Scientist Solutions: Life Science Discussions
    
Home » Forums » Cloning (Recombinant Gene Expression) » Plasmids/Cosmids/BAC/PAC » Why do you need tags in plasmids?

Thanks to our sponsors who make this site possible

Why do you need tags in plasmids?

RSS Feed

Would you like to save this topic, event, protocol or job so you can find it again easily?

Just click the "Save to My Lab Drawer" link and the item will be saved in the My Lab Drawer section of your bench space.

Available to members only. Please log in or register for your free account now.

supaboy2925
United States

Send PM
See Mini bio

Status: Frog Egg
Frog Egg
Topic Started by supaboy2925
on 12/11/2008 23:38 PM   
Reply to this post Go to the top of the page

Why do most commercially available plasmids contain tags? Can you give an example/experiment of this? Thanks!


Replies
RLS
United States

Send PM
See Mini bio

Status: Cloning Moderator
Frog Laureate
Posted By RLS
on 12/12/2008 7:07 AM   
Reply to this post Go to the top of the page

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.



As a Scientist Solutions member, you are able to register a positive vote for any topic which you believe is useful and relevant to our board or any reply which you believe is especially well worded and helpful.

By participating in the voting, you will be helping to identify the best topics & replies on the board.

You may vote once for any one post, and you may not vote for your own posts.

A post (topic or reply) will earn one "thumbs up" icon for every 10 votes received (up to 3 thumbs up), and the person who made the post will also earn two bonus points.

learn more about member points.



Click here to
Become a member & join our
community (It's easy & free)
Already a member? Please log in
User Name  
Password  
Forget Password?
Find out about our Rewards System
Scientists
Not finding the answer you need?

Post a new topic

You must be logged in to post. Log in above.
Not a member yet? Click here to register
(it's free)
Thank You to Our Sponsor