Measurable impact of RNA quality on gene expression results from quantitative PCR.
Measurable impact of RNA quality on gene expression results from quantitative PCR.
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DOI：
10.1093/nar/gkr065
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发表时间：
2011-05
影响因子：
14.9
通讯作者：
Vandesompele J
中科院分区：
文献类型：
--
作者：
Vermeulen J;De Preter K;Lefever S;Nuytens J;De Vloed F;Derveaux S;Hellemans J;Speleman F;Vandesompele J
受损的RNA质量被认为会导致基因表达研究中的不可靠结果。因此，在将样品纳入分析管道之前，认为对RNA完整性和纯度的评估至关重要。当诊断、预后或治疗结论依赖于此类分析时，这可能特别重要。在这项研究中，使用740个原发性肿瘤样本的大面板，实时定量PCR基因表达结果可用，确定了6个RNA质量参数的比较值。检测参数包括基于微流控毛细管电泳的18 S/28 SrRNA比值和RNA质量指数值、基于5′/3′比值mRNA完整性分析的HPRT 1 5′-3′定量循环差（Cq）和HPRT 1 3′ Cq值、表达的Alu重复序列的Cq值以及基于4个参考基因平均表达水平的归一化因子。在建立了评估RNA质量影响的创新分析框架后，我们观察到RNA质量对参考基因变异、两个癌症患者风险组之间预后标记基因差异表达的意义以及风险分类的可测量影响使用多基因签名。该研究为进一步合理评估基于逆转录定量PCR的结果与RNA质量的关系奠定了基础。
Compromised RNA quality is suggested to lead to unreliable results in gene expression studies. Therefore, assessment of RNA integrity and purity is deemed essential prior to including samples in the analytical pipeline. This may be of particular importance when diagnostic, prognostic or therapeutic conclusions depend on such analyses. In this study, the comparative value of six RNA quality parameters was determined using a large panel of 740 primary tumour samples for which real-time quantitative PCR gene expression results were available. The tested parameters comprise of microfluidic capillary electrophoresis based 18S/28S rRNA ratio and RNA Quality Index value, HPRT1 5′–3′ difference in quantification cycle (Cq) and HPRT1 3′ Cq value based on a 5′/3′ ratio mRNA integrity assay, the Cq value of expressed Alu repeat sequences and a normalization factor based on the mean expression level of four reference genes. Upon establishment of an innovative analytical framework to assess impact of RNA quality, we observed a measurable impact of RNA quality on the variation of the reference genes, on the significance of differential expression of prognostic marker genes between two cancer patient risk groups, and on risk classification performance using a multigene signature. This study forms the basis for further rational assessment of reverse transcription quantitative PCR based results in relation to RNA quality.
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影响因子：
14.9
作者：
Imbeaud S;Graudens E;Boulanger V;Barlet X;Zaborski P;Eveno E;Mueller O;Schroeder A;Auffray C
通讯作者：
Auffray C
影响因子：
2.7
作者：
Stangegaard, Michael;Dufva, Inge Hagh;Dufva, Martin
通讯作者：
Dufva, Martin
RTPRIMERDB：实时PCR引物和探针的门户。
影响因子：
14.9
作者：
Lefever S;Vandesompele J;Speleman F;Pattyn F
通讯作者：
Pattyn F
影响因子：
158.5
作者：
Liu, Rui;Wang, Xinhao;Clarke, Michael F.
通讯作者：
Clarke, Michael F.
影响因子：
5
作者：
Antonov, J;Goldstein, DR;Jaggi, R
通讯作者：
Jaggi, R