pKM586
(Plasmid #19978)

• Depositing Lab: David Waugh
• Publication: Kapust et al Protein Expr Purif. 2000 Jul . 19(2):312-8.

Backbone

• Vector backbone: pZS*24-MCS1
• Backbone manufacturer: Herman Bujard
• Backbone size w/o insert (bp): 4300
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 30°C
• Growth Strain(s): BL21-Pro
• Growth instructions: 30 degrees
• Copy number: Low Copy

Gene/Insert

• Gene/Insert name: TEV protease
• Species: tobacco etch virus
• Insert Size (bp): 900
• Mutation: S219D mutant
• Entrez Gene: TEVgp1 (a.k.a. TEVgp1)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: HindIII (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: n/a

Resource Information

• A portion of this plasmid was derived from a plasmid made by: A pZ derivative from - Lutz, R. and Bujard, H. (1997). Independent and tight regulation of transcriptional units in Escherichia coli via the LacR/O, the TetR/O and AraC/I1-I2 regulatory elements. Nucl. Acids Res. 25: 1203-10.

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

This TEV protease expression vector is intended to be used for controlled intracellular processing of fusion proteins (TEV protease substrates) in E. coli. It is supplied in BL21-Pro cells so that transcription of TEV protease can be regulated by anhydrotetracycline.

Replicon: pSC101
Promoter: Synthetic ªPL/tetO

Inducer: anhydrotetracycline (100 ºg/liter)
Notes: Expression of the TEV protease gene on pKM586 will be constitutive unless the host strain produces Tet repressor. Regulated expression can be achieved in DH5alphaPro or BL21Pro cells (Clontech). For best results, perform intracellular processing experiments at 30 °C.