| Summary: | 碩士 === 國立成功大學 === 化學系 === 87 === In this study, a rapid and reliable analysis of PCR products of hepatitis C virus by capillary electrophoresis with UV detection at 254 nm was developed for clinical diagnoses.
A polyacrylamide-derivatized column filled with 1% HPMC in TBE buffer containing 10μM ethidium bromide was demonstrated to resolve the PCR products of hepatitis C virus in 15 minutes, and identification of the amplified HCV fragment (145 base pair) was further assessed using a DNA Marker（fX-174-RF DNA HaeⅢ）. It was found that the use of DNA intercalating dyes enhanced the resolution and intensity. Moreover, the HCV assay established on silica tubing could be directly transferred to the DB-17 column and PMMA tubing. Results further indicate that the concentration and mass sensitivity of CE method were superior to agarose gel electrophoresis by a factor of 50 and an order of 4, respectively.
An analysis of single-strand conformation polymorphism (SSCP) of HCV amplicon in 5`-noncoding region by capillary electrophoresis was also attempted. This approach was successfully applied to separation the same base pairs but different sequence single strand DNA by high resolution of capillary electrophoresis. The influences of factors such as the applied voltage, and temperature were systematically investigated. The SSCP analysis based on capillary electrophoresis for analysis of the polymorphism of HCV is well suited in clinical diagnoses.
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