pKSJ340
(Plasmid #27125)

• Depositing Lab: Alan Finkelstein
• Publication: Jakes et al Mol Microbiol. 2010 Feb;75(3):567-78. doi: 10.1111/j.1365-2958.2009.06966.x. Epub 2009 Nov 17.

Backbone

• Vector backbone: PUC19
• Backbone manufacturer: New England Biolabs
• Backbone size w/o insert (bp): 2686
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): XL10 Gold
• Growth instructions: The plasmid can be expressed in any recA+ strain; it is currently supplied in XL10 Gold (recA-) for optimal plasmid preparations.
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: Colicin Ia + Ia immunity protein
• Species: E. coli
• Insert Size (bp): 2636

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: EcoRI (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: M13_puc_Fwd
• 3′ sequencing primer: M13_puc_Rev

Resource Information

• A portion of this plasmid was derived from a plasmid made by: From Partho Ghosh, Stroud Lab, UCSF
• Article Citing this Plasmid:
  • 1 Reference

Terms and Licenses

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

Depositor Comments

The original clone had some mutations that had been introduced during the PCR cloning of the colicin operon (into another vector). Here, the operon was moved into pUC19 and the mutations were corrected, so that the sequence is the same as the one in GenBank (M13819.1)

This plasmid contains Colicin Ia + Ia immunity protein, each with their own promoters; colicin production can be induced with mitomycin C.

The SOS promoter for colicin expression is at around residues 148-165 of the insert, in the forward direction. The immunity promoter -35 box is about 460 bp from the end of the insert; there are only about 20 bp between the terminator for the colicin gene, reading in one direction, and the terminator for the immunity protein, reading in the other.