pEB2_mScarletI_mVenusNB-MrkHR113A
(Plasmid #182292)

• Purpose: expresses c-di-GMP mutant biosensor in E. coli
• Depositing Lab: Howard Berg
• Publication: Vrabioiu et al Proc Natl Acad Sci U S A. 2022 Feb 8;119(6). pii: 2116830119. doi: 10.1073/pnas.2116830119.

Backbone

• Vector backbone: pEB2
• Backbone manufacturer: J. H. Davis, A. J. Rubin, R. T. Sauer, Design, construction and characterization of a set of insulated bacterial promoters.
• Backbone size w/o insert (bp): 3200
• Total vector size (bp): 5320
• Modifications to backbone: E. Balleza, J. M. Kim, P. Cluzel, Systematic characterization of maturation time of fluorescent proteins in living cells. Nat Methods 15, 47-51 (2018).
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: Low Copy

Gene/Insert

• Gene/Insert name: mVenusNB-MrkHR113A
• Alt name: cyclic di-GMP mutant sensor
• Alt name: c-di-GMP mutant sensor
• Alt name: Sensor*
• Species: Synthetic; K. pneumoniae
• Insert Size (bp): 1401
• Mutation: changed arginine 113 to alanine, MrkH numbering as in PMID: 27551088
• Promoter: proC
• Tag / Fusion Protein:
  • mVenusNB internal fusion to MrkH with a R113A point mutation

Cloning Information

• Cloning method: Ligation Independent Cloning
• 5′ sequencing primer: TG GACATCACCT CTCATAATG
• 3′ sequencing primer: GAG CCT TTC GTT TTA TTT GAT GC

Resource Information

• A portion of this plasmid was derived from a plasmid made by: Maria Schumacher, Duke University School of Medicine, Durham NC

Terms and Licenses

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

Depositor Comments

We obtained the codon optimized MrkH sequence from Maria Schumacher, as described in her publication PMID: 27551088