Difference between revisions of "Part:BBa K3771078"

Revision as of 02:53, 22 October 2021

Ptrc-CS

Description

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

Biology

trc promoter constitutively facilitates the expression of CS enzyme.

Fig. 1. L-cysteine sulfonic acid synthase catalyzes the reaction that turns O-phospho-L-serine into L-cysteine sulfonic acid. CDO1, L-cysteine dioxygenase; CSAD, L-cysteine sulfinic acid decarboxylase.

Usage

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

Fig. 2. Taurine production of P_trc-cdo1+P_lacI-csad +P_trc-cs in different growth mediums.

Characterization

Fig. 3. Confirmation of cs fragment by PCR. M: Marker; Lane 1: cs (1272 bp)

Fig. 4. Confirmation of pSUI fragment by digestion. M: Marker; Lane 1: pSUI (3664 bp)

Fig. 5. Transformation/CS in DH5α

Fig. 6. Confirmation of cs fragment by colony PCR. M: Marker; Lane 1~6: Different colonies of pSUI-P_trc-cs (1272 bp)

Fig. 7. Confirmation of pSUI-P_trc-CS by digestion. M: Marker; Lane 1: Colony of pSUI-P_trc-cs

Fig. 8. Confirmation of the protein expression of CS M: Marker; Lane 1: CS in DH5α with induction (~46 kDa)

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
COMPATIBLE WITH RFC[25]
1000
INCOMPATIBLE WITH RFC[1000]
Illegal SapI.rc site found at 580

@@ Line 19: / Line 19: @@
   <img src="https://2021.igem.org/wiki/images/c/c9/T--NCKU_Tainan--taurine_pathway_1.png" style="width:35%;">
   </div>
-   <p>Fig. 1 L-cysteine sulfonic acid synthase catalyzes the reaction that turns O-phospho-L-serine into L-cysteine sulfonic acid. CDO1, L-cysteine dioxygenase; CSAD, L-cysteine sulfinic acid decarboxylase.
+   <p>Fig. 1. L-cysteine sulfonic acid synthase catalyzes the reaction that turns O-phospho-L-serine into L-cysteine sulfonic acid. CDO1, L-cysteine dioxygenase; CSAD, L-cysteine sulfinic acid decarboxylase.
    </p>
@@ Line 31: / Line 31: @@
   <img src="https://2021.igem.org/wiki/images/a/aa/T--NCKU_Tainan--3.png" style="width:35%;">
   </div>
-   <p>Fig. 2 Taurine production of <i>P<sub>trc</sub>-cdo1</i>+<i>P<sub>lacI</sub>-csad</i> +<i>P<sub>trc</sub>-cs</i> in different growth mediums.</p>
+   <p>Fig. 2. Taurine production of <i>P<sub>trc</sub>-cdo1</i>+<i>P<sub>lacI</sub>-csad</i> +<i>P<sub>trc</sub>-cs</i> in different growth mediums.</p>
 <br><b style="font-size:1.3rem">Characterization</b>
@@ Line 39: / Line 39: @@
 <img src="https://2021.igem.org/wiki/images/3/39/T--NCKU_Tainan--CS-PCR.png" style="width:35%;">
 </div>
-<p align="center">Fig. 2 Confirmation of <i>cs</i> fragment by PCR. M: Marker; Lane 1: <i>cs</i> (1272 bp)</p>
+<p align="center">Fig. 3. Confirmation of <i>cs</i> fragment by PCR. M: Marker; Lane 1: <i>cs</i> (1272 bp)</p>
 <div style="width=100%; display:flex; align-items: center; justify-content: center;">
 <img src="https://2021.igem.org/wiki/images/e/e3/T--NCKU_Tainan--CS-Vactor-digestion.png" style="width:35%;">
 </div>
-<p align="center">Fig. 3 Confirmation of pSUI fragment by digestion. M: Marker; Lane 1: pSUI (3664 bp)</p>
+<p align="center">Fig. 4. Confirmation of pSUI fragment by digestion. M: Marker; Lane 1: pSUI (3664 bp)</p>
 <div style="width=100%; display:flex; align-items: center; justify-content: center;">
 <img src="https://2021.igem.org/wiki/images/3/35/T--NCKU_Tainan--CS-plate%28DH5a%29.png" style="width:35%;">
 </div>
-<p align="center">Fig. 4 Transformation/CS in DH5α</p>
+<p align="center">Fig. 5. Transformation/CS in DH5α</p>
 <div style="width=100%; display:flex; align-items: center; justify-content: center;">
 <img src="https://2021.igem.org/wiki/images/9/9b/T--NCKU_Tainan--CS-colony-PCR.png" style="width:35%;">
 </div>
-<p align="center">Fig. 5 Confirmation of <i>cs</i> fragment by colony PCR.
+<p align="center">Fig. 6. Confirmation of <i>cs</i> fragment by colony PCR.
 M: Marker; Lane 1~6: Different colonies of <i>pSUI-P<sub>trc</sub>-cs</i> (1272 bp)
 </p>
@@ Line 61: / Line 61: @@
 <img src="https://2021.igem.org/wiki/images/1/1d/T--NCKU_Tainan--CS-digestion.png" style="width:35%;">
 </div>
-<p align="center">Fig. 6 Confirmation of <i>pSUI-P<sub>trc</sub>-CS</i> by digestion.
+<p align="center">Fig. 7. Confirmation of <i>pSUI-P<sub>trc</sub>-CS</i> by digestion.
 M: Marker; Lane 1: Colony of <i>pSUI-P<sub>trc</sub>-cs</i>
 </p>
@@ Line 68: / Line 68: @@
 <img src="https://2021.igem.org/wiki/images/6/64/T--NCKU_Tainan--CS-PAGE%28DH5a%29.png" style="width:35%;">
 </div>
-<p align="center">Fig. 7 Confirmation of the protein expression of CS
+<p align="center">Fig. 8. Confirmation of the protein expression of CS
 M: Marker; Lane 1: CS in DH5α with induction (~46 kDa)
 </p>