Recently I have been using a High range DNA ladder ranging from 48kb to 10kb with manufactures instruction. But I am not able get a good separation.Bands are found to be staked at one end of the gel.I am using Invitrogen Ultra pure agarose.
What percentage gel are you running? To separate these, you will need to use a very low percentage of agarose (0.5 or 0.6%). ). The 0.4% is pretty tricky to work with. You could also do a pulse-field gel for much better separation. Also, heat the samples to 65oC before loading and run the gel very slowly, preferably overnight at low voltage (20-30 V).
Sir, thanks for your input. If you have noticed the attachment I have run in the same condition.here we are trying to avoid PFGE.
You could also try with low concentration of DNA ladder as well. Mix half volume of ladder you use and adjust the gel loading volume with loading dye.
I agree with Asha's suggestions. I also had similar problem like yours.I use fermentas 1Kb ladder. Earlier I use to load 1-2ul of ladder with no proper separation later I started loading 0.5ul ladder on 1% gel and separation of bands was very good. How much volume and concentration of ladder you are loading on your gel?
We had the same problem when we dyed the gel with Gel Red before runing the electrophoresis, but our results improved when we started to dye the gel after runing the electrophoresis in a Gel Red 3X solution.
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