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Published on: December 19, 2011
[Microanalysis of Acetaldehyde Dehydrogenase 2 Genotype Based on SNaPShot-FCA]
Shu-Shu Zeng1, Xiu-Feng Gao1, You Zhang1
1Department of Biochemistry and Molecular Biology, West China School of Basic Medical Sciences & Forensic Medicine, Sichuan University, Chengdu 610041, China.
Objective:
To establish SNaPShot-fluorescence capillary analysis (SNaPShot-FCA) assay for rapid detection of the genotype of aldehyde dehydrogenase 2 gene (ALDH2) rs671 locus.
Methods:
The genomic DNA was extracted from peripheral blood cells. Using R6G-ddATP and cy5-ddGTP as fluorescent substrates, the ALDH2 gene was amplified by SNaPShot to generate DNA products with different fluorescent dyes at the 3' end. FCA was used to detect the products separated by agarose gel electrophoresis and recovered by gel recovery kit, and the genetype of ALDH2 polymorphism was analyzed by fluorescence spectrum. The samples were tested three times repeatedly and compared with the results of DNA sequencing.
Results:
The optimal concentrations of R6G-ddATP and cy5-ddGTP were 1.4 μmol/L and 8.0 μmol/L, respectively. 106 samples were tested for ALDH2 genotype by SNaPShot-FCA under optimal conditions, including 67 of wild type (GG), 38 of hybrid type (AG), and 1 of mutant type (AA), which were consistent with the sequencing results.
Conclusion:
This study successfully established the SNaPShot-FCA for the micro-detection of ALDH2 genotype for the rapid screening and identification of ALDH2 gene.
