pJC.15A.sgRNA.TA
(Plasmid #232483)

• Purpose: sgRNA cloning backbone for expression of sgRNA in C. difficile. Contains TA system to enable maintenence without antibiotic selection.
• Depositing Lab: James Collins
• Publication: Collins et al Microbiology Resource Announcements

Backbone

• Vector backbone: pRPF185
• Vector type: Bacterial Expression, CRISPR

Growth in Bacteria

• Bacterial Resistance(s): Chloramphenicol, 25 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Growth instructions: Does not require antibiotic maintenance following conjugation into C. difficile.
• Copy number: Unknown

Gene/Insert 1

• Gene/Insert name: Toxin-antitoxin System from CD630
• Species: C. difficile
• Insert Size (bp): 615
• Promoter: native promoter

Cloning Information for Gene/Insert 1

• Cloning method: Other
• 5′ sequencing primer: cccactttcacagccgcctcaatacttataaag
• 3′ sequencing primer: ATGATTATTCATTTTTTTTATATAAACAATGAAATTCAAG

Gene/Insert 2

• Gene/Insert name: medium-copy-number p15A origin of replication
• Alt name: p15A
• Insert Size (bp): 546
• Promoter: native promoter

Cloning Information for Gene/Insert 2

• Cloning method: Other
• 5′ sequencing primer: tgctagattctaaaatagCAAATTAAGCAGAAGGCCATCC
• 3′ sequencing primer: aggcggtaatacggttcggcctaggagatacttaaca

Gene/Insert 3

• Gene/Insert name: mCherry
• Promoter: P3 constitutive promoter

Cloning Information for Gene/Insert 3

• Cloning method: Other
• 5′ sequencing primer: TATATAAAAAAAATGAATAATCATTGATTGCTCGAGC

Terms and Licenses

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

Depositor Comments

pJC.15A.sgRNA.TA carries the gene encoding the red fluorescent protein mCherry and a toxin-antitoxin system that enables plasmid maintenance without antibiotics. The mCherry gene is flanked by two BsmBI sites to allow for easy replacement with sgRNA.