Hi,
I have been doing DNA plasmid PhiX 174 RF I DNA dialysis against milliq water and purity after dialysis is 1.6. Before Dialysis i have checked and it was 1.92. For my further experiment I needed 1.92 purity (better if I get greater than 1.92). I am doing dialysis to desalt (Tris EDTA ) / i want to remove TE buffer from DNA stock.
1} Milliq water that i used was sterilized, cellulose membrane was pre-treated with 2% sodium bicarbonate and1mM EDTA .
2} Time duration for dialysis 6hrs and every 2hrs i changed milliq.
3} sterile Ultra pure Milliq ph 6.3
Dialysis clamp , forceps to hold the cellulose membrane or handling were all sterile.
4} Dialysis done in 4 degree C.
One possibility is that dialysis removed a contaminant that made the DNA appear to have higher purity than it actually had because the contaminant's absorbance spectrum matched that of DNA. It could be RNA (which degraded during dialysis) or nucleotides, for example.
Thanks Adam, but i was thinking the ph of ultra pure which is 6.3 has any effect on DNA!
Have a look at this from NanoDrop:
http://www.bio.davidson.edu/projects/gcat/protocols/NanoDrop_tip.pdf
It would be advisable to buffer your dialysis solution to avoid pH fluctuations.
in the pdf it has mentioned that ph does effect the reading of nano drop and when i sample blank on nano drop, i use milliq water. Thanks again adam.
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