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首页> 外文期刊>BMC Medical Genomics >Expression profiling with RNA from formalin-fixed, paraffin-embedded material
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Expression profiling with RNA from formalin-fixed, paraffin-embedded material

机译：用福尔马林固定的石蜡包埋材料中的RNA进行表达谱分析

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Background Molecular characterization of breast and other cancers by gene expression profiling has corroborated existing classifications and revealed novel subtypes. Most profiling studies are based on fresh frozen (FF) tumor material which is available only for a limited number of samples while thousands of tumor samples exist as formalin-fixed, paraffin-embedded (FFPE) blocks. Unfortunately, RNA derived of FFPE material is fragmented and chemically modified impairing expression measurements by standard procedures. Robust protocols for isolation of RNA from FFPE material suitable for stable and reproducible measurement of gene expression (e.g. by quantitative reverse transcriptase PCR, QPCR) remain a major challenge. Results We present a simple procedure for RNA isolation from FFPE material of diagnostic samples. The RNA is suitable for expression measurement by QPCR when used in combination with an optimized cDNA synthesis protocol and TaqMan assays specific for short amplicons. The FFPE derived RNA was compared to intact RNA isolated from the same tumors. Preliminary scores were computed from genes related to the ER response, HER2 signaling and proliferation. Correlation coefficients between intact and partially fragmented RNA from FFPE material were 0.83 to 0.97. Conclusion We developed a simple and robust method for isolating RNA from FFPE material. The RNA can be used for gene expression profiling. Expression measurements from several genes can be combined to robust scores representing the hormonal or the proliferation status of the tumor.
机译：背景技术通过基因表达谱分析对乳腺癌和其他癌症进行分子表征已证实了现有分类，并揭示了新的亚型。大多数分析研究均基于新鲜的冷冻（FF）肿瘤材料，该材料仅可用于有限数量的样品，而成千上万的肿瘤样品以福尔马林固定，石蜡包埋（FFPE）块存在。不幸的是，FFPE材料衍生的RNA被片段化，并通过标准程序化学修饰损害了表达测量。从FFPE材料中分离RNA的可靠方案（例如通过定量逆转录酶PCR，QPCR）适用于稳定和可重复的基因表达测量仍然是主要挑战。结果我们提供了一种从诊断样品的FFPE材料中分离RNA的简单程序。与优化的cDNA合成方案和针对短扩增子的TaqMan测定法结合使用时，RNA适合通过QPCR进行表达测量。将FFPE衍生的RNA与从相同肿瘤中分离的完整RNA进行比较。根据与ER反应，HER2信号传导和增殖相关的基因计算出初步得分。来自FFPE材料的完整RNA和部分片段RNA的相关系数为0.83至0.97。结论我们开发了一种从FFPE材料中分离RNA的简单而可靠的方法。 RNA可用于基因表达谱分析。可以将来自多个基因的表达测量结果结合起来，得出代表肿瘤荷尔蒙或增殖状态的稳健评分。

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