Solid-Phase Hybridization Assay for Detection of Mutated Cancer DNA by Fluorescence

Maria Taskova, Kira Astakhova*

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingBook chapterResearchpeer-review

Abstract

We report a straightforward protocol for the detection of mutated DNA extracted from cancer cells. The assay combines a step-wise solid-phase hybridization and a readout by fluorescence emission. We detect a single-nucleotide polymorphism in two human oncogenes, BRAF and EGFR, and reach a limit of the detection of 300 pM by conventional fluorometry. The protocol described herein may be used as a foundation for development of automatic optimized assays capable for detection of mutant DNA and RNA in vitro and in cells.
Original languageEnglish
Title of host publicationNucleic Acid Detection and Structural Investigations : Methods and Protocols
EditorsKira Astakhova , Syeda Atia Bukhari
Number of pages8
Volume2063
PublisherSpringer
Publication date2020
Pages37-44
Chapter4
ISBN (Print)978-1-0716-0137-2
ISBN (Electronic)978-1-0716-0138-9
DOIs
Publication statusPublished - 2020
SeriesMethods in Molecular Biology
ISSN1064-3745

Keywords

  • DNA oncogene
  • Fluorescent oligonucleotides
  • Hybridization assay
  • Fluorometry

Cite this

Taskova, M., & Astakhova, K. (2020). Solid-Phase Hybridization Assay for Detection of Mutated Cancer DNA by Fluorescence. In K. A., & S. A. B. (Eds.), Nucleic Acid Detection and Structural Investigations: Methods and Protocols (Vol. 2063, pp. 37-44). Springer. Methods in Molecular Biology https://doi.org/10.1007/978-1-0716-0138-9_4
Taskova, Maria ; Astakhova, Kira. / Solid-Phase Hybridization Assay for Detection of Mutated Cancer DNA by Fluorescence. Nucleic Acid Detection and Structural Investigations: Methods and Protocols. editor / Kira Astakhova ; Syeda Atia Bukhari. Vol. 2063 Springer, 2020. pp. 37-44 (Methods in Molecular Biology).
@inbook{99175b483fb74897bc3b709e3f34591e,
title = "Solid-Phase Hybridization Assay for Detection of Mutated Cancer DNA by Fluorescence",
abstract = "We report a straightforward protocol for the detection of mutated DNA extracted from cancer cells. The assay combines a step-wise solid-phase hybridization and a readout by fluorescence emission. We detect a single-nucleotide polymorphism in two human oncogenes, BRAF and EGFR, and reach a limit of the detection of 300 pM by conventional fluorometry. The protocol described herein may be used as a foundation for development of automatic optimized assays capable for detection of mutant DNA and RNA in vitro and in cells.",
keywords = "DNA oncogene, Fluorescent oligonucleotides, Hybridization assay, Fluorometry",
author = "Maria Taskova and Kira Astakhova",
year = "2020",
doi = "10.1007/978-1-0716-0138-9_4",
language = "English",
isbn = "978-1-0716-0137-2",
volume = "2063",
pages = "37--44",
editor = "{Kira Astakhova} and {Syeda Atia Bukhari}",
booktitle = "Nucleic Acid Detection and Structural Investigations",
publisher = "Springer",

}

Taskova, M & Astakhova, K 2020, Solid-Phase Hybridization Assay for Detection of Mutated Cancer DNA by Fluorescence. in KA & SAB (eds), Nucleic Acid Detection and Structural Investigations: Methods and Protocols. vol. 2063, Springer, Methods in Molecular Biology, pp. 37-44. https://doi.org/10.1007/978-1-0716-0138-9_4

Solid-Phase Hybridization Assay for Detection of Mutated Cancer DNA by Fluorescence. / Taskova, Maria; Astakhova, Kira.

Nucleic Acid Detection and Structural Investigations: Methods and Protocols. ed. / Kira Astakhova; Syeda Atia Bukhari. Vol. 2063 Springer, 2020. p. 37-44 (Methods in Molecular Biology).

Research output: Chapter in Book/Report/Conference proceedingBook chapterResearchpeer-review

TY - CHAP

T1 - Solid-Phase Hybridization Assay for Detection of Mutated Cancer DNA by Fluorescence

AU - Taskova, Maria

AU - Astakhova, Kira

PY - 2020

Y1 - 2020

N2 - We report a straightforward protocol for the detection of mutated DNA extracted from cancer cells. The assay combines a step-wise solid-phase hybridization and a readout by fluorescence emission. We detect a single-nucleotide polymorphism in two human oncogenes, BRAF and EGFR, and reach a limit of the detection of 300 pM by conventional fluorometry. The protocol described herein may be used as a foundation for development of automatic optimized assays capable for detection of mutant DNA and RNA in vitro and in cells.

AB - We report a straightforward protocol for the detection of mutated DNA extracted from cancer cells. The assay combines a step-wise solid-phase hybridization and a readout by fluorescence emission. We detect a single-nucleotide polymorphism in two human oncogenes, BRAF and EGFR, and reach a limit of the detection of 300 pM by conventional fluorometry. The protocol described herein may be used as a foundation for development of automatic optimized assays capable for detection of mutant DNA and RNA in vitro and in cells.

KW - DNA oncogene

KW - Fluorescent oligonucleotides

KW - Hybridization assay

KW - Fluorometry

U2 - 10.1007/978-1-0716-0138-9_4

DO - 10.1007/978-1-0716-0138-9_4

M3 - Book chapter

SN - 978-1-0716-0137-2

VL - 2063

SP - 37

EP - 44

BT - Nucleic Acid Detection and Structural Investigations

A2 - , Kira Astakhova

A2 - , Syeda Atia Bukhari

PB - Springer

ER -

Taskova M, Astakhova K. Solid-Phase Hybridization Assay for Detection of Mutated Cancer DNA by Fluorescence. In KA, SAB, editors, Nucleic Acid Detection and Structural Investigations: Methods and Protocols. Vol. 2063. Springer. 2020. p. 37-44. (Methods in Molecular Biology). https://doi.org/10.1007/978-1-0716-0138-9_4