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CHAPTER 4 – Assay Setup
4.1 General PCR Considerations
It is important to minimize sample contamination and PCR product carryover when setting up a
new experiment. Please take note of the following precautions:
o
Wear a clean lab coat, clean gloves, and eye protection when preparing samples for PCR
amplification. Change gloves whenever you think there may be a contamination
carryover.
o
Ensure there are separate areas for PCR setup and PCR amplification. Avoid bringing
amplified PCR products into the PCR setup area.
o
Open and close all sample tubes carefully. Avoid splash-back of the PCR samples.
o
Use aerosol resistant pipette tips and change tips after each use.
o
Ensure that reaction and reagent components are always capped when not in use.
o
Clean lab benches and equipment periodically with 70% ethanol or 10% bleach followed
by 70% ethanol.
4.2 Create a New Experiment
To set up a new assay procedure, log into your account through the appropriate web address.
On the home screen, click on the green icon titled
Create A New Experiment
. You will then see
a box asking for the experiment name. Input the information and click on
Create Experiment
.
Figure 4.2
Figure 4.1
You will be brought to the assay setup screen. The protocol parameters must be defined on this
page prior to running the experiment. The default cycling conditions are shown in Figure 4.3. In
the default protocol, data is collected during the annealing step after completion of each cycling
stage.
