A rapid micro-polymerase chain reaction system for hepatitis C virus amplification

Abstract

A rapid micro-polymerase chain reaction (μ-PCR) system was integrated to amplify the complementary DNA (cDNA) molecules of hepatitis C virus (HCV). This system consists of a rapid thermal cycling system and a μ-PCR chip fabricated by MEMS fabrication techniques. This rapid μ-PCR system is verified by using serum samples from patients with chronic hepatitis C. The HCV amplicon of the rapid μ-PCR system was analyzed by slab gel electrophoresis with separation of DNA marker in parallel. The μ-PCR chip was fabricated on silicon wafer and Pyrex glass using photolithography, wet etching, and anodic bonding methods. Using silicon material to fabricate the reaction well improves the temperature uniformity of sample and helps to reach the desired temperature faster. The rapid close loop thermal cycling system comprises power supplies, a thermal generator, a computer control PID controller, and a data acquisition subsystem. The thermoelectric (TE) cooler is used to work as the thermal generator and a heat sink by controlling the polarity of supplied power. The μ-PCR system was verified with traditional PCR equipment by loading the same PCR mixture with HCV cDNA and running the same cycle numbers, then comparing both HCV amplicon in slab gel electrophoresis. The HCV amplicon from the μ-PCR system shows a DNA fragment with an expected size of 145 bp. The background is lower with the μ-PCR system than that with the traditional PCR equipment. Comparing the traditional PCR equipment which needs 5.5 h for 30 cycles to gain the detectable amount of HCV amplicon in slab gel separation, this μ-PCR system takes 30 min to finish the 30 thermal cycles. This work has demonstrated that this rapid μ-PCR system can provide rapid heat generation and dissipation, improved temperature uniformity in DNA amplification.