Hello everyone. I would like to clarify and get advice on single-stranded DNA electrophoresis. A 2% solution of agarose gel was used, which contained about 12 µl of ethidium bromide. The DNA marker (or ladder) was from 10-300 bp. DNA samples were used with a 4X buffer for loading in the amount of 1 µl of buffer and 4 µl of DNA sample. 1 sample from the ladder (5.5 OE), 2 (10.0) and the extreme (5.5 OE). 1X TBE was used as a buffer solution. The strength of the intense field was 90 V. And so on for 1.5 hours. But the ladder, alas, did not work. The lines with the samples are blurred at the end. There is an assumption that the concentration of agar must be increased and the voltage reduced. But honestly I don't know. Do you have any suggestions
What is the nature of your dna samples?
It looks like your ladder is separating well but a lower voltage would cause less thermal diffusion so better separation but the dna samples are running at a tiny size. It looks too small even to be RNA so I am thinking that your dna samples are very degraded and that you have a storage or isolation problem allowing such a degree of degradation
Paul Rutland These are synthetic DNA samples of small size. Some are 5.5 OE, and others are 10. Phosphoramidites of nucleosides served as monomers during synthesis. Samples are stored in the freezer at -20 0C. Thawed 1-2 times a week
Maxim Kutyrev Please ignore my previous about degradation and RNA. I misunderstood what you are doing. For samples that small or unusually charged due to solvation effects and possibly by secondary structure if you want to use agarose then I would use 3% agarose (3% low gelling or at least 2% low gelling and 1% ordinary grade agarose,) run at low voltage to minimise thermal diffusion and run the gel less far ( shorter time) to maximise the band intensity. Is there a good reason for not running PAGE gels? ....they separate oligos better than agarose and if using denaturing page secondary structure is minimised. Are these gels diagnostic/analytical or preparative ? .If preparative and you are trying to separate small moieties and salts perhaps size exclusion mini columns would be applicable using something like sephadex g25
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