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mutpRL-TK 1x perfect to bulged siRNA
(Plasmid #40765)

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This material is available to academics and nonprofits only.
Item Catalog # Description Quantity Price (USD)
Plasmid 40765 Standard format: Plasmid sent in bacteria as agar stab 1 $85
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Backbone

  • Vector backbone
    mutpRL-TK
  • Backbone manufacturer
    Sharp Lab (http://web.mit.edu/sharplab/RNAi/Cloning.html)
  • Modifications to backbone
    pRL-TK (Promega) was digested with XbaI, and the following ligated in: ctagattccgagatatcggtaatgggcc This produced a modified vector, still containing an XbaI site, but now also containing an ApaI site, to allow for directional cloning of 3' UTR inserts. Final vector: TAATTctagattccgagatatcggtaatgggccCTAGA
  • Vector type
    Luciferase

Growth in Bacteria

  • Bacterial Resistance(s)
    Ampicillin, 100 μg/mL
  • Growth Temperature
    37°C
  • Growth Strain(s)
    Top10
  • Copy number
    High Copy

Gene/Insert

  • Gene/Insert name
    CXCR4 small RNA perfect to bulged target site
  • Mutation
    mutated from TAG to CTC at nt 387–389 of the Renilla luciferase open reading frame to generate mismatches with seed positions 5, 6, and 7 of the siRNA guide strand by PCR-directed mutagenesis
  • Promoter TK
  • Tag / Fusion Protein
    • luciferase (N terminal on backbone)

Cloning Information

  • Cloning method Restriction Enzyme
  • 5′ cloning site XbaI (not destroyed)
  • 3′ cloning site ApaI (not destroyed)
  • 5′ sequencing primer CCAATGCTATTGTTGAAGGTGCCAAG
  • 3′ sequencing primer CACTGCATTCTAGTTGTGGTTTGTCC
    • (Common Sequencing Primers)

Terms and Licenses

  • Academic/Nonprofit Terms
  • Industry Terms
    • Not Available to Industry
Trademarks:
  • Zeocin® is an InvivoGen trademark.
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:

    mutpRL-TK 1x perfect to bulged siRNA was a gift from Phillip Zamore (Addgene plasmid # 40765 ; http://n2t.net/addgene:40765 ; RRID:Addgene_40765)

  • For your References section:

    Argonaute protein identity and pairing geometry determine cooperativity in mammalian RNA silencing. Broderick JA, Salomon WE, Ryder SP, Aronin N, Zamore PD. RNA. 2011 Oct;17(10):1858-69. Epub 2011 Aug 30. 10.1261/rna.2778911 PubMed 21878547
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