pEC3112 (pSpy2C)
(Plasmid #220251)

• Purpose: (Empty Backbone) replicative E. coli-S. pyogenes shuttle plasmid for heterologous gene expression
• 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: pSpy2C
• Backbone size (bp): 6869
• Modifications to backbone: See associated article for detailed construction information.
• Vector type: Bacterial Expression

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, use 5 µg/mL chloramphenicol in THY medium. The plasmid has high copy numbers in E. coli and low copy numbers in S. pyogenes.
• Copy number: High Copy

Cloning Information

• Cloning method: Golden Gate
• 5′ sequencing primer: GTAAAACGACGGCCAGTC
• 3′ sequencing primer: CAGGAAACAGCTATGAC

Terms and Licenses

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

Depositor Comments

The multiple cloning site (EcoRI, XbaI, SpeI and PstI) contains the Plac_mrfp cassette. The mrfp reporter allows for red-white screening during sub-cloning in E. coli (DH5a). When the Plac_mrfp cassette is replaced with the insert of interest, colonies will appear white.

Resource Information: The chloramphenicol resistance cassette was taken from pIB166 (Plasmid #90189) and the MCS featuring the Plac_mrfp cassette from pBS3Clux (Plasmid #55172). The pUC origin for replication in E. coli was taken from pIB184-Km (Plasmid #90195) and the repDEG replicon for replication in Gram-positives from pMSP3535 (Plasmid #46886)

Please download the file linked below to obtain a well annotated map of the plasmid.