Difference between revisions of "Part:BBa K4800003"
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<p style="font-size: 180%; font-weight: bold;">Build: | <p style="font-size: 180%; font-weight: bold;">Build: | ||
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<p><img src="https://static.igem.wiki/teams/4800/wiki/parts/ptrc-ycia-sfp-mmcar-yahk-png.png" width="80%" height="80%"></p> | <p><img src="https://static.igem.wiki/teams/4800/wiki/parts/ptrc-ycia-sfp-mmcar-yahk-png.png" width="80%" height="80%"></p> | ||
<div align="center"> | <div align="center"> | ||
| − | <strong> | + | <strong>fig1.ptrc-ycia-sfp-mmcar-yahk</strong> |
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<p>1.Experimental methods | <p>1.Experimental methods | ||
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1.2Construction of PRSFDuet-YciA-sfp-MmCAR-YahK | 1.2Construction of PRSFDuet-YciA-sfp-MmCAR-YahK | ||
The PRSFDuet plasmid was extracted from laboratory conserved strains. The fragment Ptrc-YciA-sfp-MmCAR-sfp-Yahk was amplified by PCR using PCWJ-YciA-sfp-MmCAR-YahK as a template, and Ptrc-YciA-MmCAR-sfp-Yahk was ligated by In-fusion cloning to the vector PRSFDuet that was linearized using PCR for homologous recombination ligation. | The PRSFDuet plasmid was extracted from laboratory conserved strains. The fragment Ptrc-YciA-sfp-MmCAR-sfp-Yahk was amplified by PCR using PCWJ-YciA-sfp-MmCAR-YahK as a template, and Ptrc-YciA-MmCAR-sfp-Yahk was ligated by In-fusion cloning to the vector PRSFDuet that was linearized using PCR for homologous recombination ligation. | ||
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<p style="font-size: 180%; font-weight: bold;">Test: | <p style="font-size: 180%; font-weight: bold;">Test: | ||
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<p><img src="https://static.igem.wiki/teams/4800/wiki/parts/catalytic-generation-of-1-5-pdo-by-bl21-de3-containing-different-plasmids-png.png" width="80%" height="80%"></p> | <p><img src="https://static.igem.wiki/teams/4800/wiki/parts/catalytic-generation-of-1-5-pdo-by-bl21-de3-containing-different-plasmids-png.png" width="80%" height="80%"></p> | ||
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| − | <strong> | + | <strong>fig2.Catalytic generation of 1,5-PDO by BL21(DE3) containing different plasmids</strong> |
</div> | </div> | ||
<p>By HPLC data, it was seen that BL21 imported into PRSFDuet-YciA-sfp-MmCAR-Yahk catalyzed the production of more 1,5-PDO.</p > | <p>By HPLC data, it was seen that BL21 imported into PRSFDuet-YciA-sfp-MmCAR-Yahk catalyzed the production of more 1,5-PDO.</p > | ||
| − | <img src="https://static.igem.wiki/teams/4800/wiki/parts/fermentation-of-ka30-containing-different-plasmids-to-produce-1-5-pdo-png.png" width="80%" height="80%"></p> | + | <p><img src="https://static.igem.wiki/teams/4800/wiki/parts/fermentation-of-ka30-containing-different-plasmids-to-produce-1-5-pdo-png.png" width="80%" height="80%"></p> |
<div align="center"> | <div align="center"> | ||
| − | <strong> | + | <strong>fig3.Fermentation of KA30 containing different plasmids to produce 1,5-PDO</strong> |
</div> | </div> | ||
| − | <p>By comparing the fermentation production of 1,5-PDO content with the previous KA30 with PCWJ+PTRC99a, we can clearly see that KA30 with PRSFDuet+PTRC99a produced more 1,5-PDO, therefore, we decided to use both vectors, PRSFDuet and PTRC99a, in all our future experiments. | + | <p>By comparing the fermentation production of 1,5-PDO content with the previous KA30 with PCWJ+PTRC99a, we can clearly see that KA30 with PRSFDuet+PTRC99a produced more 1,5-PDO, therefore, we decided to use both vectors, PRSFDuet and PTRC99a, in all our future experiments.</p > |
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| − | </p > | + | |
</body> | </body> | ||
Revision as of 16:18, 6 October 2023
Ptrc-sfp-MmCAR-YahK
(1)Ptrc is a promoter.(2)YahK is a acyl-CoA thioester hydrolase from Haemophilus influenzae.(3) Sfp is a phosphopantetheinyl transferase from Bacilus subtilis.(4) MmCAR is a Carboxylic acid reductase from Mycobacterium marinum.(5) YahK is a NADPH-dependent aldehyde reductase.
Sequence and Features
Assembly Compatibility:
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 1332
Illegal PstI site found at 1767
Illegal PstI site found at 1794
Illegal PstI site found at 3063
Illegal PstI site found at 3090
Illegal PstI site found at 3483
Illegal PstI site found at 3979
Illegal PstI site found at 4623 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 1332
Illegal NheI site found at 4891
Illegal PstI site found at 1767
Illegal PstI site found at 1794
Illegal PstI site found at 3063
Illegal PstI site found at 3090
Illegal PstI site found at 3483
Illegal PstI site found at 3979
Illegal PstI site found at 4623 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 1332
Illegal BglII site found at 2321
Illegal BglII site found at 2947
Illegal BglII site found at 3223
Illegal BglII site found at 3914
Illegal BamHI site found at 1304
Illegal XhoI site found at 4863 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 1332
Illegal PstI site found at 1767
Illegal PstI site found at 1794
Illegal PstI site found at 3063
Illegal PstI site found at 3090
Illegal PstI site found at 3483
Illegal PstI site found at 3979
Illegal PstI site found at 4623 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 1332
Illegal PstI site found at 1767
Illegal PstI site found at 1794
Illegal PstI site found at 3063
Illegal PstI site found at 3090
Illegal PstI site found at 3483
Illegal PstI site found at 3979
Illegal PstI site found at 4623
Illegal NgoMIV site found at 3591
Illegal NgoMIV site found at 3653
Illegal NgoMIV site found at 3870
Illegal NgoMIV site found at 4317
Illegal AgeI site found at 1497
Illegal AgeI site found at 2508
Illegal AgeI site found at 2805
Illegal AgeI site found at 3690
Illegal AgeI site found at 3699
Illegal AgeI site found at 4374 - 1000COMPATIBLE WITH RFC[1000]