Part:BBa_K4632002:Design
Cry3A-like toxin
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 1813
Illegal PstI site found at 295
Illegal PstI site found at 304
Illegal PstI site found at 1456 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 1813
Illegal PstI site found at 295
Illegal PstI site found at 304
Illegal PstI site found at 1456 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 1813
Illegal BglII site found at 533 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 1813
Illegal PstI site found at 295
Illegal PstI site found at 304
Illegal PstI site found at 1456 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 1813
Illegal PstI site found at 295
Illegal PstI site found at 304
Illegal PstI site found at 1456
Illegal AgeI site found at 1495 - 1000COMPATIBLE WITH RFC[1000]
Design Notes
3. What we have done? (SCAU-China 2023)
In our design, the coding gene fragment for the active Cry3A-like toxin will be transformed into E. coli by the pET-30a vector, to confer the ability to produce Cry3A-like toxin.
To enable its secretion, a signal peptide sequence, OmpA, was added to the N-terminal of the Cry3A-like toxin. OmpA is a well-studied signal peptide in E.coli for the secretion of foreign proteins. (Figure 1 )
Furthermore, we fused a 6×His tag to the C-terminus of Cry3A-like toxin to facilitate subsequent protein purification and Western blot-specific characterization experiments.(Figure 2 )
Figure 1.Diagram of Cry3A-like Toxin circuit design
Figure 2.pET30a-OmpA-Cry3A-like toxin with a 6×His tag
Source
We obtained the sequence of the active Cry protoxin of UTD-001 from the original literature. After codon optimization, we entrusted GUANGZHOU IGE BIOTECHNOLOGY LTD to synthesize the gene fragment OmpA-active Cry protoxin of UTD-001.
References
[1] Lee A. Bulla, Jr.Mehmet Candas Formicidae (ant) control using Bacillus thuringiensis toxin US 6,551,800B1[P]. 2003-04-22.