Tõnisson Neeme, Zernant Jana, Kurg Ants, Pavel Hendrik, Slavin Georg, Roomere Hanno, Meiel Aune, Hainaut Pierre, Metspalu Andres
Asper, Ltd., 3 Oru Street, 51014 Tartu, Estonia.
Proc Natl Acad Sci U S A. 2002 Apr 16;99(8):5503-8. doi: 10.1073/pnas.082100599.
Identification of mutations in the tumor suppressor gene TP53 has implications for the molecular epidemiology and for the molecular pathology of human cancer. We have developed and evaluated an arrayed primer extension assay for covering both strands of a region of the coding sequence containing more than 95% of the mutations described so far in TP53. On average, 97.5% of the arrayed TP53 gene sequence can be analyzed from either sense or antisense strands, and 81% from both strands. A patient DNA sample is amplified and annealed to arrayed primers, which then promote DNA polymerase extension reactions with four fluorescently labeled dideoxynucleotides. The TP53 gene chip spans exons 2-9 plus two introns from both strands. The performance of the assay was evaluated by using freshly extracted genomic DNA, as well as DNA extracted from archival (paraffin-embedded) DNA samples. The arrayed primer extension-based TP53 gene test provides an accurate and efficient tool for DNA sequence analysis of this frequently mutated gene for both research and clinical applications.
肿瘤抑制基因TP53突变的鉴定对人类癌症的分子流行病学和分子病理学具有重要意义。我们开发并评估了一种阵列引物延伸检测方法，用于覆盖编码序列区域的两条链，该区域包含了迄今为止TP53中描述的95%以上的突变。平均而言，从正义链或反义链可分析阵列TP53基因序列的97.5%，从两条链可分析81%。患者DNA样本经扩增后与阵列引物退火，然后引物促进DNA聚合酶与四种荧光标记的双脱氧核苷酸发生延伸反应。TP53基因芯片跨越外显子2至9以及两条链的两个内含子。通过使用新鲜提取的基因组DNA以及从存档（石蜡包埋）DNA样本中提取的DNA来评估该检测方法的性能。基于阵列引物延伸的TP53基因检测为该频繁突变基因的DNA序列分析提供了一种准确且高效的工具，可用于研究和临床应用。
