Part:BBa_K2255003
Multi-Tag (Rfc 25)
This tag provides multiple fixation sites: a Strep-tag [1] and a histidine tag. Between those tag is a TEV protease cutting site [2].
This part was designed with the Freiburg (Rfc25) extension. Thus, it contains the restriction sites NgoMIV and AgeI that are compatible and allow the absence of a start and stop codon, which ease the assembly of multiple protein domain.
Usage and Biology
This part is usefull in our to purify the [http://2017.igem.org/Team:Aix-Marseille/DEPS enzyme] that use to disrupt the biofilm.
GFP purification with a Ni column
Purification of [sfGFP] fusionned with this tag (BBa_K2255003), with the [Ni-NTA Spin Kit] (Quiagen). As you can see, mainly sfGFP is present after elution and most of the protein are gone with the wash. Thus, the his-tag of this part is functionnal.
References
- ↑ Schmidt, Thomas GM; Skerra, Arne (2007). "The Strep-tag system for one-step purification and high-affinity detection or capturing of proteins". Nature Protocols. 2 (6): 1528–35. PMID 17571060. doi:10.1038/nprot.2007.209
- ↑ Parks TD, Leuther KK, Howard ED, Johnston SA, Dougherty WG (February 1994). "Release of proteins and peptides from fusion proteins using a recombinant plant virus proteinase". Anal. Biochem. 216 (2): 413–7. PMID 8179197. doi:10.1006/abio.1994.1060
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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