Fluorescence-based DHPLC for allelic quantification by single-nucleotide primer extension

Kenjiro Kosaki, Hiroshi Yoshihashi, Yuko Ohashi, Rika Kosaki, Taichi Suzuki, Nobutake Matsuo

研究成果: Article

7 引用 (Scopus)

抄録

We have investigated the possibility of determining quantitatively the alleles of binary DNA polymorphisms by single-nucleotide primer extension (SNuPE) and fluorescence-based DHPLC. Using a polymorphism of interest to our group, ROX-labeled dideoxy CTP (ROX-ddCTP) was incorporated at the 3′ end of the primer annealed to the template adjacent to the polymorphic site. The primer extension product was then resolved from the unincorporated dye terminator by ion-pair reversed-phase liquid chromatography. The signal intensity of incorporated ROX-ddCTP correlated well over one order of magnitude with the relative amount of the C-allele present in the genomic DNA template. We conclude that SNuPE, when combined with fluorescence-based DHPLC, can accurately determine the relative molar proportion of one allele in total DNA.

元の言語English
ページ(範囲)111-119
ページ数9
ジャーナルJournal of Biochemical and Biophysical Methods
47
発行部数1-2
DOI
出版物ステータスPublished - 2001 1 30

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Cytidine Triphosphate
Nucleotides
Fluorescence
Alleles
Polymorphism
DNA
Liquid chromatography
Reverse-Phase Chromatography
Single Nucleotide Polymorphism
Coloring Agents
Ions

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics

これを引用

Fluorescence-based DHPLC for allelic quantification by single-nucleotide primer extension. / Kosaki, Kenjiro; Yoshihashi, Hiroshi; Ohashi, Yuko; Kosaki, Rika; Suzuki, Taichi; Matsuo, Nobutake.

:: Journal of Biochemical and Biophysical Methods, 巻 47, 番号 1-2, 30.01.2001, p. 111-119.

研究成果: Article

Kosaki, Kenjiro ; Yoshihashi, Hiroshi ; Ohashi, Yuko ; Kosaki, Rika ; Suzuki, Taichi ; Matsuo, Nobutake. / Fluorescence-based DHPLC for allelic quantification by single-nucleotide primer extension. :: Journal of Biochemical and Biophysical Methods. 2001 ; 巻 47, 番号 1-2. pp. 111-119.
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