Identification of mutations by RNA conformational polymorphism "bar code" analysis.
Identification of mutations by RNA conformational polymorphism "bar code" analysis.
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通过RNA构象多态性“条形码”分析鉴定突变。
DOI：
10.1006/geno.1995.0024
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Danenberg,PV
中科院分区：
文献类型：
--
作者：
Lenz,HJ;Danenberg,KD;Schnieders,B;Banerjee,D;Bertino,JR;Leichman,L;Danenberg,PV
DNA单链构象多态性（SSCP）分析被广泛用于检测临床标本中的点突变。用cRNA代替DNA进行SSCP分析已被证明可以提高突变检测频率。RNA可以以多种亚稳定构象存在，其在非变性电泳凝胶上显示为条带模式。单碱基突变不仅可以引起主要条带的迁移率变化，而且可以导致某些构象的丢失和新构象的出现。在许多情况下，与特定碱基序列相关的独特RNA SSCP模式允许视觉识别点突变。然而，在某些情况下，序列中单个碱基变化的RNA SSCP模式对于突变的阳性鉴定来说没有足够的不同。通过在转录反应中加入3′-脱氧核苷酸，提高了RNA SSCP的检测能力。转录反应中链终止核苷酸的存在形成了许多新的RNA片段，从而产生了每个RNA序列所特有的复杂带型（“条形码”）。应用该方法分析了结肠癌患者的p53突变。
DNA single-strand conformational polymorphism (SSCP) analysis is widely used for detection of point mutations in clinical specimens. Performing SSCP analysis with cRNA instead of DNA has been shown to improve mutation detection frequency. RNA can exist in numerous meta-stable conformations, which appear as patterns of bands on nondenaturing electrophoresis gels. Single base mutations can cause not only mobility shifts of major bands, but also loss of some conformations and appearance of new conformations. Unique RNA SSCP patterns associated with specific base sequences in many cases allow visual identification of point mutations. However, in some cases, the RNA SSCP pattern of a single base change in a sequence is not sufficiently different for a positive identification of the mutation. Improvement in the detection capability of RNA SSCP was obtained by adding 3′-deoxynucleotides to the transcription reaction. The presence of chain-terminating nucleotides in the transcription reaction formed numerous new RNA fragments, thereby generating complex band patterns (“bar codes”) unique to each RNA sequence. This method was applied to analyzing p53 mutations in patients with colon cancer.