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Molecular Biology

, Volume 52, Issue 6, pp 865–871 | Cite as

Multiplex Assay to Evaluate the Genetic Risk of Developing Human Melanoma

  • D. O. Fesenko
  • I. S. Abramov
  • V. E. Shershov
  • V. E. Kuznetsova
  • S. A. Surzhikov
  • I. V. Grechishnikova
  • V. E. Barsky
  • A. V. Chudinov
  • T. V. Nasedkina
GENOMICS. TRANSCRIPTOMICS
  • 9 Downloads

Abstract

A genotyping procedure based on single-step PCR and subsequent allele-specific hybridization on a hydrogel biochip was developed to address the polymorphisms of HERC2, OCA2, SLC24A4, SLC45A2, TYR, IRF4, MC1R, MITF, PIGU, MYH7B, NCOA6, and CDK10. Amplified gene fragments were fluorescently labeled in PCR, and fluorescent signals from biochip cells were detected to evaluate how efficiently the PCR product formed a perfect duplex with an immobilized probe. The analytical characteristics of hybridization analysis were estimated for several fluorophores with different optical spectra. Cyanine dyes fluorescing in the range of Cy5 and Cy7 were synthesized for the purpose and used as 5'-tags of universal primers in single-step PCR. A Cy7 analog fluorescing in the near infrared range was found to increase the sensitivity of hybridization analysis by producing a lower background signal in the cases where target gene amplification was low.

Keywords:

melanoma susceptibility genotyping biochip single nucleotide polymorphism cyanine dye 

Notes

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Copyright information

© Pleiades Publishing, Inc. 2018

Authors and Affiliations

  • D. O. Fesenko
    • 1
  • I. S. Abramov
    • 1
  • V. E. Shershov
    • 1
  • V. E. Kuznetsova
    • 1
  • S. A. Surzhikov
    • 1
  • I. V. Grechishnikova
    • 1
  • V. E. Barsky
    • 1
  • A. V. Chudinov
    • 1
  • T. V. Nasedkina
    • 1
  1. 1.Engelhardt Institute of Molecular Genetics, Russian Academy of SciencesMoscowRussia

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