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-   -   Non-specific band in positive control but negative control not (http://www.molecularstation.com/forum/pcr-polymerase-chain-reaction-forum/83101-non-specific-band-positive-control-but-negative-control-not.html)

Ruby_tran 07-05-2011 07:31 AM

Non-specific band in positive control but negative control not
 
Dear guys,

My previous PCRs were OK but recently, I have got a problem of nonspecific band in positve control. I'm sure this is not contamination in reagents because negative control is no problem. So i tend to think may be positive DNA was contaminated. But non-specific band is about 1000kb :frusty: and target band is 123bp:frusty:

If my PCR process have problem, previous PCR results would have non-specific band. But they were not.

PCR reaction
940C for 4 min
Denaturation : 94C for 2 mins
Annealing : 68C for 2 mins 35 cycles
Extension : 72C for 1 mins
72C for 10 mins

load DNA 5ul

May be because concentration DNA was too much? It's was about 28 ng/ul:shakehead:


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