期刊
BIOSENSORS & BIOELECTRONICS
卷 36, 期 1, 页码 89-94出版社
ELSEVIER ADVANCED TECHNOLOGY
DOI: 10.1016/j.bios.2012.03.045
关键词
Single-nucleotide polymorphism; Colorimetric genotyping; Gap ligationreaction; Magnetic bead; Horseradish peroxidase
类别
资金
- A*STAR-ANR Program
Herein we report a new strategy for highly sensitive and selective colorimatric assay for genotyping of single-nucleotide polymorphisms (SNPs). It is based on the use of a specific gap ligation reaction, horseradish peroxidase (HRP) for signal amplification, and magnetic beads for the easy separation of the ligated product. Briefly, oligonucleotide capture probe functionalized magnetic beads are first hybridized to a target DNA. Biotinylated oligonucleotide detection probes are then allowed to hybridize to the already captured target DNA. A subsequent ligation at the mutation point joins the two probes together. The introduction of streptavidin-conjugated HRP and a simple magnetic separation allow colorimetric genotyping of SNPs. The assay is able to discriminate one copy of mutant in 1000 copies of wild-type KRAS oncogene at 30 picomolar. The detection limit of the assay is further improved to 1 femtomolar by incorporating a ligation chain reaction amplification step, offering an excellent opportunity for the development of a simple and highly sensitive diagnostic tool. (C) 2012 Elsevier B.V. All rights reserved.
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