pEC2861 (pSpy1C-PgyrA_ffluc)
(Plasmid #218511)

• Purpose: replicative E. coli-S. pyogenes shuttle plasmid, constitutive expression of firefly luciferase reporter
• Depositing Lab: Emmanuelle Charpentier
• Publication: Lautenschlager et al Front Bioeng Biotechnol. 2024 Jun 7;12:1395659. doi: 10.3389/fbioe.2024.1395659. eCollection 2024.

Backbone

• Vector backbone: pSpy1C
• Total vector size (bp): 5773
• Modifications to backbone: The Plac_mrfp cassette on pSpy1C was replaced with the PgyrA promoter from S. pyogenes and the ffluc reporter gene from pLZ12Km2-P23R:TA:ffluc. See associated publication for detailed construction information.
• Vector type: Luciferase

Growth in Bacteria

• Bacterial Resistance(s): Chloramphenicol, 25 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Growth instructions: For E.coli, use 20 µg/mL chloramphenicol in LB medium. For S. pyogenes, also use 20 µg/mL chloramphenicol in THY medium.
• Copy number: Low Copy

Gene/Insert

• Gene/Insert name: ffluc
• Species: Photinus pyralis
• Insert Size (bp): 1653
• Mutation: deletion of Plac_mrfp cassette
• GenBank ID: XM_031473197
• Promoter: PgyrA

Cloning Information

• Cloning method: Gibson Cloning
• 5′ sequencing primer: GTAAAACGACGGCCAGTC
• 3′ sequencing primer: CAGGAAACAGCTATGAC

Resource Information

• Supplemental Documents:
  • pSpy1C-PgyrA_ffluc.gb

Terms and Licenses

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

Depositor Comments

This high-copy replicative shuttle plasmid features the constitutive PgyrA promoter from S. pyogenes driving expression of the Firefly luciferase reporter. Luminescence can be detected by adding luciferin to the culture. See associated publication for more details.

Resource Information: The PgyrA promoter was taken from the genome of S. pyogenes SF370. The ffluc reporter gene was taken from pLZ12Km2-P23R:TA:ffluc (Addgene plasmid #88900).

Please download the detailed plasmid map using the file linked below.