I had set up overnight ligation reaction at 16 degree celsius in a PCR machine but the lid temperature was high around 95 degrees. Will it affect the efficeincy of ligation if done in PCR in this way?
Amy Klocko
the ligation should work fine but the lid can be cooler to save wear and tear on your PCR machine. The 95 lid is meant to prevent condensation during a standard PCR cycle.
if the ligation is challenging you could try alternative ligases that work at room temperature.
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