Hello colleagues, I found the Eco system real time PCR illumina device in the lab. The last people who used the device are already out... So after the initial excitation about using it, I discovered that the sample sizes are quite smaller than the normal PCR samples I used to make in Eppendorf devices. So is the real-time PCR sample preparation the same for this device as any other PCR? or is it different? any and all information is highly welcome, I went over the user guide several times, it generously explains how to use the device, except for the part pertaining to the actual sample preparation. Thanks a million in advance
The sample preparation for real-time PCR on the Illumina Eco system does follow the same general principles as standard real-time PCR. However, there are some specific requirements to adapt the protocol for the Eco system:
- Reaction volume is very small - only 10-20 μL per well compared to 20-50 μL for standard PCR plates. So reagents need to be scaled down accordingly.
- Special 384-well plates are used. Make sure to use compatible optical plates provided by Illumina.
- More concentrated primer stocks may be needed to accommodate the smaller reaction volumes. 100 μM primer stocks are common.
- Always include positive and negative controls.
The actual real-time PCR reagents (Taq polymerase, buffers, dNTPs) and cycling conditions can remain the same as standard PCR. Just the preparation needs to be adapted for the smaller 384-well Eco plates.
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