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pRK792
(Plasmid #8830)

Ordering

Item Catalog # Description Quantity Price (USD)
Plasmid 8830 Standard format: Plasmid sent in bacteria as agar stab 1 $65

This material is available to academics and nonprofits only.

Backbone

  • Vector backbone
    pMal-C2
  • Backbone manufacturer
    New England Biolabs
  • Backbone size w/o insert (bp) 6700
  • Vector type
    Bacterial Expression

Growth in Bacteria

  • Bacterial Resistance(s)
    Chloramphenicol and Ampicillin
  • Growth Temperature
    37°C
  • Growth Strain(s)
    BL21(DE3)-RIL
  • Growth instructions
    Shifting the temperature from 37C to 30C during induction maximizes the yield of soluble TEV protease. Ampicillin for pRK792; Chloramphenicol for pRIL
  • Copy number
    Low Copy

Gene/Insert

  • Gene/Insert name
    TEV protease, S219P mutant
  • Alt name
    tobacco etch virus protease
  • Insert Size (bp)
    750
  • Mutation
    S219P mutation
  • Tags / Fusion Proteins
    • MBP (cleaved) (N terminal on backbone)
    • His (N terminal on insert)
    • polyarginine (C terminal on insert)

Cloning Information

  • Cloning method Restriction Enzyme
  • 5′ cloning site SacI (not destroyed)
  • 3′ cloning site BamHI (not destroyed)
  • 5′ sequencing primer N/A
  • 3′ sequencing primer M13-F20
  • (Common Sequencing Primers)

Resource Information

  • A portion of this plasmid was derived from a plasmid made by
    ATCC
  • Terms and Licenses
  • Articles Citing this Plasmid

Depositor Comments

pRK792 overproduces the catalytic domain of tobbaco etch virus (TEV) protease in the form of an MBP fusion protein that cleaves itself in vivo to yield a TEV protease catalytic domain with an N-terminal His-tag and a C-terminal polyarginine tag. The TEV protease produced by this strain is the autoinactivation-resistant mutant S219P.The pRIL plasmid (Stratagene) improves the yield of TEV protease by increasing the supply of argU tRNA in the cell (for AGG and AGA codons). The catalytic activity of the S219P mutant is approximately 2-fold less than that of the wild-type protease. Bacterial strain: E.coli BL21(DE3)-RIL (Stratagene).

How to cite this plasmid ( Back to top)

These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the plasmids were described, and include Addgene in the Materials and Methods of your future publications.

  • For your Materials & Methods section:

    pRK792 was a gift from David Waugh (Addgene plasmid # 8830 ; http://n2t.net/addgene:8830 ; RRID:Addgene_8830)
  • For your References section:

    Tobacco etch virus protease: mechanism of autolysis and rational design of stable mutants with wild-type catalytic proficiency. Kapust RB, Tozser J, Fox JD, Anderson DE, Cherry S, Copeland TD, Waugh DS. Protein Eng. 2001 Dec . 14(12):993-1000. 10.1093/protein/14.12.993 PubMed 11809930