epitope tagging

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l_cremin
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epitope tagging

Post by l_cremin » Mon Nov 07, 2005 11:07 pm

Does anyone know a good website, or can tell me what tagging methods there are for recombinant DNA tagging and advantages and disadvantages of each!?

thanks!

sdekivit
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Post by sdekivit » Tue Nov 08, 2005 5:20 pm

famous tags are the V5-epitope and the histidine-tag. In a tagging experiment, a vector is used: the pYES2.1/V5-His-TOPO vector.

The insert contains the DNA code for a V5-epitope and a histidine-tail behind a GAL1-promotersite.

--> this is advantageous for purifying methods and western blotting. When purifying for example a protein that contains a histidine-tag, affinitychromatography is easily used, using a Ni-NTA-column where the histidinetag will bind.

In western blotting, identification of a protein of interest can be visualized due to the V5-epitope, where a specific antibody can bind to. This can be visualized by using a secondary labelled antibody for example.

l_cremin
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Post by l_cremin » Tue Nov 08, 2005 7:28 pm

Thanks ever so much for your help....much appreciated!

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