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pCM184
(Plasmid #46012)

Ordering

Item Catalog # Description Quantity Price (USD)
Plasmid 46012 Standard format: Plasmid sent in bacteria as agar stab 1 $65

This material is available to academics and nonprofits only.

Backbone

  • Vector backbone
    pAYC61
  • Backbone manufacturer
    Lidstrom Lab (PMID 8021187)
  • Backbone size (bp) 6000
  • Modifications to backbone
    To insert loxP bounded antibiotic resistance gene: The 1.3-kb HincII fragment bearing the kanamycin resistance cassette from pUC4K (Vieira, 1982) was inserted into pLox1 (Palmeros, 2000), which had been cut with XbaI and blunted, to create pCM161. To introduce convenient multiple cloning sites, the loxP-bounded kanamycin cassette of pCM161 was amplified with following primer pair, CM-ufkMCS, 5′-TGACGTCTAGATCTGAATTCAGCTGTACAATTGGTACCATGGATGCATATGGCGGCCGCA-3′, and CM-dfkMCS, 5′-GACTAGTGAGCTCACCGGTTAACACGCGTACGTAGGGCCCGCGGTATCGATAAGCTGGATCC-3′. The resulting 1.4-kb PCR product was purified and cloned into pCR2.1 (Invitrogen, Carlsbad, CA, USA) to create pCM183. To preserve useful cloning sites, pAYC61 was cut with EcoRI and SmaI, blunted using T4 DNA polymerase, and selfligated to produce pCM182. Finally, the 1.4-kb AatII-SpeI fragment from pCM183 containing the loxP-flanked kanamycin cassette was ligated between the AatII and XbaI sites of pCM182 to create pCM184.
  • Vector type
    Cre/Lox
  • Promoter na

Growth in Bacteria

  • Bacterial Resistance(s)
    Ampicillin and Kanamycin
  • Growth Temperature
    37°C
  • Growth Strain(s)
    DH5alpha
  • Copy number
    Unknown

Cloning Information

Resource Information

Depositor Comments

This allelic exchange vector is used with Cre expression vectors (Addgene plasmids 45863 or 45864) to generate unmarked mutants in a broad bacterial host range.

The full plasmid sequence is approximate and there may be small discrepancies present.

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:

    pCM184 was a gift from Mary Lidstrom (Addgene plasmid # 46012 ; http://n2t.net/addgene:46012 ; RRID:Addgene_46012)
  • For your References section:

    Broad-host-range cre-lox system for antibiotic marker recycling in gram-negative bacteria. Marx CJ, Lidstrom ME. Biotechniques. 2002 Nov;33(5):1062-7. 10.2144/02335rr01 PubMed 12449384