The pre-programmed Annealing/Extension+ plate read step at 60ºC for 1 min is sub-optimal for my qPCR assay (low efficiency, NTC amplification, etc). When I tried to set conditions for my primers (annealing @55ºC, 15 sec; extension@72ºC, 15 sec) that already worked in a LightCycler 480, the error 1404 warn me that the run was aborted due to insufficient time for plate reading. However, it did allow me to reduce time of the pre-programmed step (60ºC, 30 sec). What could I be doing wrong?
Its too late to answer, but I faced this error now so posting my solution. This error is due to short annealing or extension time. For data collection at a particular step on ABi 7500 instrument, minimum 30 sec time is required. So, in PCR program, if the time duration of the step where data collection is enabled, is less than 30 sec, the machine will give this error. So keep minimum 30 sec time at the step where you acquire data, and issue will be resolved.
I second that of Uma. I have seen a similar error on the ABI 7500DX but that was because we tried to change a preset diagnostic profile. When we set up our own profile we had no problem. I much prefer the LC480 over the ABI systems except for the software. The new Roche PCR software is so much nicer.
Its too late to answer, but I faced this error now so posting my solution. This error is due to short annealing or extension time. For data collection at a particular step on ABi 7500 instrument, minimum 30 sec time is required. So, in PCR program, if the time duration of the step where data collection is enabled, is less than 30 sec, the machine will give this error. So keep minimum 30 sec time at the step where you acquire data, and issue will be resolved.
Thx Muhammad Shakeel , you are right, I have tried such changes before your answer and the error has disappeared !
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