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Old 04-03-2014, 06:15 PM   #1
FredTam1
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Default Hi-Di Formamide accidentally added to sequencing rxn mix

Hi! Ok, my sequencing reaction was all set up great, I cleaned with with "Centrisep" plates from Princeton Separations, and as a last step I fill any empty columns on my plate with Hi-Di formamide so as to not dry out my ABI 3730.

Question, I accidentally just went through and added Hi-Di Formamide to samples actually containing sequencing reaction that I needed. Will this in any way hurt or destroy the DNA, ruin the reaction, will I get OK results? Thank you so much in advance!
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Old 04-04-2014, 05:39 AM   #2
bbeitzel
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You might get lower intensity fluorescence due to loading less of the diluted DNA into the capillaries, but other than that it should be fine. We used to dry down our BigDye reactions and then resuspend the pellet in HiDi formamide before running.
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Old 04-05-2014, 01:44 PM   #3
FredTam1
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Yes, thanks. After freaking out I read a lot about Hi-Di and its actually used to denature the DNA right before going into the capillaries on the sequencer. So it's actually meant to augment the whole process, the data came out clean.
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