Waha A, Rollbrocker B, Wiestler O D, von Deimling A
Institut für Neuropathologie, Universitätskliniken, Bonn, Germany.
Diagn Mol Pathol. 1996 Jun;5(2):147-50. doi: 10.1097/00019606-199606000-00010.
A differential polymerase chain reaction (PCR) protocol was established for semiquantitative, nonradioactive detection of gene amplification using a DNA sequencer. Oncogene fragments and control DNA sequences were simultaneously PCR-amplified using fluorescent-labelled primers. Analysis of the PCR products allowed quantitative assessment of gene copy numbers in this coamplification assay. Using this approach, we examined a series of 132 brain tumors for amplification of the epidermal growth factor receptor (EGFR) gene. The same set of tumors was also analyzed by Southern blotting and hybridization with a radiolabelled EGFR probe. Both methods yielded virtually identical results. This technique has a great potential for nonradioactive screening of large tumor panels of oncogene amplification.
建立了一种差异聚合酶链反应（PCR）方案，用于使用DNA测序仪进行基因扩增的半定量、非放射性检测。使用荧光标记引物同时对癌基因片段和对照DNA序列进行PCR扩增。对PCR产物的分析允许在这种共扩增试验中对基因拷贝数进行定量评估。使用这种方法，我们检测了一系列132例脑肿瘤中表皮生长因子受体（EGFR）基因的扩增情况。同一组肿瘤也通过Southern印迹法并用放射性标记的EGFR探针进行杂交分析。两种方法产生的结果几乎相同。该技术在对大量肿瘤样本进行癌基因扩增的非放射性筛查方面具有很大潜力。
