Part:BBa_K1597002

tasA, amyloid-like fibers

tasA is resposible for the amyloid-like fibers that are one of the main constituents of the extracellular matrix of Bacillus subtilis biofilms. After the expression of the tapA-sipW-tasA operon, TasA amyloid-like fibres are synthesised. These fibers attach to the cell wall. Together with extracellular polysaccharides, TasA amyloid-like fibers promote formation of cell clusters, where these amyloid-like fibers resemble bundles of cell chains. This effect results in a more robust biofilm.

tasA is responsible for amyloid-like fibers that are one of the main constituents of the extracellular matrix of B. subtilis biofilms. In our construct we added the salt inducible promoter before tasA before integration in B. subtilis. With this construct we want to induce the production of tasA with different concentration salt.

Both B. subtilis ComI strain with tasA overexpression (TASA) and B. subtilis ComI (COMI)were grown on Msgg media with or without salt. After 24 and 48 hours thioflavin S (which is an amyloid fiber staining) was added to the biofilms. After 15 min incubation at room temperature the biofilms were photographed with white light and fluorescence.

With the induction of salt, TASA shows more amyloid fibers are present after 24 hours.This is not the case for COMI. After 48 hours the differences are less visible, which could be explained that tasA is a natural gene in bacillus which is expressed in a later phase of biofilm production. The natural gene is not yet fully activated after 24 hours, where it is with salt induction in the TASA. This means that the salt promoter is activated with salt and produces extra tasA.

Another method is to measure tasA expression with thioflavin S over time in both B. subtilis comI strain with tasA overexpression (tasA) and B. subtilis comI (ComI). Both strains were grown on SSM media with different concentrations of salt, ranging from 0M salt up to 1 M salt. Over time the fluorescence and the OD were measured. The plates were incubated during measurement an in between at 37 °C. Strains grown on a salt concentration above 0,5M showed little growth. Therefore these results are not shown here. A bar chart was plotted after 1 hour and after 3 hours for both B. subtilis comI and tasA mutant. After one hour the tasA mutant show a higher fluorescence than the B. subtilis comI control. After 3 hours the tasA mutant shows, with an induction of 0,1M; 0,2M and 0,3M salt, a significant increase than the control. This confirms that both the salt-inducible <a href="https://parts.igem.org/Part:BBa_K1597000">(BBa_K1597000)</a> and the tasA biobrick function in B. subtilis.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]