Team:Edinburgh OG/Improve

PhagED: a molecular toolkit to re-sensitise ESKAPE pathogens

 

 

 

 

 

Our Best Improved Part ! ! !
Registry Number Brief Description Key Details Length Sequence information
BBa_K2739000 ProR-PhaR The phasin autoregulation system with native promoter 658bp Original Sequence obtained from: NCBI(AM260479.1) and Monitoring and kinetic analysis of the molecular interactions by which a repressor protein, PhaR, binds to target DNAs and poly[(R)-3-hydroxybutyrate] (2013) . Sequence is codon optimised for E.coli.
Previous Parts
BBa_K108015 PhaR (From previous team) The phasin autoregulation system 552bp Source of sequence :Regulation of phasin expression and polyhydroxyalkanoate (PHA) granule formation in Ralstonia eutropha H16 (2002) and NCBI (No further indication). No codon optimisation for E.coli. No result or user experience provided.
BBa_K108014 ProP (From previous team) The native promoter and RBS of phasin autoregulation system 234bp Source of sequence :Regulation of phasin expression and polyhydroxyalkanoate (PHA) granule formation in Ralstonia eutropha H16 (2002) and NCBI (No further indication). No codon optimisation for E.coli. No result or user experience provided.

The sequence of PhaR promoter is more properly selected in comparison with the previous parts. Previous publication: Monitoring and kinetic analysis of the molecular interactions by which a repressor protein, PhaR, binds to target DNAs and poly[(R)-3-hydroxybutyrate] (2013) is served as reference guideline and the selected functional promoter sequence is 134bp shorter than the previous part.

The template sequence of PhaR in this project is based on NCBI(AM260479.1), being 4bp difference to the previous part.

Figure 1. Sequence alignment of NCBI(AM260479.1) and BBa_K108015. The 4bp difference is indicated by red color.

The sequence is codon optimised to E.coli via IDT. (See the Registry design page for the detail alignment)

This composite part has allow us to investigate A) its relation in PHA production through PHA operon; B) its function on PHA production with Phasin (BBa_K2739005); C) Assess the secretion function of Phasin-HlyA is compatible with phaR and if the improvement of phaR of PHA production is remained in this parts composition (BBa_K2739006);.

Figure 2. Diagram of other PhaR Biobricks development.