Biology Forum › Molecular Biology › AGOROSE GEL PROBLEM
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- August 18, 2008 at 11:46 am #9981
biolog35Participantı have really interesting problem.ı digested my plasmid DNA(2.5ug).To control digestion ı loaded 5 ul rxn mixture into well and ı see an excellent band.After checkıng dıgestıon ı dephosphorylated plasmıd DNA and loaded all mıxture into another weel but ı could not see any band 🙁 ı could not understand reason.any idea? please
- August 19, 2008 at 2:19 pm #85573CatParticipant
You need to purify your DNA prior to running your gel. Salts and other impurities can interfere with the electrophoresis.
- August 19, 2008 at 5:09 pm #85576canalonParticipantquote Cat:You need to purify your DNA prior to running your gel. Salts and other impurities can interfere with the electrophoresis.
😯 Never heard that. I use electrophoresis to purify DNA…
As for the original question. Besides contamination of reagents by DNAse (it happened to someone I knew), there is no reason the DNA should disappear. The question is how much DNA in "all the mixture"? And did you had a purification step between the digestion and the dephosphorylation. If something went wrong at that step, that might explain your problems
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