AGOROSE GEL PROBLEM

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biolog35
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AGOROSE GEL PROBLEM

Post by biolog35 » Mon Aug 18, 2008 11:46 am

ı 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

Cat
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Post by Cat » Tue Aug 19, 2008 2:19 pm

You need to purify your DNA prior to running your gel. Salts and other impurities can interfere with the electrophoresis.

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canalon
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Re:

Post by canalon » Tue Aug 19, 2008 5:09 pm

Cat wrote:You need to purify your DNA prior to running your gel. Salts and other impurities can interfere with the electrophoresis.

:shock: 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
Patrick

Science has proof without any certainty. Creationists have certainty without
any proof. (Ashley Montague)

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