Coding

Part:BBa_K4632018

Designed by: Haolong Lai   Group: iGEM23_SCAU-China   (2023-10-09)


LM35 (T222I)


LM35 (T222I) is a mutant of LasR prepared through error-prone polymerase chain reaction (mutagenesis), aiming to reduce its sensitivity to 3OC8HSL and 3OC12HSL to avoid activation by autoinducers produced by EsaI.

LM35 exhibits the following characteristics:

(1) Reduced Sensitivity to 3OC12HSL: LM35 displays a significantly reduced sensitivity to 3OC12HSL, requiring a higher concentration of 3OC12HSL compared to the wild-type LasR to achieve half-saturated expression of the las promoter.

(2)Sensitivity to 3OC8HSL: LM35 exhibits a higher sensitivity to 3OC8HSL and can be activated when the concentration of 3OC8HSL is greater than 1000 nM.

(3)Sensitivity to 3OC6HSL: LM35 also shows reduced sensitivity to 3OC6HSL and cannot be activated by 3OC6HSL.

What have we done? (SCAU-China 2023)

We optimized the codons of the provided sequence for Escherichia coli expression. We successfully characterized its performance in E. coli TOP10. For more detailed characterization results, please refer to our composite_part-BBa_K4632024[1]

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal XbaI site found at 21
    Illegal PstI site found at 113
    Illegal PstI site found at 363
    Illegal PstI site found at 521
    Illegal PstI site found at 599
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 113
    Illegal PstI site found at 363
    Illegal PstI site found at 521
    Illegal PstI site found at 599
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 633
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal XbaI site found at 21
    Illegal PstI site found at 113
    Illegal PstI site found at 363
    Illegal PstI site found at 521
    Illegal PstI site found at 599
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal XbaI site found at 21
    Illegal PstI site found at 113
    Illegal PstI site found at 363
    Illegal PstI site found at 521
    Illegal PstI site found at 599
  • 1000
    COMPATIBLE WITH RFC[1000]

References

Jiang W, He X, Luo Y et al. Two Completely Orthogonal Quorum Sensing Systems with Self-Produced Autoinducers Enable Automatic Delayed Cascade Control[J]. ACS SYNTH BIOL, 2020,9(9):2588-2599.



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