Difference between revisions of "Part:BBa K3771003"
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<br><b style="font-size:1.3rem">Description
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<br><i>PlacI-csad</i> is a composite part consisting of the lacI promoter and the csad sequences. This part was used in in vivo testing of taurine production. The sequence for csad enzyme and lacI promoter were ligated and transformed into E. coli to calculate taurine production using high-performance liquid chromatography (HPLC).<br>
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<br><b style="font-size:1.3rem">Biology
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<br><i>lacI</i> promoter constitutively facilitates the expression of CSAD enzyme.<br>
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<html><div style="width=100%; display:flex; align-items: center; justify-content: center;">
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<img src="https://2021.igem.org/wiki/images/c/c9/T--NCKU_Tainan--taurine_pathway_1.png" style="width:35%;"></html>
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===Usage and Biology===
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<br><b style="font-size:1.3rem">Usage
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<br>CDO1 was used in in vivo testing of taurine production. The sequence for CDO1 enzyme and trc promoter were ligated and transformed into E. coli to calculate taurine production using high-performance liquid chromatography (HPLC).
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<html><div style="width=100%; display:flex; align-items: center; justify-content: center;">
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<img src="https://2021.igem.org/wiki/images/6/6b/T--NCKU_Tainan--invivo1.png" style="width:35%;"></html>
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<br><b style="font-size:1.3rem">Characterization
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<br>The CDO1 fragment was synthesized by IDT and amplified by PCR. Agarose gel electrophoresis result is shown in Fig. 2.<br>
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<img src="https://2021.igem.org/wiki/images/8/86/T--NCKU_Tainan--CSAD-PCR.png" style="width:35%;"></html>
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<p>Fig. 2 Confirmation of csad fragment by PCR. M: Marker; Lane 1: csad (1368 bp)</p>
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<img src="https://2021.igem.org/wiki/images/4/48/T--NCKU_Tainan--CSAD1-Vactor-digestion.png" style="width:35%;"></html>
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<p>Fig. 2 Confirmation of pSAA fragment by digestion. M: Marker; Lane 1: pSAA (2531 bp)</p>
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<img src="https://2021.igem.org/wiki/images/4/4e/T--NCKU_Tainan--CDO1-PAGE.png" style="width:35%;"></html>
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<p>Fig. 3 Confirmation of protein expression of CDO1.M: Marker; Lane1: CDO1 in DH5α without induction; Lane2: CDO1 in DH5α with induction (~22 kDa)</p>
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<html><div style="width=100%; display:flex; align-items: center; justify-content: center;">
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<img src="https://2021.igem.org/wiki/images/a/aa/T--NCKU_Tainan--3.png" style="width:35%;"></html>
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<p>Fig. 4 Taurine production of Ptrc-cdo1 +PlacI-csad +Ptrc-cs in different growth mediums.</p>
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<br><b style="font-size:1.3rem">References
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<br>Joo Y-C, Ko YJ, You SK, et al. Creating a New Pathway in Corynebacterium glutamicum for the Production of Taurine as a Food Additive. Journal of Agricultural and Food Chemistry. 2018;66(51):13454-13463. doi:10.1021/acs.jafc.8b05093<br>
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<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Revision as of 10:58, 18 October 2021


CSAD


Description
PlacI-csad is a composite part consisting of the lacI promoter and the csad sequences. This part was used in in vivo testing of taurine production. The sequence for csad enzyme and lacI promoter were ligated and transformed into E. coli to calculate taurine production using high-performance liquid chromatography (HPLC).


Biology
lacI promoter constitutively facilitates the expression of CSAD enzyme.


Usage
CDO1 was used in in vivo testing of taurine production. The sequence for CDO1 enzyme and trc promoter were ligated and transformed into E. coli to calculate taurine production using high-performance liquid chromatography (HPLC).



Characterization
The CDO1 fragment was synthesized by IDT and amplified by PCR. Agarose gel electrophoresis result is shown in Fig. 2.

Fig. 2 Confirmation of csad fragment by PCR. M: Marker; Lane 1: csad (1368 bp)

Fig. 2 Confirmation of pSAA fragment by digestion. M: Marker; Lane 1: pSAA (2531 bp)

Fig. 3 Confirmation of protein expression of CDO1.M: Marker; Lane1: CDO1 in DH5α without induction; Lane2: CDO1 in DH5α with induction (~22 kDa)

Fig. 4 Taurine production of Ptrc-cdo1 +PlacI-csad +Ptrc-cs in different growth mediums.




References
Joo Y-C, Ko YJ, You SK, et al. Creating a New Pathway in Corynebacterium glutamicum for the Production of Taurine as a Food Additive. Journal of Agricultural and Food Chemistry. 2018;66(51):13454-13463. doi:10.1021/acs.jafc.8b05093


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
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 250
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 35