Publication: Highly sensitive EGFR mutation detection by specific amplification of mutant alleles
Issued Date
2014-02-01
Resource Type
ISSN
10960945
00144800
00144800
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2-s2.0-84892475210
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Mahidol University
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SCOPUS
Bibliographic Citation
Experimental and Molecular Pathology. Vol.96, No.1 (2014), 85-91
Suggested Citation
Nalin Leelatian, Pichpisith Boonchoo, Sitsom Wijitburaphat, Kanya Moolsuwan, Pattara Wongjaroen, Priyakorn Chinnasang, Komsan Anyamaneeratch, Ruchira Ruangchira-urai, Naravat Poungvarin Highly sensitive EGFR mutation detection by specific amplification of mutant alleles. Experimental and Molecular Pathology. Vol.96, No.1 (2014), 85-91. doi:10.1016/j.yexmp.2013.12.006 Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/33306
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Title
Highly sensitive EGFR mutation detection by specific amplification of mutant alleles
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Abstract
Mutations in the tyrosine kinase domain of the epidermal growth factor receptor (EGFR) gene predict benefit from tyrosine kinase inhibitors in patients suffering from non-small-cell lung cancer. In this study, we developed a fast, simple, cost-effective and highly sensitive assay for detection of five clinically important EGFR mutations in exon 19 (2235_2249del and 2236_2250del), exon 20 (C2369T) and exon 21 (T2573G and c.2573_2574 TG. >. GT). We designed EGFR mutation detection assays by combining allele-specific PCR amplification with the detection of SYBR Green I fluorescence, and optimized PCR conditions to specifically amplify mutant alleles. These one-step assays were able to detect the mutations at levels as low as 1.5 mutant copies in a DNA sample. Commercially available probe-based allele-specific PCR exhibited relatively poor performance when detecting very low copies of mutated DNA, especially in exon 19 and 20. Our assays offered dramatically less reagent cost than that of the commercial kit and generated results in less than 90. min after DNA extraction. These protocols can also be applied to conventional thermal cyclers followed by gel electrophoresis detection. © 2013 Elsevier Inc.