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| − | [[Image:Part icon tag.png]]
| + | #redirect [[Help:Proteins]] |
| − | <small>Browse [https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=Tag Tag parts]!</small>
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| − | <hr>
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| − | ==Introduction to Tags==
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| − | [https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=Tag (Degradation) tags] are genetic additions to the end of a sequence that modify expressed proteins in different ways such as marking the protein for faster degradation (see below).
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| − | Tags are also used for binding various particles in solution (making them easier to purify in solution from other proteins in a cell). Famous examples of the latter are the [http://en.wikipedia.org/wiki/His-tag "His tag"] [which binds to nickel (Ni++) or cobalt(Co++) ions attached to a matrix] and the [http://www-users.med.cornell.edu/~jawagne/FLAG-tag.html "FLAG tag"] (which binds to an "AntiFLAG" antibody). Both of these tags are commercially available.
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| − | ==Degradation Tags==
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| − | These genetic tags mark a protein for degradation, thus decreasing a protein's half-life (see each tag's specifications for their respective half-lives).
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| − | One of the useful aspects of genetic tags is the ability to detect gene activity in a time-sensitive manner.
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| − | Tags can be found on:
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| − | * [[Image:Part icon reporter.png]][https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=reporter Reporters],
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| − | * [[Image:Part icon cds.png]][https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=coding Protein-coding parts],
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| − | * [[Image:Part icon generator.png]][https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=generator Protein Generators], and
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| − | * [[Image:Part icon inverter.png]][https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=inverter Inverters].<br>
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| − | ''Note'': Tags cannot currently be used separately as [[An Introduction to BioBricks|BioBricks]] because currently all of our [https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=Coding Protein-coding parts] end in the double stop codon "<b>TAATAA</b>".
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| − | ===Mechanism of Use===
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| − | Currently all of our [https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=Tag tags] operate through the use of protein-degrading enzymes (proteases) within the cell.
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| − | They do so by coding for a sequence of about eleven amino acids at the C-terminus of a protein. This sequence is normally generated in ''E. coli'' when a ribosome gets stuck on a broken ("truncated") mRNA. Without a normal termination codon, the ribosome can't detach from the defective mRNA. A special type of RNA known as ssRA ("small stable RNA A") or tmRNA ("transfer-messenger RNA") rescues the ribosome by adding the degradation tag followed by a stop codon. This allows the ribosome to break free and continue functioning. The tagged, incomplete protein gets degraded by the proteases ClpXP or ClpAP. When the BioBrick protein-coding sequence ends in a similar degradation tag, the ClpXP or ClpAP proteases can attack it, resulting in a shorter half-life. No such system is yet known for yeast, however ClpXP has been expressed in yeast successfully, so the standard sSRA system is easily portable.
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| − | Although originally the number of amino acids encoding for a ssRA/tmRNA tag was ''eleven'' (the first sequence that encoded for destruction via ssRA tagging was <b>AANDENYALAA</b> by [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=8584937 Keiler ''et al.'']), a subsequent study by [http://www.pubmedcentral.gov/articlerender.fcgi?tool=pubmed&pubmedid=9603842 Andersen ''et al.''] tested the efficacy of mutating the last ''three'' amino acids of that system. Thus our tags (<b>AAV, ASV, LVA, LAA</b>) are classified by only three amino acids.
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| − | ===Future Plans===
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| − | Our future plans for tags involve building proteins using BioBricks. To read more on this project, visit [https://dspace.mit.edu/handle/1721.1/32535 Ira Phillip's work on BioBrick modification].
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| − | ===References===
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| − | # Jabri and Nature News. "Tag, you're degraded". 2003. [http://www.nature.com/nsmb/journal/v10/n9/full/nsb0903-676.html article] | + | |
| − | # Karzai, A.W.. "The SsrA−SmpB system for protein tagging, directed degradation and ribosome rescue". 2000. [http://www.nature.com/nsmb/journal/v7/n6/full/nsb0600_449.html Review article]
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| − | # Keiler, K.C. ''et al.'' "Role of a peptide-tagging system in degradation of proteins synthesized from damaged messenger RNA." 1996. [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=8584937 Pubmed]
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| − | # Andersen, J.B. ''et al.'' "New Unstable Variants of Green Fluorescent Protein for Studies of Transient Gene Expression in Bacteria". 1998. [http://www.pubmedcentral.gov/articlerender.fcgi?tool=pubmed&pubmedid=9603842 link]
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