Kriegshäuser Gernot, Fabjani Gerhild, Ziegler Barbara, Zöchbauer-Müller Sabine, End Adelheid, Zeillinger Robert
ViennaLab Diagnostics GmbH, 1120 Vienna, Austria.
Int J Mol Sci. 2011;12(12):8530-8. doi: 10.3390/ijms12128530. Epub 2011 Nov 29.
This study is aimed at evaluating the potential of a biochip assay to sensitively detect KRAS mutation in DNA from non-small cell lung cancer (NSCLC) tissue samples. The assay covers 10 mutations in codons 12 and 13 of the KRAS gene, and is based on mutant-enriched PCR followed by reverse-hybridization of biotinylated amplification products to an array of sequence-specific probes immobilized on the tip of a rectangular plastic stick (biochip). Biochip hybridization identified 17 (21%) samples to carry a KRAS mutation of which 16 (33%) were adenocarcinomas and 1 (3%) was a squamous cell carcinoma. All mutations were confirmed by DNA sequencing. Using 10 ng of starting DNA, the biochip assay demonstrated a detection limit of 1% mutant sequence in a background of wild-type DNA. Our results suggest that the biochip assay is a sensitive alternative to protocols currently in use for KRAS mutation testing on limited quantity samples.
本研究旨在评估一种生物芯片检测法灵敏检测非小细胞肺癌（NSCLC）组织样本DNA中KRAS突变的潜力。该检测法涵盖KRAS基因第12和13密码子的10种突变，基于突变富集PCR，随后将生物素化的扩增产物与固定在矩形塑料棒尖端的一系列序列特异性探针进行反向杂交（生物芯片）。生物芯片杂交鉴定出17个（21%）样本携带KRAS突变，其中16个（33%）为腺癌，1个（3%）为鳞状细胞癌。所有突变均经DNA测序确认。使用10 ng起始DNA，生物芯片检测法在野生型DNA背景下显示出1%突变序列的检测限。我们的结果表明，对于有限数量样本的KRAS突变检测，生物芯片检测法是目前所用检测方案的一种灵敏替代方法。
