Polymerase chain reaction thermal cycling using the programmed tilt displacements of capillary tubes

Abstract

A thermal cycling method, whereby capillary tubes holding polymerase chain reactions are subjected to programmed tilt displacements so that they are moved using gravity over three spatial regions (I, II, and III) kept at different constant temperatures to facilitate deoxyribonucleic acid (DNA) denaturation, annealing, and extension, is described. At tilt speeds in excess of 0.2 rad/s, the standard deviation of static coefficient of friction values was below 0.03, indicating in sync movement of multiple capillary tubes over the holding platform. The travel time during the acceleration phase and under constant velocity between adjacent regions (I to II and II to III) and distant regions (III to I) was 0.03 s and 0.31 s, respectively. The deviations in temperature did not exceed 0.05 °C from the average at the prescribed denaturing, annealing, and extension temperatures applied. DNA amplification was determined by optical readings, the fluorescence signal was found to increase twofold after 30 thermal cycles, and 1.16 × 106 DNA copies/μl could be detected. The approach also overcomes problems associated with thermal inertia, sample adhesion, sample blockage, and handling of the reaction vessels encountered in the other thermal cycling schemes used.

Low-cost Imaging of Fluorescent DNA in Agarose Gel Electrophoresis using Raspberry Pi cameras

“…The reaction samples from each vial was transferred to 75 □μl capillary glass tubes (Hirschmann) that were sealed at both ends using putty. The tubes were then loaded in a tilting platform thermal cycler system for PCR thermal cycling amplification [21]. The PCR protocol parameters applied for thermal cycling are given in Table 1.…”

Low-Cost Imaging of Fluorescent DNA in Agarose Gel Electrophoresis Using Raspberry Pi Cameras

“…The reaction samples from each vial was transferred to 75 µl capillary glass tubes (Hirschmann) that were sealed at both ends using putty. The tubes were then loaded in a tilting platform thermal cycler system for PCR thermal cycling ampli cation [21]. The PCR protocol parameters applied for thermal cycling are given in Table 1.…”

Filterless fluorescent DNA sensing in capillary tube PCR

“…The reaction samples from each vial were transferred to 75 µl capillary glass tubes (Hirschmann) having inner and outer diameters of 1.07 mm and 1.23 mm respectively that were sealed at both ends using putty. The tubes were then loaded in a tilting platform thermal cycler system for PCR thermal cycling ampli cation [14]. The PCR protocol parameters applied for thermal cycling are given in Table 1.…”

“…Continuous-ow PCR is performed by pumping the mixture through a uidic channel into regions that are kept at different temperatures [12], while convective PCR involves the circulation of reactants between the hot and cool zones of the system that correspond to denaturation, annealing and extension [13]. More recently, an approach that applies programmed tilt displacements, so that the reactants held in capillary tubes are displaced by gravitational force to and from closely adjacent holding regions kept at different constant temperatures, has been shown to overcome many of the issues encountered with the other approaches [14][15][16].…”