Plasmid Guide
GenBank files of each plasmid’s sequence are available for download below.
Each plasmid has one distinct sequence but may have several cloning options based on a combination of different restriction enzymes. Reference the tables for detailed breakdowns of the elements that are included or excluded in each cloning option.
BacMam Vectors
BacMam NT H8-CL7-eGFP: Enables target proteins to be labeled with N-terminal His8, CL7, and eGFP tags. Protease cleavage removes all tags.
Sequence file (GenBank format)
BacMam CT eGFP-CL7-H8: Enables target proteins to be labeled with C-terminal eGFP, CL7, and His8 tags. Protease cleavage removes all tags.
Sequence file (GenBank format)
Multi-Subunit Vector
Plasmid 1: Contains the ttRNAP protein – a model DNA/RNA-binding multi-subunit protein – with a C-terminal CL7 tag.
- Options 1-3 - Bsu36I/SwaI, HindIII/SfiI, and AhdI/SpeI - excise subunits 1, 2, and 3, respectively Sequence file (GenBank format)
Single Subunit Vectors
Plasmid 1: Contains the ttRNAP protein – a model DNA/RNA-binding multi-subunit protein – with a C-terminal CL7 tag.
- Option 4 - Bsu36I/SpeI - excises all 3 subunits in a 10 kb restriction fragment Sequence file (GenBank format)
Plasmid 3: Serves as a template for tagging target proteins with an N-terminal CL7 tag.
- Option 1 - HindIII/XhoI Insertion Site- Trx | CL7 | SUMO | SRT | Gene of Interest Sequence file (GenBank format)
- Option 2 - KpnI/XhoI Insertion Site- Trx | CL7 | SUMO | Gene of Interest Sequence file (GenBank format)
- Option 3 - Bsu36I/XhoI Insertion Site- Trx | CL7 | SUMO | Gene of Interest Sequence file (GenBank format)
Plasmid 4: Serves as a control; it encodes the CL7 model (CL7M) protein, which directly binds the Im7 protein immobilized on the Im7 Resin.
- Plasmid 4 Sequence file (GenBank format)
Group 1: Vectors for Membrane Proteins
Plasmid 8: Plasmid #8 is used for the expression of membrane proteins using an engineered signal peptide. It includes a C-terminal His8 tag and CL7 tags with cleavage sites before the His8 tag and in between the His8 tag and the CL7 tag.
- Plasmid 8 Sequence file (GenBank format)
- Plasmid 9 Sequence file (GenBank format)
- Plasmid 10 Sequence file (GenBank format)
Group 20: N-terminal CL7 and Expression Tags
Plasmid 21: Enables target proteins to be tagged with N-terminal CL7 and His8. There are three options for the 5’ placement of target genes, and XhoI is the 3’ cloning site.
- Option 1 - HindIII/XhoI Insertion Site – Trx | CL7 | PSC | 44-Amino Acid Linker | PSC | His8 Tag | SUMO | 13-Amino Acid Linker | SRT | Gene of Interest Sequence file (GenBank format)
- Option 2 - KpnI/XhoI Insertion Site – Trx | CL7 | PSC | 44-Amino Acid Linker | PSC | His8 Tag | SUMO | 3-Amino Acid Linker | Gene of Interest Sequence file (GenBank format)
- Option 3 - Bsu36I/XhoI Insertion Site – Trx | CL7 | PSC | 44-Amino Acid Linker | PSC | His8 Tag | SUMO | Gene of Interest Sequence file (GenBank format)
Plasmid 22: Enables the tagging of target proteins with a cleavable CL7 at the N-terminus and an uncleavable His8 at the C-terminus. There are three options for the 5’ placement of target genes, and SpeI is the 3’ cloning site.
- Option 1 - HindIII/SpeI Insertion Site – Trx | CL7 | 20-Amino Acid Linker | SUMO | SRT | 13-Amino Acid Linker | Gene of Interest | His8 Sequence file (GenBank format)
- Option 2 - KpnI/SpeI Insertion Site – Trx | CL7 | 20-Amino Acid Linker | SUMO | 3 aa | Gene of Interest | His8 Sequence file (GenBank format)
- Option 3 - Bsu36I/SpeI Insertion Site – Trx | CL7 | 20-Amino Acid Linker | SUMO | Gene of Interest | His8 Sequence file (GenBank format)
- Option 4 - HindIII/XhoI Insertion Site – Trx | CL7 | 20-Amino Acid Linker | SUMO | SRT | 13-Amino Acid Linker | Gene of Interest Sequence file (GenBank format)
- Option 5 - KpnI/XhoI Insertion Site – Trx | CL7 | 20-Amino Acid Linker | SUMO | 3 aa | Gene of Interest Sequence file (GenBank format)
- Option 6 - Bsu36I/XhoI Insertion Site – Trx | CL7 | 20-Amino Acid Linker | SUMO | Gene of Interest Sequence file (GenBank format)
Group 30: C-terminal CL7 and N-terminal Expression Tags
Plasmid 31: Enables the labeling of target proteins with both cleavable N-terminal His8 and cleavable C-terminal CL7 tags.
- Option 1 - HindIII/SpeI Insertion Site – Trx | SUMO | His8 | TEV | Gene of Interest | SRT | PSC | PSC | PSC | CL7 Sequence file (GenBank format)
- Option 2 - KpnI/SpeI Insertion Site – Trx | SUMO | His8 | Gene of Interest | SRT | PSC | PSC | PSC | CL7 Sequence file (GenBank format)
- Option 3 - Bsu36I/SpeI Insertion Site – Trx | SUMO | Gene of Interest | SRT | PSC | PSC | PSC | CL7 Sequence file (GenBank format)
- Sequence file (GenBank format)
Plasmid 33: Enables target proteins to be labeled with C-terminal His8 and CL7, retaining the His8 tag depending on the protease utilized.
- Option 1 - HindIII/SpeI Insertion Site – Trx | SUMO | Gene of Interest | SRT | His8 | PSC | PSC | PSC | CL7 Sequence file (GenBank format)
- Option 2 - KpnI/SpeI Insertion Site – Trx | SUMO | Gene of Interest | SRT | His8 | PSC | PSC | PSC | CL7 Sequence file (GenBank format)
- Option 3 - Bsu36I/SpeI Insertion Site – Trx | SUMO | Gene of Interest | SRT | His8 | PSC | PSC | PSC | CL7 Sequence file (GenBank format)
Plasmid 34: Enables target proteins to be labeled with C-terminal His8 and CL7. Protease cleavage removes both tags.
- Option 1 - HindIII/SpeI Insertion Site – Trx | SUMO | Gene of Interest | SRT | PSC | PSC | PSC | His8 | CL7 Sequence file (GenBank format)
- Option 2 - KpnI/SpeI Insertion Site – Trx | SUMO | Gene of Interest | SRT | PSC | PSC | PSC | His8 | CL7 Sequence file (GenBank format)
- Option 3 - Bsu36I/SpeI Insertion Site – Trx | SUMO | Gene of Interest | SRT | PSC | PSC | PSC | His8 | CL7 Sequence file (GenBank format)
