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pMSP3535VA
(Plasmid #46887)

Ordering

Item Catalog # Description Quantity Price (USD)
Plasmid 46887 Standard format: Plasmid sent in bacteria as agar stab 1 $75

This material is available to academics and nonprofits only.

Backbone

  • Vector backbone
    pMSP3535VA
  • Backbone size (bp) 8278
  • Modifications to backbone
    See associated article for detailed construction information
  • Vector type
    Bacterial Expression ; Nisin-inducible expression; Gram-positive Bacterial Shuttle Vector
  • Promoter PnisA

Growth in Bacteria

  • Bacterial Resistance(s)
    Kanamycin
  • Growth Temperature
    30°C
  • Growth Strain(s)
    Stbl3
  • Growth instructions
    For E. coli, grow on Luria Broth (LB) agar containing 50 μg/mL kanamycin. For E. faecalis, grow on Brain Heart Infusion (BHI) medium containing 1000 μg/mL kanamycin.
  • Copy number
    Low Copy

Cloning Information

  • Cloning method Restriction Enzyme
  • 5′ sequencing primer Unknown
  • 3′ sequencing primer T7; pBR322ori-F (5'-GGGAAACGCCTGGTATCTTT-3')
  • (Common Sequencing Primers)

Resource Information

  • Addgene Notes
  • A portion of this plasmid was derived from a plasmid made by
    The pNisA promoter and the nisin-sensing genes come from the Netherlands Dairy Research Institute (NIZO) (de Ruyter, Kuipers, and de Vos, 1996. PMID: 8837421)
  • Terms and Licenses
  • Industry Terms
    • Not Available to Industry
  • Article Citing this Plasmid

Depositor Comments

This plasmid utilizes the NIsin Controlled Expression (NICE) system. NICE® is a registered trademark of NIZO food research BV, P.O.Box 20, 6710 BA Ede, The Netherlands.

pMSP3535VA contains the Nisin-inducible PnisA promoter, and the pVA380-1 replicon for expression in gram-positive bacteria.

For protein expression in E. faecalis, induce with 10-25 ng/mL nisin (Sigma, N-5764). See associated article for instructions on how to make nisin solution.

Note: Nucleotide sequence encoding the signal sequence and one-third of the mature NisA (nisin) peptide remains associated with the PnisA promoter. The depositing laboratory recommends adding of a Stop codon to the 5' end of inserted sequences to avoid unintentional gene fusions.

Note: The theoretical full sequence uploaded here is NOT fully accurate - the NheI site at position 4414 has been lost, along with approximately 500bp of sequence in this region (compare the depositor's uploaded maps). The actual size of this plasmid is 8278 as listed here. The depositing lab has not resequenced the region in question, but the discrepancy does not affect plasmid function.

Note: This plasmid has been shown to accumulate deletions when replicated in E. coli (or gram-negative bacteria). Screening transformants by restriction digest is recommended.

Addgene's quality control sequencing finds discrepancies with the depositor's provided sequence, but they are not thought to affect plasmid function.

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:

    pMSP3535VA was a gift from Gary Dunny (Addgene plasmid # 46887 ; http://n2t.net/addgene:46887 ; RRID:Addgene_46887)
  • For your References section:

    Improved vectors for nisin-controlled expression in gram-positive bacteria. Bryan EM, Bae T, Kleerebezem M, Dunny GM. Plasmid. 2000 Sep;44(2):183-90. 10.1006/plas.2000.1484 PubMed 10964628