experiments using quantitative reverse transcription polymerase chain reaction (qRT-PCR) in canine mammary tumors. The transcription level of the candidates was measured in 22 histologically characterized excised tumor specimens from mammary gland tissue and 22 samples of non-neoplastic mammary tissue samples from the same individuals. Results were used to rank candidate reference genes using the GeNorm tool. It was determined that in samples of canine mammary gland tissue, a combination of hypoxanthine-phosphoribosyl transferase, ATP-synthase subunit 5B, ribosomal protein L32 and ubiquitin yields stable reference gene expression levels, whereas the use of glycerin aldehyde dehydrogenase or ribosomal RNA is unsuitable for normalization of qRT-PCR results in this tissue type.