Coding

Part:BBa_K2010002:Design

Designed by: William Cho   Group: iGEM16_Harvard_BioDesign   (2016-06-21)
Revision as of 06:52, 8 October 2016 by Williamcho (Talk | contribs) (→‎Source)

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pelB + PETase (PET-degrading enzyme, origin I. sakaiensis)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 112
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 528


Design Notes

This sequence was optimized for E. coli K12 using the Codon Optimization Tool from the IDT website.

The T7 promoter was used in order to have strong, inducible transcription, in conjunction with NEB’s T7 Express lysY/Iq Competent E. coli.

RBS 34 was used for standard efficiency.

The GGS linker was used to prevent the His-tag from interfering with the folding of PETase.

The His-tag was included for His-tag purification.

The pelB tag was included for excretion of PETase.

Source

T7 promoter: BBa_I712074

RBS: BBa_B0034

PETase, originally named ISF6_4831, comes from Ideonella sakaiensis, identified in "A bacterium that degrades and assimilates poly(ethylene terephthalate)" (Yoshida et al. 2016).

References