pX330-U6-(negative selection module)-Chimeric_BB-CBh-hSpCas9
(Plasmid #223321)

• Purpose: It contains a ccdB-CMr negative selection module flanked by BpiI (BbsI) sites that allows excluding clones without correct insertion of sgRNA. Human codon optimized Cas9 expressing plasmid.
• Depositing Lab: Jonathan Arias
• Publication: Petruškevičiūtė et al Stem Cell Res. 2024 Dec 12:103628. doi: 10.1016/j.scr.2024.103628.

Backbone

• Vector backbone: pX330_42230
• Vector type: Mammalian Expression, CRISPR, Synthetic Biology

Growth in Bacteria

• Bacterial Resistance(s): Chloramphenicol and Ampicillin, 25 & 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): ccdB Survival
• Copy number: High Copy

Gene/Insert 1

• Gene/Insert name: SpCas9 codon optimized for human
• Species: H. sapiens (human), Synthetic
• Insert Size (bp): 4125
• Entrez Gene: cas9 (a.k.a. SD90_RS03825, SD90_03830)
• Promoter: CBh

Cloning Information for Gene/Insert 1

• Cloning method: Other
• 5′ sequencing primer: ccdB-fw

Gene/Insert 2

• Gene/Insert name: chloramphenicol resistance protein
• Species: Synthetic
• Insert Size (bp): 681
• GenBank ID: ABO28845.1

Cloning Information for Gene/Insert 2

• Cloning method: Gibson Cloning
• 5′ sequencing primer: hU6-F

Gene/Insert 3

• Gene/Insert name: CcdB
• Species: Synthetic
• Insert Size (bp): 306
• GenBank ID: ABO28846.1

Cloning Information for Gene/Insert 3

• Cloning method: Gibson Cloning
• 5′ sequencing primer: hU6-F

Resource Information

• Supplemental Documents:
  • 223321_pX330-U6-(negative selection module)-Chimeric_BB-CBh-hSpCas9.gb
• A portion of this plasmid was derived from a plasmid made by: Expression vector from Cong et al. 10.1126/science.1231143 that carries the chloramphenicol resistance protein ABO28845.1 and the negative selection marker ccdB ABO28846.1 for efficient cloning of sgRNA targeting sequences.

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

Expression vector from Cong et al. 10.1126/science.1231143 that carries the chloramphenicol resistance protein ABO28845.1 and the negative selection marker ccdB ABO28846.1 for efficient cloning of sgRNA targeting sequences.