Temp Gradient PCR - (Aug/31/2009 )
Hi all, Could someone tell me abt temperature gradient PCR? As i understand its a pcr performed with different annealing temperatures and then the final product seen on an agarose gel to see at which temp the primer performs the best. I know this is the basic explanation, can you provide more details?
Thanx in adv
Gradient PCR is generally performed to standardize the annealing temp's. The highest annealing temp at which you see your desired product is optimised by this process to prevent non specific amplification.
You need to have a thermocycler that will generate a temperature gradient, obviously. Usually this is achieved by blowing air along the block; this cools one end of the block more than the other. many cyclers allow you to change the amount of change.