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Vol 45(2011) N 5 p. 797-803; M.A. Emelyanova1, F.A. Amosenko2, A.V. Chudinov1, S.A. Surzhikov1, T.P. Kazubskaya3, L.N. Lyubchenko3, T.V. Nasedkina1* Detection of KRAS Mutations in Tumor Cells Using Biochips 1Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, 119991 Russia2Research Centre for Medical Genetics, Russian Academy of Medical Sciences, Moscow, 115478 Russia 3Blokhin Cancer Research Center, Russian Academy of Medical Sciences, Moscow, 115478 Russia *nased@eimb.ru Received - 2010-12-08; Accepted - 2011-01-20 Somatic mutations in the KRAS gene are important markers of some types of tumors, for example, pancreatic cancer, and may be useful in early diagnostics. A biochip has been developed which allows determining most frequent mutations in 12, 13, and 61 codons of the KRAS gene. To increase the sensitivity of the method and to enable the analysis of minor fractions of tumor cells in clinical samples, the method of blocking wild type sequence PCR amplification by LNA-oligonucleotides has been used. The product of LNA-clamp PCR was further hybridized with oligonucleotide probes, immobilized on the biochip. The biochip was tested with 42 clinical DNA samples from patients with pancreatic cancer, mostly duct adenocarcinomas. As reference methods, RFLP analysis and sequencing were used. The developed approach allows detecting somatic mutations in the KRAS gene if the portion of tumor cells with mutation is at least 1% of the whole cell population. somatic mutations, tumor, KRAS gene, biochips |
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