PB-CAG-dCasRx-RBM25-IRES-BleoR
(Plasmid #221002)

• Purpose: Stable expression of dCasRx-RBM25 when co-transfected with piggyBac transposase. Zeocin selection. CAG promoter. Use with lentiviral gRNA delivery (Addgene #138150) to stably modulate splicing.
• Depositing Labs: Benjamin Blencowe, Mikko Taipale
• Publication: Efficient, specific, and combinatorial control of endogenous exon splicing with dCasRx-RBM25 (unpublished)

Backbone

• Vector backbone: PB_tre_dCas9_VP160 (Addgene #126031)
• Vector type: Mammalian Expression, CRISPR
• Selectable markers: Zeocin

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: dCasRx-RBM25-IRES-BleoR
• Alt name: Human RBM25 fused C-terminal of dCasRx, followed by IRES-BleoR selection cassette
• Species: H. sapiens (human)
• Promoter: CAG
• Tags / Fusion Proteins:
  • NLS (N terminal on insert)
  • NLS-HA (C terminal on insert)

Cloning Information

• Cloning method: Gibson Cloning

Resource Information

• A portion of this plasmid was derived from a plasmid made by: dCasRx coding sequence from Konermann et al., 2018 (Addgene #109050)

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • piggyBac Limited Use Label License
• Industry Terms:
  • Not Available to Industry

Depositor Comments

Co-transfect with piggyBac transposase followed by Zeocin selection to generate polyclonal cells expressing dCasRx-RBM25. We recommend a high ratio of transposon to transposase for highest expression levels. Please see our paper for additional details.