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pCS2+MT-hToca-1-wt
(Plasmid #33030)

Ordering

Item Catalog # Description Quantity Price (USD)
Plasmid 33030 Standard format: Plasmid sent in bacteria as agar stab 1 $85

This material is available to academics and nonprofits only.

Backbone

  • Vector backbone
    pCS2+MT
  • Backbone size w/o insert (bp) 4350
  • Vector type
    Mammalian Expression

Growth in Bacteria

  • Bacterial Resistance(s)
    Ampicillin, 100 μg/mL
  • Growth Temperature
    37°C
  • Growth Strain(s)
    DH5alpha
  • Copy number
    High Copy

Gene/Insert

  • Gene/Insert name
    Toca-1
  • Species
    H. sapiens (human)
  • Entrez Gene
    FNBP1L (a.k.a. C1orf39, TOCA1)
  • Promoter simian CMV IE94
  • Tag / Fusion Protein
    • 6xMyc (N terminal on backbone)

Cloning Information

  • Cloning method Restriction Enzyme
  • 5′ cloning site none (unknown if destroyed)
  • 3′ cloning site none (unknown if destroyed)
  • 5′ sequencing primer SP6
  • 3′ sequencing primer M13 reverse
  • (Common Sequencing Primers)

Resource Information

Terms and Licenses

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

Depositor Comments

Molecular cloning of Toca-1 was difficult, because the cDNA encoding human Toca-1 is lethal to E. coli. The toxicity problem was not resolved by using vectors with low copy numbers or with minimal transcriptional activity, suggesting that the source of toxicity was not the translated protein product but rather the DNA sequence itself. The toxic region of the Toca-1 cDNA was localized to codon 127. The Xenopus tropicalis homolog of Toca-1 is completely nontoxic when cloned using E. coli and is identical in amino acid sequence to human Toca-1 between amino acids 124 and 127, but these amino acids are encoded by different codons. Replacing codons 126 and 127 in the human Toca-1 with the X. Tropicalis sequence (which constitute silent mutations) completely ameliorated the toxicity problem and allowed us to propagate the human Toca-1 clone in E. coli without difficulty.

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:

    pCS2+MT-hToca-1-wt was a gift from Marc Kirschner (Addgene plasmid # 33030 ; http://n2t.net/addgene:33030 ; RRID:Addgene_33030)
  • For your References section:

    Toca-1 mediates Cdc42-dependent actin nucleation by activating the N-WASP-WIP complex. Ho HY, Rohatgi R, Lebensohn AM, Le Ma, Li J, Gygi SP, Kirschner MW. Cell. 2004 Jul 23;118(2):203-16. 10.1016/j.cell.2004.06.027 PubMed 15260990