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An Optimized Electrophoresis System for Tandem SSCP and Heteroduplex Analysis of p53 Gene Exons 5-9 on Glass Microfluidic Chips

Published

Author(s)

Christa N. Hestekin, J P. Jakupciak, Thomas N. Chiesl, C D. O'Connell, Annelise E. Barron, C W. Kan

Abstract

With the sequencing of the human genome, there is a growing need for rapid and sensitive genotyping methods that can be incorporated into the clinical setting. DNA-based methods, such as single strand conformational polymorphism (SSCP) and heteroduplex analysis (HA), are commonly used in a research setting to examine mutations in cancer-related genes, but need optimization to achieve the high sensitivity needed for a clinical setting. Here we investigate the importance of several parameters such as polymer matrix, wall coating, and electric field strength on the mutation detection and sensitivity of microchip electrophoresis-SSCP/HA for exons 5-9 of the p53 gene. By looking at the effect of concentration and molecular weight of the polymer matrix, linear polyacrylamide, we determined that 8% 600 kDa was the optimum polymer for providing high resolution of the mutation detection. In addition, we found that including a small amount of the polymer wall coating, poly-N-hydroxyethylacrylamide, improved resolution, decreased loading times, and extended coating lifetime.
Citation
Electrophoresis
Volume
27
Issue
19

Keywords

capillary electrophoresis, genetic screening, microchip, p53, polymer

Citation

Hestekin, C. , Jakupciak, J. , Chiesl, T. , O'Connell, C. , Barron, A. and Kan, C. (2006), An Optimized Electrophoresis System for Tandem SSCP and Heteroduplex Analysis of p53 Gene Exons 5-9 on Glass Microfluidic Chips, Electrophoresis (Accessed May 25, 2024)

Issues

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Created September 30, 2006, Updated October 12, 2021