pSpdCas9-hudTET1CD-T2A-mCherry(PX458)-SghuTSDR-5
(Plasmid #129045)

• Purpose: inactivated control (targeted DNA demethylation_human_TSDR), expression of dCas9-hudTET1CD-T2A-mCherry sgRNA5 targeting human TSDR
• Depositing Lab: Julia K Polansky
• Publication: Kressler et al Front Immunol. 2021 Jan 7;11:609891. doi: 10.3389/fimmu.2020.609891. eCollection 2020.

Backbone

• Vector backbone: PX458
• Backbone manufacturer: Feng Zhang (Addgene plasmid # 48138)
• Vector type: Mammalian Expression

Growth in Bacteria

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

Gene/Insert

• Gene/Insert name: dCas9-hudTET1CD, SgRNA5 (huTSDR)
• Alt name: dCas9-hudTET1CD-T2A-mCherry
• gRNA/shRNA sequence: TGGGTGCTGGTGGATGTGGT
• Species: Synthetic
• Mutation: dCas9 (D10A;H840A), catalytic domain huTET1 inactive (H1671Y; D1673A)
• Promoter: CbH (for dCas9-hudTET1CD-T2A-EGFP) U6 (for sgRNA)
• Tags / Fusion Proteins:
  • HA-Tag, NLS (N terminal on insert)
  • T2A-mCherry (C terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: Unknown (unknown if destroyed)
• 3′ cloning site: Unknown (unknown if destroyed)
• 5′ sequencing primer: SgRNA-cloning-site: hU6-F (GAGGGCCTATTTCCCATGATT)

Resource Information

• Supplemental Documents:
  • 21-pSpdCas9-hudTET1CD-T2A-mCherry-SghuTSDR-5.gb
• A portion of this plasmid was derived from a plasmid made by: mCherry (BbsI removed) from Nicola Patron (Addgene plasmid # 50316); HA tag-NLS-dCas9 from Rudolf Jaenisch (Addgene plasmid # 48223); hudTET1CD from Keith Joung (Addgene plasmid # 49965).

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • Takara Bio Limited Use Label License (formerly Clontech)
• Industry Terms:
  • Not Available to Industry