Skip to main content
This website uses cookies to ensure you get the best experience. By continuing to use this site, you agree to the use of cookies.

Please note: Your browser does not support the features used on Addgene's website. You may not be able to create an account or request plasmids through this website until you upgrade your browser. Learn more

Please note: Your browser does not fully support some of the features used on Addgene's website. If you run into any problems registering, depositing, or ordering please contact us at [email protected]. Learn more

AICSDP-45: TTN-mEGFP
(Plasmid #114412)

Ordering

Item Catalog # Description Quantity Price (USD)
Plasmid 114412 Standard format: Plasmid sent in bacteria as agar stab 1 $85

This material is available to academics and nonprofits only.

Backbone

  • Vector backbone
    pUC57
  • Vector type
    Mammalian Expression, CRISPR ; Donor Template

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
    TTN Homology Arms with linker-mEGFP and Cas9-excisable selection cassette
  • Species
    H. sapiens (human)
  • Insert Size (bp)
    5709
  • Mutation
    homology arms contain point mutations to disrupt crRNA binding sites used, and WTC-specific homozygous SNP(s)
  • Entrez Gene
    TTN (a.k.a. CMD1G, CMH9, CMPD4, CMYP5, EOMFC, HMERF, LGMD2J, LGMDR10, MYLK5, SALMY, TMD)
  • Tag / Fusion Protein
    • linker-mEGFP (C terminal on insert)

Cloning Information

  • Cloning method Unknown

Resource Information

Terms and Licenses

Trademarks:
  • Zeocin® is an InvivoGen trademark.

Depositor Comments

This plasmid has been used with locus-specific CRISPR/Cas9 in a two-step editing process to add a mEGFP tag to the C-terminus of human TTN in WTC human induced pluripotent stem cells by the Allen Institute for Cell Science. After initial HDR, stable expression of CAGGS-driven mCherry will indicate cells with integrated donor sequence. This selection cassette is designed to be removed with subsequent CRISPR/Cas9-mediated excision as previously described (https://doi.org/10.1101/342881). A linker of 3-21 amino acids will be produced after excision. To enrich for edited cells, see our protocol for fluorsecence-assisted cell sorting and subcloning of hiPSCs (https://www.allencell.org/instructional-videos-and-tutorials-for-cell-methods.html). Further, we recommend PCR-based assays for identifying precisely edited clones as previously described (https://www.molbiolcell.org/doi/abs/10.1091/mbc.e17-03-0209).

For more information on the entire plasmid collection, please see https://www.addgene.org/allen-institute-cell-science/.

Please visit https://www.biorxiv.org/content/early/2018/06/09/342881 for the bioRxiv preprint.

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:

    AICSDP-45: TTN-mEGFP was a gift from Allen Institute for Cell Science (Addgene plasmid # 114412 ; http://n2t.net/addgene:114412 ; RRID:Addgene_114412)
  • For your References section:

    Fluorescent Gene Tagging of Transcriptionally Silent Genes in hiPSCs. Roberts B, Hendershott MC, Arakaki J, Gerbin KA, Malik H, Nelson A, Gehring J, Hookway C, Ludmann SA, Yang R, Haupt A, Grancharova T, Valencia V, Fuqua MA, Tucker A, Rafelski SM, Gunawardane RN. Stem Cell Reports. 2019 May 14;12(5):1145-1158. doi: 10.1016/j.stemcr.2019.03.001. Epub 2019 Apr 4. 10.1016/j.stemcr.2019.03.001 PubMed 30956114