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PCRmon wiki

Portable PCR system for decentralized testing applications.

Pipetting setup for PCR

Pipetting tools for setting up PCR reactions
Figure 1: The system shown consists of a reaction master mix tube that can contain the sample, mineral oil tube (for keeping the reaction mix at bottom of PCR tube during thermocycling), pipettor tool, wide-bore pipette for aspirating and dispensing mineral oil, and a small volume pipette for aspirating and dispensing the reaction master mix. Also shown are PCR tubes that fit in the PCRmon thermocycler. The mineral oil is layered on top of the aspirated reaction mix.

How the pipettor works
Figure 2: The pipettor is able to securely attach to different types of pipette tips. By squeezing the top part of the silicone tubing, it is possible to aspirate and dispense very small volumes. To consistently aspirate the same volume, a marking on the pipette tip can be drawn as a reference.

Accurately aspirating microliter volumes of reaction mix
Figure 3: In order to transfer the reaction from the source well into the PCR test tube, you need to aspirate then dispense the sample. This example demonstrates how it is easy to accurately aspirate microliter volumes of reaction mix (in this action the volume aspirated is 10µl). In order to aspirate, first, you squeeze the tubing at the top of the pipette using your index finger and thumb in order to reduce the volume in the pipette, then you move the pipette into the tube containing the reaction mix that you want to aspirate and carefully reduce the squeezing pressure in order to create suction to aspirate the volume enough to reach the marketing. Then you move the pipette tip to the empty PCR tube and dispense the volume.

Accurately aspirating mineral oil
Figure 4: Because the PCRmon thermal block does not have a heated lid, mineral oil is needed in order to keep the volume of the reaction mix at the bottom of the PCR tube during thermocycling. The smaller pipettes used for aspirating the reaction mix have too small of an orifice for this relatively viscous mineral oil so a larger bore pipette tip is better for this. The volume of the mineral oil added to the reaction mix is not as critical, you can aspirate the total amount of volume (just squeeze and release entirely when the pipette tip is in the mineral oil), this produces around 30μl of mineral oil in the tube.
pcrmon.txt · Last modified: 2021/01/25 20:09 (external edit)