Everything is standardized as per protocol, but still this smearing is there, need suggestions from PCR experts.
DNA extraction by the magnetic beads-based method.
tried colony Pcr.
PCR conditions optimized,
Different concentrations of both Primer and template were checked.
Gel, Run-time, Voltage everything checked.
what could be the other reason to remove Smearing.
Gel Pic attached
Hello Guido Bognolo, obviously some degree of smearing is likely with any diffusion process, and this will be more the higher up the slide it has had to travel, but hopefully it can be minimized.
Can you verify with a monodisperse MW material how sharp a line the gel gives?
What degree of polydispersity would you expect from the DNA fragments acquired?
Smearing on a DNA gel can be caused by various factors. In your gel picture DNA ladder is not entirely clear. I think there are contaminants in electrophoresis buffer that interfere with DNA migration and cause smearing. Choose the appropriate agarose percentage based on the size of your DNA fragments and with fresh electrophoresis buffer.
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