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Why do you need tags in plasmids?

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supaboy2925
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Topic Started by supaboy2925
on 12/11/2008 23:38 PM
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Why do most commercially available plasmids contain tags? Can you give an example/experiment of this? Thanks!

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RLS
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Posted By RLS on 12/12/2008 7:07 AM
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Are you referring to tags such as a polyhistidine tag, TAP tag, or Myc? Tags such as the polyhistidine (often seen as 'His tag') are placed in frame at either the N-terminus or C-terminus of your protein coding sequence in a vector, which forms a metal-binding site for affinity purification of recombinant fusion protein on metal-chelating resin (such as Qiagen's Ni-NTA, Clontech's Talon Metal affinity resin, or Invitrogen's ProBond). His tags are found in a number of the commercially available protein expression vectors.

Tags such as the TAP, or tandem affinity purification, also allows creation of a fusion protein with a TAP tag on the end, typically for the end purpose of studying protein-protein interactions (often used to purify protein complexes from yeast). The TAP tag is made up of a CBP tag, a TEV protease recognition site, and a ProtA tag. Initially, cell extract is applied to IgG beads, where the ProtA tag binds, followed by cleavage with the protease for release from the beads. Next, the eluate is applied to calmodulin beads, where the CBP tag binds. Elution from the calmodulin beads results in a nice preparation of your protein of interest and proteins interacting with your protein which can be analyzed further.

Other tags, such as the c-myc are epitope tags, where one can determine the expression of the protein of interest in a cell lysate, for example, using Anti-Myc antibodies in a Western blot.

There are many, many tags out there! I hope this helps answer your question.