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smearing in negative control as well - (Mar/14/2006 )

hi,
I have been doing a pcr for amplifying a 396bp product,i was getting it earlier,then it started smearing,now there is even smearing in negative controls where no dna is added,how to correct this,can it be due to dNTPs?
Kindly help

-niveda-

QUOTE (niveda @ Mar 14 2006, 12:00 PM)
hi,
I have been doing a pcr for amplifying a 396bp product,i was getting it earlier,then it started smearing,now there is even smearing in negative controls where no dna is added,how to correct this,can it be due to dNTPs?
Kindly help


do you use BSA in your batch? - whats the size of your smearing?

-dersven-

Hi,I dont use BSA, the msear is in the range of 500-1000bp.

-niveda-

looks a little big for primer-dimer

so...either there is contamination of your stuff (replace reagents, water, tips, clean pipettors, etc till you find the source) or there is something in your mix that is showing up on the gel.

could you please post more details about your protocol? not necessarily cycling conditions, but tell us what you add, etc

before anything else, I would try 'clean' water and cleaning up your pipets

-aimikins-