Part:BBa_K3634007:Design
Heme Oxygenase (ho1) (Optimised for E.coli)
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 390
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
The sequence from BBa_K2328062 was taken and subject to the IDT codon optimisation tool. As a result of optimisation, two illegal restriction sites were introduced to the part. A PstI site at 300bp was removed by in silico site mutagenesis (t297a, TCT (Ser) to TCA (Ser)). The second illegal restriction site, EcoR1, at 469bp was removed also by in silico site mutagenesis (a471g, GAA (Glu) to GAG (Glu)). This allowed for the part to be used at RFC[10] and RFC[1000] standard with codon optimisation.
Source
The ho1 gene is found in Synechocystis sp. PCC 6803 and has been previously categorised in BioBrick form by J.J. Tabor (BBa_I15008). In 2017, TJU China removed the pre-existing barcode attached to the sequence (BBa_K2328062) before codon optimising the part for intestinal bacteria (BBa_K2328003). Although these intestinal bacteria included E.coli, the St Andrews iGEM 2020 team decided to specifically optimise BBa_K2328062 for E.coli only using the IDT codon optimisation tool to allow maximum expression in their chosen E.coli chassis.
References
UTAustin iGEM 2004 - https://parts.igem.org/Part:BBa_I15008
TJU China iGEM 2017 - https://parts.igem.org/Part:BBa_K2328062
IDT Codon Optimisation Tool - https://eu.idtdna.com/CodonOpt
UniProtKB: P72849 (HO1_SYNY3) - https://www.uniprot.org/uniprot/P72849#