Coding

Part:BBa_K4212008:Design

Designed by: Shirin Bamezai   Group: iGEM22_Imperial_College_London   (2022-09-29)


GerA_operon


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 3506
    Illegal PstI site found at 521
    Illegal PstI site found at 3457
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 3506
    Illegal PstI site found at 521
    Illegal PstI site found at 3457
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 3506
    Illegal BglII site found at 3038
    Illegal XhoI site found at 628
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 3506
    Illegal PstI site found at 521
    Illegal PstI site found at 3457
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 3506
    Illegal PstI site found at 521
    Illegal PstI site found at 3457
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 2693


Design Notes

Codon optimised to remove recognition sites for BbsI, BsaI and BsmbI to enable assembly in Golden Gate system.

Usage and Biology

Spores are known to respond by initiating the germination cascade to a class of nutrients called germinants, including L-alanine, asparagine and simple sugars such as glucose or fructose. The putative set of proteins responsible for the recognition of these germinants and the subsequent initiation of the germination cascade are known as germinant receptors, sitting on the inner membrane of the spore. In B.subtilis, three receptors are believed to control the majority of germination: GerA, GerB and GerK. The former is capable of inducing complete germination of the bacterial spore in response to L-alanine alone, while the other two require stimulation from a variety of different germinants and their signaling often doesn’t result in full germination. Due to its orthogonality towards a single germinant, in addition to its superior germination-inducing capacities, GerA was chosen in this project as the target for the development of a chitin-sensing receptors.

Figure 8. The schematic design of GerA

Source

Native sequence from B. subtilis genome.

References

[1] Paidhungat, M. & Setlow, P. (2000) Role of Ger Proteins in Nutrient and Nonnutrient Triggering of Spore Germination in Bacillus subtilis. Journal of Bacteriology. 182 (9), 2513–2519. doi:10.1128/JB.182.9.2513-2519.2000. [2] Igarashi, T. & Setlow, P. (2005) Interaction between Individual Protein Components of the GerA and GerB Nutrient Receptors That Trigger Germination of Bacillus subtilis Spores. Journal of Bacteriology. 187 (7), 2513–2518. doi:10.1128/JB.187.7.2513-2518.2005.