How to discover the source of contamination and get rid of it?
I work with ISSR markers. I am struggling a problem for more than 2 months. Consistently the negative control in the PCR reactions are showing contamination. I alread changed the reagents twice, including the water. I always autoclave the tubes and pipette tips. I do the PCR reactions in the flow. Before starting, I place tips, gloves, pipettes, water and the tubes for 15 minutes in UV; nevertheless, I still get bands in the negative controls and in all negative control displays a different pattern of bands. I have already used new reagents, exchanging one at a time, tubes, pipettes, water. I do sterilize the flow with bleach. However, I have not found out the source of contamination. The worst is that in the same reaction of negative control, using the same mix, some reactions appear bands and others do not.
Does anyone have any idea what is the problem? Any solution?