Difference between revisions of "Part:BBa K3989010"
Line 9: Line 9:
 
[[File:ClyA_in_omp8_OMV.jpeg|500px]]
 
[[File:ClyA_in_omp8_OMV.jpeg|500px]]
  
−
Figure 1. ClyA protein with or without elicitor fused exists in outer membrane vesicles of E. coli omp8 strain.
+
Figure 1. ClyA protein with or without elicitor fused exists in outer membrane vesicles of <i>E. coli<i> omp8 strain.
  
−
The immunogenicity of engineered OMVs was tested by Seedling Growth Inhibition (SGI) assay (Figure 2 & 3）, in which lower seedling weight means higher immune responses are triggered (due to the trade-off between immunity and growth). Two OMV-producing <i>E. coli<i> strains were used, omp8 and <i>tolB<i>. In both strain, ClyA-flg22 and ClyA-elf18 can trigger stronger immune responses.
+
The immunogenicity of engineered OMVs was tested by Seedling Growth Inhibition (SGI) assay (Figure 2 & 3）, in which lower seedling weight indicates that higher immune responses are triggered (due to the trade-off between immunity and growth). Two OMV-producing <i>E. coli<i> strains were used, omp8 and <i>tolB<i>. In both strain, ClyA-flg22 and ClyA-elf18 can trigger stronger immune responses.
  
 
[[File:omp8 OMVs on col-0.jpeg|500px]]
 
[[File:omp8 OMVs on col-0.jpeg|500px]]

Revision as of 17:49, 16 October 2021


plant immune elicitor flg22 fused with ClyA for OMV display

In this construct, outer membrane-associated protein ClyA (Part:BBa_K811000) is used to display the plant immune elicitor flg22 (Part:BBa_K3286138) on the surface of bacterial outermembrane vesicles (OMVs) to trigger plant immune responses. To make flg22 better recognized by plant immune receptor FLS2, we put mutiple linkers and a 3×FLAG tag between ClyA and flg22 to minimize the steric hindrance.

By doing western blot using anti-FLAG antibody, we proved that ClyA-elicitor fusion proteins indeed exist in OMVs (Figure 1).

ClyA in omp8 OMV.jpeg

Figure 1. ClyA protein with or without elicitor fused exists in outer membrane vesicles of E. coli<i> omp8 strain.

The immunogenicity of engineered OMVs was tested by Seedling Growth Inhibition (SGI) assay (Figure 2 & 3）, in which lower seedling weight indicates that higher immune responses are triggered (due to the trade-off between immunity and growth). Two OMV-producing <i>E. coli<i> strains were used, omp8 and <i>tolB<i>. In both strain, ClyA-flg22 and ClyA-elf18 can trigger stronger immune responses.

Omp8 OMVs on col-0.jpeg

Figure 2.

TolB OMVs on col-0.jpeg

Figure 3.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 1105
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 1105
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 1105
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 1105
    Illegal NgoMIV site found at 1099
    Illegal AgeI site found at 1063
  • 1000
    COMPATIBLE WITH RFC[1000]