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Sensitive detection of low-abundance in-frame deletions in EGFR exon 19 using novel wild-type blockers in real-time PCR.
Ren, Xiao-Dong; Liu, Ding-Yuan; Guo, Hai-Qin; Wang, Liu; Zhao, Na; Su, Ning; Wei, Kun; Ren, Sai; Qu, Xue-Mei; Dai, Xiao-Tian; Huang, Qing.
Afiliação
  • Ren XD; Department of Laboratory Medicine, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, 400038, P.R. China.
  • Liu DY; Department of Laboratory Medicine, Institute of Surgery Research, Daping Hospital, Third Military Medical University (Army Medical University), Chongqing, 400042, P.R. China.
  • Guo HQ; Department of Pulmonology, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, 400038, P.R. China.
  • Wang L; Department of Pulmonology, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, 400038, P.R. China.
  • Zhao N; Department of Laboratory Medicine, Institute of Surgery Research, Daping Hospital, Third Military Medical University (Army Medical University), Chongqing, 400042, P.R. China.
  • Su N; Department of Laboratory Medicine, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, 400038, P.R. China.
  • Wei K; Department of Laboratory Medicine, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, 400038, P.R. China.
  • Ren S; Department of Laboratory Medicine, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, 400038, P.R. China.
  • Qu XM; Department of Laboratory Medicine, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, 400038, P.R. China.
  • Dai XT; Department of Laboratory Medicine, Institute of Surgery Research, Daping Hospital, Third Military Medical University (Army Medical University), Chongqing, 400042, P.R. China.
  • Huang Q; Department of Pulmonology, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, 400038, P.R. China. xtdai2013@163.com.
Sci Rep ; 9(1): 8276, 2019 06 04.
Article em En | MEDLINE | ID: mdl-31164704
ABSTRACT
Epidermal growth factor receptor (EGFR) mutations are associated with response of tyrosine kinase inhibitors (TKIs) for patients with advanced non-small cell lung cancer (NSCLC). However, the existing methods for detection of samples having rare mutations(i.e. ~0.01%) have limits in terms of specificity, time consumption or cost. In the current study, novel wild-type blocking (WTB) oligonucleotides modified with phosphorothioate or inverted dT at the 5'-termini were designed to precisely detect 11 common deletion mutations in exon 19 of EGFR gene (E19del) using a WTB-PCR assay. And internal competitive leptin amplifications were further applied to enhance the specificity of the WTB-PCR system. Our results showed that WTB-PCR could completely block amplification of wild-type EGFR when 200 ng of DNA was used as template. Furthermore, the current WTB-PCR assay facilitated the detection of E19del mutations with a selectivity of 0.01% and sensitivity as low as a single copy. And, the results showed that the current WTB-PCR system exceeded detection limits afforded by the ARMS-PCR assay. In conclusion, the current WTB-PCR strategy represents a simple and cost-effective method to precisely detect various low-abundance deletion mutations.
Assuntos

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Oligonucleotídeos / Carcinoma Pulmonar de Células não Pequenas / Inibidores de Proteínas Quinases Tipo de estudo: Diagnostic_studies Limite: Adult / Aged / Aged80 / Female / Humans / Male / Middle aged Idioma: En Revista: Sci Rep Ano de publicação: 2019 Tipo de documento: Article País de publicação: ENGLAND / ESCOCIA / GB / GREAT BRITAIN / INGLATERRA / REINO UNIDO / SCOTLAND / UK / UNITED KINGDOM

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Oligonucleotídeos / Carcinoma Pulmonar de Células não Pequenas / Inibidores de Proteínas Quinases Tipo de estudo: Diagnostic_studies Limite: Adult / Aged / Aged80 / Female / Humans / Male / Middle aged Idioma: En Revista: Sci Rep Ano de publicação: 2019 Tipo de documento: Article País de publicação: ENGLAND / ESCOCIA / GB / GREAT BRITAIN / INGLATERRA / REINO UNIDO / SCOTLAND / UK / UNITED KINGDOM