A GeNorm algorithm-based selection of reference genes for quantitative real-time PCR in skin biopsies of healthy dogs and dogs with atopic dermatitis
A GeNorm algorithm-based selection of reference genes for quantitative real-time PCR in skin biopsies of healthy dogs and dogs with atopic dermatitis
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DOI：
10.1016/j.vetimm.2008.12.004
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发表时间：
2009-05-15
影响因子：
1.8
通讯作者：
Penning, Louis C.
中科院分区：
文献类型：
--
作者：
Schlotter, Yvette M.;Veenhof, Eveline Z.;Penning, Louis C.
定量真实的时间PCR（Q-PCR）是研究mRNA表达水平的首选方法。由于Q-PCR非常灵敏，因此使用稳定表达的参考基因对数据进行标准化至关重要。参考基因的稳定性取决于组织和感兴趣的物种。因此，必须对研究中的每个新组织和种属进行参考基因稳定性评价。使用GeNorm软件在速冻犬皮肤活检中分析B2M、GAPDH、HPRT、SRPR、hnRNPH、GUSB、RPL 8、RPS 5和RPS 19的稳定性。包括健康犬（n = 7）和确诊特应性皮炎的犬（n = 28）。分析了病变和非病变皮肤。研究表明，除GUSB和HPRT外，最适合作为犬皮肤参考基因的是核糖体基因产物RPL 8、RPS5和RPS19。少至三个参考基因将揭示高度可靠的Q-PCR计算。(c)2008 Elsevier B.V.保留所有权利。
Quantitative real time PCR (Q-PCR) is the method of choice to study mRNA expression levels. Since Q-PCR is very sensitive, normalization of the data with stably expressed reference genes if of utmost importance. The stability of reference genes depends on the tissue and the species of interest. Therefore, evaluation of the stability of reference genes must be performed for each new tissue and species under study. The stability of B2M, GAPDH, HPRT, SRPR, hnRNPH, GUSB, RPL8, RPS5, and RPS19 was analyzed with the GeNorm software in snap frozen canine skin biopsies. Healthy dogs (n = 7) and dogs with confirmed atopic dermatitis (n = 28) were included. Lesional and non-lesional skin was analyzed. The study indicated that the most appropriate reference genes in canine skin are the ribosomal gene products RPL8, RPS5 and RPS19 besides GUSB and HPRT. As little as three reference genes will reveal highly reliable Q-PCR calculations. (c) 2008 Elsevier B.V. All rights reserved.