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Proceedings Paper

An ECL-PCR method for quantitative detection of point mutation
Author(s): Debin Zhu; Da Xing; Xingyan Shen; Qun Chen; Jinfeng Liu
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Paper Abstract

A new method for identification of point mutations was proposed. Polymerase chain reaction (PCR) amplification of a sequence from genomic DNA was followed by digestion with a kind of restriction enzyme, which only cut the wild-type amplicon containing its recognition site. Reaction products were detected by electrochemiluminescence (ECL) assay after adsorption of the resulting DNA duplexes to the solid phase. One strand of PCR products carries biotin to be bound on a streptavidin-coated microbead for sample selection. Another strand carries Ru(bpy)32+ (TBR) to react with tripropylamine (TPA) to emit light for ECL detection. The method was applied to detect a specific point mutation in H-ras oncogene in T24 cell line. The results show that the detection limit for H-ras amplicon is 100 fmol and the linear range is more than 3 orders of magnitude, thus, make quantitative analysis possible. The genotype can be clearly discriminated. Results of the study suggest that ECL-PCR is a feasible quantitative method for safe, sensitive and rapid detection of point mutation in human genes.

Paper Details

Date Published: 4 April 2005
PDF: 8 pages
Proc. SPIE 5704, Genetically Engineered and Optical Probes for Biomedical Applications III, (4 April 2005); doi: 10.1117/12.588909
Show Author Affiliations
Debin Zhu, South China Normal Univ. (China)
Da Xing, South China Normal Univ. (China)
Xingyan Shen, South China Normal Univ. (China)
Qun Chen, South China Normal Univ. (China)
Jinfeng Liu, South China Normal Univ. (China)

Published in SPIE Proceedings Vol. 5704:
Genetically Engineered and Optical Probes for Biomedical Applications III
Darryl J. Bornhop; Samuel I. Achilefu; Ramesh Raghavachari; Alexander P. Savitsky, Editor(s)

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