Validation of real-time RT-PCR assays for mRNA quantification in baboons
Validation of real-time RT-PCR assays for mRNA quantification in baboons
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DOI：
10.1016/j.cyto.2005.07.002
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发表时间：
2005-09-21
期刊：
影响因子：
3.8
通讯作者：
D'Hooghe, T
中科院分区：
文献类型：
--
作者：
Overbergh, L;Kyama, CM;D'Hooghe, T
实时逆转录聚合酶链式反应已被广泛应用于基础研究和临床诊断，例如用于定量测定人体组织和组织活检中的RNA水平。在本研究中，我们提供了一种验证用于实时RT-PCR定量检测狒狒样本的引物/探针的策略。该方法以TaqMan系统为基础，使用已为人类实时RT-PCR设计和验证的引物/探针。这一策略的准确性的一个先决条件是人和狒狒的PCR反应具有相似的扩增效率。我们提出了两种不同的方法，即根据稀释曲线的斜率计算PCR效率或使用线性回归方法来比较人和狒狒样本的扩增效率。综上所述，通过一个简单的验证实验，基于人类序列的实时聚合酶链式反应分析方法可以很容易地应用于狒狒样本的分析。(C)2005爱思唯尔有限公司。保留所有权利。
Real-time RT-PCR has been used widely, both in fundamental research and in clinical diagnostics, for instance for quantification of RNA levels in human tissues and tissue biopsies. In the present study we provide a strategy to validate primers/probes for realtime RT-PCR quantification of baboon samples. The method is based on the TaqMan system and uses primers/probes that have been designed and validated for human real-time RT-PCR. A prerequisite for the accuracy of this strategy is a similar amplification efficiency between human and baboon PCR reactions. We propose two different methods, i.e. by calculating PCR efficiencies from the slope of a dilution curve or by using the linear regression method, to compare the amplification efficiency between human and baboon samples. In conclusion, by performing a simple validation experiment, real-time PCR assays based on human sequences, which are easily available, can be applied for analysis of baboon samples. (c) 2005 Elsevier Ltd. All rights reserved.