Last data update: 04 July 2022 09:25 CEST
Plasmid name: pET-28a-MBP-TEV-Canf7 (LMBP 12828)
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|Price category:||Cat. 2 (cf. price list)|
|Status:||GeneCorner core plasmid|
(View with Genome Compiler)
Canis lupus familiaris NPC intracellular cholesterol transporter 2 cDNA (NPC2, CE1, Can f 7, Can f7, GeneID 403920); mature sequence|
Escherichia coli maltose ABC transporter periplasmic binding protein cDNA (malE, GeneID 948538); mature sequence, N-terminal
Thrombin recognition site; N-terminal
Tobacco etch virus (TEV) protease recognition site; N-terminal
Histidine tag (His-tag); N-terminal
Escherichia coli DNA-binding transcriptional repressor LacI cDNA (lacI, GeneID 945007)
Histidine tag (His-tag); C-terminal
|Promoter:||Phage T7 gene 10 promoter (T7g10) with lac operator (T7lac)
Escherichia coli lac repressor promoter (lacI)
|Ribosome binding site (RBS) of the phage T7 gene 10 (T7g10)|
|Terminator:||Phage T7 gene 10 terminator (T7g10)|
|Selection marker:||Neomycin (neo; kanamycin (kan))|
|Replicon:||Escherichia coli plasmid pMB1 origin
Phage f1 origin
|Host range:||Escherichia coli|
|Further information:||The plasmid was constructed by cloning the dog NPC2 coding sequence, optimised for E. coli, into the BamHI/XhoI opened pET28a-MBP-TEV vector.
The plasmid was designed to express the malE-NPC2 fusion protein in E. coli BL21(DE3). The plasmid encodes for the mature form of the E. coli maltose binding protein, which does not contain a signal sequence, hence the protein will remain in the cytosol.
The N-terminal His-tag can be removed via a thrombin recognition site, while the maltose binding protein can be separated from the dog NPC2 allergen via the TEV protease recognition site. The His-tag at the C-terminal end of the allergen is not functional in this plasmid.
The amino acid sequence of the dog NPC2 allergen corresponds with GenBank accession number NP_001003242.2.
Other name of the plasmid is pET-28a(+)-MBP-TEV-Canf7.
|EMBL Accession number:||NP_001003242.2, view at EMBL, GenBank, DDBJ|
|Latest sequence update:||15/06/2021|
|Authenticity test:||Restriction enzyme pattern analysed at GeneCorner: BamHI/XhoI, BglI/NcoI, BglII/EcoRV, DraI and PvuII.
The dog NPC2 coding sequence has also been sequenced at BCCM/GeneCorner.
|History of deposit:||This plasmid was deposited by Dr. G. Zvirblis (1).
(1) Institute of Biotechnology, Life Science Center, Vilnius University, Vilnius, Lithuania
|Restricted distribution:||- The depositor will be informed of the customer's identity upon release of a sample outside the depositor's department or outside the departments in which BCCM/GeneCorner is embedded, namely UGent-DBMB and VIB-IRC.
- BCCM MTA
|Distributed as:||H/P active culture or plasmid DNA|
|Host for distribution:||Escherichia coli K12 DH5αT1R|
|Related host reference:||Killmann et al., J. Bacteriol. 178 (1996), 6913-6920 [PMID: 8955314] [DOI: 10.1128/jb.178.23.6913-6920.1996]
|Cultivation medium:||LB-Lennox + kanamycin (50 μg/ml)|
|Other culture collection numbers:||-|
Refer in your Materials and Methods:
|pET-28a-MBP-TEV-Canf7 (LMBP 12828) is available at BCCM/GeneCorner. This plasmid was deposited by Dr. G. Zvirblis .|
Note: Up-to-date, validated data are enclosed with the biological material. Nevertheless, these data are a snapshot at a given moment; further updates are always possible.