I know you have to keep all the reagents in a freezer but I don't know why, please help.
In PCR , amplification of selected DNA fragment can occure upto 25 th cycle
In this reaction temp is elevated for the activity of Taq DNA Polymerase which is upto 80 degree celcius . So for maintaining the stability of all other reagents concern to product formation, cold condition is maintained
Hello Daniela Riquelme
It is a good practice, but not necessary to keep PCR reagents on ice when you are preparing a mixture for PCR. None of the PCR components degrade at room temperature. But it is always recommended that you keep original concentrated vials of PCR reagents on ice and only the master mix may be kept at room temperature.
Taq DNA polymerase is extremely heat resistant with a half-life of 40 minutes at 95°C. It is optimally catalytically active at 70–75°C. When you prepare the master mix, addition of Taq DNA polymerase must be done at the very last.
All PCR reagents come together in a single tube only for a short time before loading on the PCR machine, and so preparing the master mix at room temperature does not make much of a difference. But keeping the original PCR reagents including Taq DNA polymerase on ice during PCR master mix preparation is recommended for long term stability of these reagents just to avoid thermal degradation.
Best.
- Badges
- Science topic
- Similar topics
- Medicine
- Physiology
- Biosignals
- Biomedical Signal Processing
- Statistical Analysis