pcr contamination - (Oct/02/2007 )
hi every body
i have the same problem i have no amplified band after pcr carryover i repeat the pcr but i have the same result
all i have is primer dimer if the contamination is the same resone
can i put the water, buffer & dNTPs in uv 2 get rid of contamination or i will lose them
thx u very much
If you think that it is your water, buffer or dNTPs that are contaminated, I would just chuck them out and get fresh ones. I don't know whether UV would do the trick or not- I suspect not, and I think it might damage your dNTPs??
Can anyone else elaborate???
do you mean you have a PCR product in the negative water only controls? If contamination, I suggest you throw everything away and start with fresh solutions and reagents.
do you mean you don't have any PCR products in both your samples and negative controls?
i thank you for ur reply
i mean i have no result in my pcr with my primer control all i have is primer dimer
i dont know the reasone. if this because the contamination or what
i used master mix instead taq, buffer & dntps to minimizes the risk of contamination
but have no band appear than thx u again /regards
do you mean to say that your PCR reaction has not worked? Your negative controls are clean? (no PCR bands)
If so you will have optimise your PCR conditions. How big is the desired product and what are you tm of your primers? Try reducing your annealing temperature. Perhaps try a gradient PCR to find an optimal temperature. Also try increasing your mg ion concentration. Since you are using a master mix, you will have to find a vial of magnesium salt solution and added in some. Try increasing Mg ion content by 50%.
If that doesn't work there are several other things you can try.