see also:

https://www.researchgate.net/post/How_to_decide_the_annealing_temperature_range_in_touchdown_PCR

https://www.researchgate.net/post/What_is_Touch_Down_PCR_When_is_it_used

Thank guys for the suggestion but I can get the specific band through TD-PCR in normal cycler but not in Bio-Rad IQ5 2 steps real time PCR.

If I understand you correctly, you are using different PCR conditions in the two systems. I would suggest to design new primers for the qPCR (or use the same PCR conditions in the qPCR for the primers you have).

Sabine Strehi, I used the same PCR conditions in qPCR with the same primers but it does not give me any specific quantification.

Guys, I managed to optimised the real time TD-PCR protocol. Thanks for the advice and suggestions..