PCR failing - (Sep/22/2004 )
hello to anyone who has had this problem.
i have pcr products from a run of 6 to 7 samples and conducted RFLP and got the correct size products. but when i repeat the procedure for a larger sample..say 45 samples...it fails??
Did you cool your samples?
It's necessary...at big numbers of samlpe RFLP ..
It's necessary...at big numbers of samlpe RFLP ..
helllo, yes i did. even when i repeated some of the samples again (in small numbers), the pcr failed
actually, what do you mean by cool down....holding? if so for how long? 
Hi,
seeing your PCR failed the second time round, even with the small number of samples, it sounds to me like soemthing in your reaction has gone off. If your other PCRs (if you are running any) work, and it's just this one, then your DNA is being degraded. otherwise it could be any of your reagents, dNTPs and Taq being the first ones to check.
Try running out your DNA to check integrity, or spec it for A260.280 ratio. Also change to fresh reagents.
Good luck!
seeing your PCR failed the second time round, even with the small number of samples, it sounds to me like soemthing in your reaction has gone off. If your other PCRs (if you are running any) work, and it's just this one, then your DNA is being degraded. otherwise it could be any of your reagents, dNTPs and Taq being the first ones to check.
Try running out your DNA to check integrity, or spec it for A260.280 ratio. Also change to fresh reagents.
Good luck!
helllo..i have check the dNTP's and Taq...they appear fine as it's also worked on another PCR...i have also checked the quality of the DNA (quality a bit poor). i'm currently running another PCR on another SNP...so i hope it works!
my last PCR failed again.... 
wat can i do...how low can you use Taq once it has been opened. the one i am currently using was opened in late July?
Hi,
it shouldn't matter when you open your Taq, but rather how it is stored and manipulated (ie, at -20C, always on ice, etc).
Is ther a PCR which always works in your lab you could try?
Or maybe try another vial of Taq?
Hello, i'm just set up another pcr that worked...to check to see if it works. i have a feeling it 's the primers? maybe fresh stock has to be made (was made not long ago ). thanks..i will keep trying
only the controls worked.....stuck!!!