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In some of the steps of incubation, when the incubation is 50oC,...

Pragnya Das May 1, 2022

Protocol: circFL-seq, A Full-length circRNA Sequencing Method

In some of the steps of incubation, when the incubation is 50oC, the lid temp of the thermal cycler is 50oC, while when the incubation is 22oC, the lid temp is 105oC, when the incubation is 37oC, the lid temp is 37oC. Why is that so? how do you decide what should be the Lid temp?

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Zelin Liu Author Answered May 3, 2022

Peking University Health Science Center Beijing

Thank you for your question. I think A1.2 "Place samples in a thermocycler with a heated lid set to approximately 105°C. Heat the mixture to 95°C for 2 min, slowly cool to 22°C (-0.1°C/s), and incubate at 22°C for an additional 5 min" is the reason for your confusion.
Acutually, the 105°C lid is set for the high tempture reaction during the temperature gradient decline from 95°C to 22°C.

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