hHS1-M7A,H102A
(Plasmid #159171)

• Purpose: Human codon-optimized cytosolic labile heme reporter
• Depositing Lab: Amit Reddi
• Publication: Hanna et al Proc Natl Acad Sci U S A. 2016 Jul 5;113(27):7539-44. doi: 10.1073/pnas.1523802113. Epub 2016 May 31.

Backbone

• Vector backbone: pcDNA3-eGFP
• Backbone manufacturer: Doug Golenbock (Addgene #13031)
• Backbone size w/o insert (bp): 6146
• Total vector size (bp): 7906
• Vector type: Mammalian Expression

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: hHS1-M7A,H102A
• Species: H. sapiens (human)
• Insert Size (bp): 1760
• Mutation: Mutated both Met 7 and His 102 of the cyt b562 module of hHS1 to Ala (M7A and H102A, respectively), creates weak affinity heme sensor.
• Promoter: CMV

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: BamHI (not destroyed)
• 3′ cloning site: XhoI (not destroyed)
• 5′ sequencing primer: CMV-F or T7

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • Evrogen Limited Use Label License for FPs
• Industry Terms:
  • Not Available to Industry

Depositor Comments

The first-generation heme sensor, HS1, consists of a heme-binding domain, the His/Met coordinating 4-alpha-helical bundle hemoprotein cytochrome b562 (Cyt b562) (13), fused to a pair of fluorescent proteins, EGFP and Katushka 2 (mKATE2)