Difference between revisions of "Part:BBa K5002345"

 
 
Line 5: Line 5:
 
The discharge of printing and dyeing wastewater has a very bad impact on the environment, we wanted to reduce the content of harmful substances in printing and dyeing wastewater to the standard of discharge. Therefore, we decided construct a cellulose synthesis operon and a group of plant cellulose discomposing genes. Cellulose synthase (acsAB) is composed of two domains: acsAB is the catalytic domain, and the cyclic di-GMP binding domain.
 
The discharge of printing and dyeing wastewater has a very bad impact on the environment, we wanted to reduce the content of harmful substances in printing and dyeing wastewater to the standard of discharge. Therefore, we decided construct a cellulose synthesis operon and a group of plant cellulose discomposing genes. Cellulose synthase (acsAB) is composed of two domains: acsAB is the catalytic domain, and the cyclic di-GMP binding domain.
  
−
<!-- Add more about the biology of this part here
+
<!-- Add more about the biology of this part here-->
 
===Usage and Biology===
 
===Usage and Biology===
 +
We synthesized acsAB gene encoding cellulose synthase and inserted it into pET23b vector. The constructed plasmid was introduced into Escherichia coli Rosetta for bacterial cellulose production.
 +
<html>
 +
<div style="display:flex; flex-direction: column; align-items: center;">
 +
<img src="https://static.igem.wiki/teams/5002/wiki/part/new-basic-sliver/image-13.png" style="width: 500px;margin: 0 auto" />
 +
<p style="font-size: 98%; line-height: 1.4em;">Figure 1  Design of the acsAB.</p >
 +
</div>
 +
</html>
 +
 +
===Characterization===
 +
We expressed the bacterial cellulose synthase with E.coli Rosetta. We broke them with the ultrasonic wave. Then, we added NaOH into the to dissolves the non-cellulosic components and precipitates the cellulose. After removing the remaining NaOH, we weighed the dried cellulose to determine the cellulose content.
 +
<html>
 +
<div style="display:flex; flex-direction: column; align-items: center;">
 +
<img src="https://static.igem.wiki/teams/5002/wiki/part/new-basic-sliver/image-14.png" style="width: 300px;margin: 0 auto" />
 +
<p style="font-size: 98%; line-height: 1.4em;">Figure 2  Gel electrophoresis of acsAB . </p >
 +
</div>
 +
</html>
 +
 +
<html>
 +
<div style="display:flex; flex-direction: column; align-items: center;">
 +
<img src="https://static.igem.wiki/teams/5002/wiki/part/new-basic-sliver/image-15.png" style="width: 300px;margin: 0 auto" />
 +
<p style="font-size: 98%; line-height: 1.4em;">Figure 3  Cellulose synthesis capacity test of AcsAB.</p >
 +
</div>
 +
</html>
 +
 +
As the picture shown, in bacterial precipitation samples, engineered E. coli expressing AcsAB produced about 1.71g/L bacterial cellulose in LB medium. But in the culture medium samples, there was almost no detectable presence of cellulose. The results indicate that our acsAB gene's ability to express bacterial cellulose synthase is significantly improved.
 +
===Potential application direction===
 +
This experiment demonstrates that the artificially synthesized AcsAB can be expressed normally in Escherichia coli, producing bacterial cellulose. In the future, this technology can be applied to the production of adsorbent materials and the treatment of wastewater in textile printing and dyeing factories, thereby addressing global environmental challenges and having promising prospects for development.
 +
 +
  
 
<!-- -->
 
<!-- -->

Latest revision as of 11:34, 12 October 2023


a cellulose synthesis operon+plant cellulose discomposing genes

The discharge of printing and dyeing wastewater has a very bad impact on the environment, we wanted to reduce the content of harmful substances in printing and dyeing wastewater to the standard of discharge. Therefore, we decided construct a cellulose synthesis operon and a group of plant cellulose discomposing genes. Cellulose synthase (acsAB) is composed of two domains: acsAB is the catalytic domain, and the cyclic di-GMP binding domain.

Usage and Biology

We synthesized acsAB gene encoding cellulose synthase and inserted it into pET23b vector. The constructed plasmid was introduced into Escherichia coli Rosetta for bacterial cellulose production.

Figure 1 Design of the acsAB.

Characterization

We expressed the bacterial cellulose synthase with E.coli Rosetta. We broke them with the ultrasonic wave. Then, we added NaOH into the to dissolves the non-cellulosic components and precipitates the cellulose. After removing the remaining NaOH, we weighed the dried cellulose to determine the cellulose content.

Figure 2 Gel electrophoresis of acsAB .

Figure 3 Cellulose synthesis capacity test of AcsAB.

As the picture shown, in bacterial precipitation samples, engineered E. coli expressing AcsAB produced about 1.71g/L bacterial cellulose in LB medium. But in the culture medium samples, there was almost no detectable presence of cellulose. The results indicate that our acsAB gene's ability to express bacterial cellulose synthase is significantly improved.

Potential application direction

This experiment demonstrates that the artificially synthesized AcsAB can be expressed normally in Escherichia coli, producing bacterial cellulose. In the future, this technology can be applied to the production of adsorbent materials and the treatment of wastewater in textile printing and dyeing factories, thereby addressing global environmental challenges and having promising prospects for development.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 185
    Illegal BglII site found at 1764
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 799
    Illegal NgoMIV site found at 1803
    Illegal NgoMIV site found at 1872
    Illegal NgoMIV site found at 1891
    Illegal AgeI site found at 1750
    Illegal AgeI site found at 2383
    Illegal AgeI site found at 3382
    Illegal AgeI site found at 4231
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 1369
    Illegal BsaI.rc site found at 2617
    Illegal SapI.rc site found at 439