Effects of RNA degradation on gene expression analysis of human postmortem tissues
Effects of RNA degradation on gene expression analysis of human postmortem tissues
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DOI：
10.1096/fj.04-3552fje
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发表时间：
2005-06-01
期刊：
影响因子：
4.8
通讯作者：
Hevezi, P
中科院分区：
文献类型：
--
作者：
Lee, J;Hever, A;Hevezi, P
利用Affymetrix GeneChip平台构建正常人体基因表达数据库。死后人体组织是此类研究的生物材料的宝贵来源，但在收获此类组织之前，它们的使用需要一些延迟。我们首先评估了从死后 3-5 小时获得的组织中获得的 RNA 质量，发现了组织和供体依赖性的变异。从大脑区域提取的 RNA 质量高于从肠道提取的 RNA，而死亡原因是供体依赖性差异的一个重要因素。为了避免这些变量，我们使用大鼠十二指肠来确定 RNA 降解对基因表达分析的影响。令人惊讶的是，即使是表现出显着 RNA 降解的样品也产生了稳健的基因表达结果，与在特定信号强度截止条件下使用完整样品获得的结果相当。我们将这些发现扩展到我们的人类表达数据库并获得了类似的结果，这表明偏向转录本 3' 端进行检测的 Affymetrix 平台可以容忍显着的 RNA 降解，同时仍然产生高质量的表达数据。我们生成的身体指数表达数据库是一个有价值的研究工具。作为潜在用途的例子，我们报告了四个潜在治疗靶点——CCL27、GPR22、GPR113和GPR128——以及一组胸腺特异性基因的新表达位点，其中包括三个以前与胸腺无关的基因。
The Affymetrix GeneChip platform was used to build a gene expression database of the normal human body. Postmortem human tissues represent a valuable source of biological materials for this type of study, but their use entails some delays before harvesting such tissues. We first evaluated the RNA quality obtained from tissues obtained 3-5 h postmortem and found variations that were both tissue and donor-dependent. RNAs extracted from brain regions were of higher quality than those obtained from the gut, while the cause of death was a significant factor in donor-dependent differences. To avoid these variables, we used rat duodenum to determine the effects of RNA degradation on the analysis of gene expression. Surprisingly, even samples exhibiting significant RNA degradation yielded robust gene expression results, comparable to those obtained using intact samples at a certain signal intensity cutoff. We extended these findings to our human expression database and obtained similar results, indicating that the Affymetrix platform, which is biased to the 3' end of transcripts for detection, can tolerate significant RNA degradation, while still yielding high quality expression data. Our resulting body index expression database is a valuable research tool. As examples of potential uses, we report novel expression sites for four potential therapeutic targets-CCL27, GPR22, GPR113 and GPR128-and as well as a set of thymus-specific genes, including three not previously associated with the thymus.