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

A rapid micro-polymerase chain reaction (mu -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 mu -PCR chip f abricated by MEMS fabrication techniques. This rapid mu -PCR system is veri fied by using serum samples from patients with chronic hepatitis C. The HCV amplicon of the rapid mu -PCR system was analyzed by slab gel electrophore sis with separation of DNA marker in parallel. The mu -PCR chip was fabrica ted on silicon wafer and Pyrex glass using photolithography, wet etching, a nd 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 com prises power supplies, a thermal generator, a computer control PID controll er, and a data acquisition subsystem. The thermoelectric (TE) cooler is use d to work as the thermal generator and a heat sink by controlling the polar ity of supplied power. The mu -PCR system was verified with traditional PCR equipment by loading the same PCR mixture with HCV cDNA and running the sa me cycle numbers, then comparing both HCV amplicon in slab gel electrophore sis. The HCV amplicon from the mu -PCR system shows a DNA fragment with an expected size of 145 bp. The background is lower with the mu -PCR system th an that with the traditional PCR equipment. Comparing the traditional PCR e quipment which needs 5.5 h for 30 cycles to gain the detectable amount of H CV amplicon in slab gel separation, this mu -PCR system takes 30 min to fin ish the 30 thermal cycles. This work has demonstrated that this rapid mu -P CR system can provide rapid heat generation and dissipation, improved tempe rature uniformity in DNA amplification. (C) 2000 Elsevier Science B.V. All rights reserved.