Plasmid #31 enables the labeling of target proteins with both cleavable N-terminal His8 and cleavable C-terminal CL7 tags.
Transcription is induced with IPTG and driven by the T7 RNA polymerase. The plasmid is designed for expression in E. Coli.
The C-terminal CL7 tag is downstream of three PreScission protease (PSC) and one Sortase A (SRT) cleavage site.
Plasmid #31 includes a C-terminal SUMO cleavage site, upstream of the His8 tag; a Tobacco Etch Virus (TEV) cleavage site between the His8 tag and target protein; a SRT site downstream of the target protein; and three PSC cleavage sites between the SRT and CL7 tag.
This plasmid includes N-terminal Trx and His8 tags.
- HindIII/SpeI Insertion Site – Trx | SUMO | His8 | TEV | Gene of Interest | SRT | PSC | PSC | PSC | CL7
- KpnI/SpeI Insertion Site – Trx | SUMO | His8 | Gene of Interest | SRT | PSC | PSC | PSC | CL7
- Bsu36I/SpeI Insertion Site – Trx | SUMO | Gene of Interest | SRT | PSC | PSC | PSC | CL7
This insertion scheme maintains the Gene of Interest’s wildtype sequence, without adding any extra residues. The N-terminus of the Gene of Interest must include the following to complete the SUMO C-terminal sequence: