Hello ,
We need to do ddPCR (Eva green chemistry) for a primer pair that yields a 429 bp amplicon. It is a single band on gel. I understand that there are reason for restriction amplicon size in real time PCR. Is there any such restriction for ddPCR. If yes please let me know the reasons.
Hi Deepak,
as written in this page (https://www.researchgate.net/topic/Droplet-Digital-PCR), amplicons must be between 75 and 100 bp length.
fred
Between 60-200 bp. Larger amplicons will have a lower amplification efficiency and will have a brighter fluorescence amplitude when using EvaGreen. However, This can be a problem with longer amplicons because there is more variability in fluorescence with larger amplicons, so it can be harder for the system to determine whether the droplet is positive or negative.
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