Dear Magda, might be that your polymerase contains traces of DNA from the host it was produced in. To confirm, add some DNAse to an aliquot of your mastermix, incubate say 1hr at 37 degC (or overnight if you like) and then kill the DNAse by heating to 95degC for say 10 minutes (longe is not so critical, but polymerase of course will decrease in activity). Then redo your assay. Include a control that demonstrates that DNAse is really gone; a control plasmid in a reasonable concentration should be amplified.
Alternatively/additinally, you might try to switch polymerase batch/ brand/type, too. Hope that helps, Wo _______________________________________________ Methods mailing list Methods@net.bio.net http://www.bio.net/biomail/listinfo/methods
