Difference between revisions of "Part:BBa K4632002:Design"
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+ | <h2>Design Notes</h2> | ||
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+ | '''3. What we have done? (SCAU-China 2023)''' | ||
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+ | <p>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. </p> | ||
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+ | <p>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 )</p> | ||
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+ | <p>'''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 )</p> | ||
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+ | ''https://static.igem.wiki/teams/4632/wiki/wiki/registry-part/part-1-7-1.png'' | ||
+ | <p><strong>Figure 1.</strong>Diagram of Cry3A-like Toxin circuit design</p> | ||
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+ | ''https://static.igem.wiki/teams/4632/wiki/wiki/registry-part/part-1-20-1.png'' | ||
+ | <p><strong>Figure 2.</strong>pET30a-OmpA-Cry3A-like toxin with a 6×His tag</p> | ||
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+ | <h2>Source</h2> | ||
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+ | 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. | ||
+ | <h2>References</h2> | ||
<html> | <html> | ||
+ | [1] Lee A. Bulla, Jr.Mehmet Candas Formicidae (ant) control using <i>Bacillus thuringiensis</i> toxin US 6,551,800B1[P]. 2003-04-22. | ||
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Latest revision as of 09:30, 11 October 2023
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.