Rapid and automated sample preparation for nucleic acid extraction on a microfluidic CD (compact disk)

摘要

Rapid and automated preparation of PCR (polymerase chain reaction)-ready genomic DNA was demonstrated on a multiplexed CD (compact disk) platform by using hard-to-lyse bacterial spores. Cell disruption is carried out while beadcell suspensions are pushed back and forth in center-tapered lysing chambers by angular oscillation of the disk - keystone effect. During this lysis period, the cell suspensions are securely held within the lysing chambers by heatactivated wax valves. Upon application of a remote heat to the disk in motion, the wax valves release lysate solutions into centrifuge chambers where cell debris are separated by an elevated rotation of the disk. Only debris-free DNA extract is then transferred to collection chambers by capillary-assisted siphon and collected for heating that inactivates PCR inhibitors. Lysing capacity was evaluated using a real-time PCR assay to monitor the efficiency of Bacillus globigii spore lysis. PCR analysis showed that 5 minutes' CD lysis run gave spore lysis efficiency similar to that obtained with a popular commercial DNA extraction kit (i.e., IDI-lysis kit from GeneOhm Sciences Inc.) which is highly efficient for microbial cell and spore lysis. This work will contribute to the development of an integrated CD-based assay for rapid diagnosis of infectious diseases.