Difference between revisions of "Part:BBa K3838345"
 
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<partinfo>BBa_K3838345 short</partinfo>
 
<partinfo>BBa_K3838345 short</partinfo>
  
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The overproduction of Superoxide dismutase (SOD) and reactive oxygen species (ROS) is considered to be one of the pathogenesis of IBD. The use of SOD can significantly reduce the peroxidation reaction in the inflamed colon. Studies have shown that the decreased expression of Cu/Zn-SOD has been observed in IBD patients. SOD can not be directly treated, so carrier transport is needed. Therefore, we selected for heteroexpression of superoxide dismutase (SOD) gene from Lactobacillus casei Lc18.We used the P170 promoter to express our protein and constructed a plasmid containing RcfB gene to initiate P170 activity.
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The overproduction of Superoxide dismutase (SOD) and reactive oxygen species (ROS) is considered to be one of the pathogenesis of IBD. The use of SOD can significantly reduce the peroxidation reaction in the inflamed colon. Studies have shown that the decreased expression of Cu/Zn-SOD has been observed in IBD patients. SOD can not be directly treated, so carrier transport is needed. Therefore, we selected for heteroexpression of superoxide dismutase (SOD) gene from Lactobacillus casei Lc18.We used the Prcf promoter to express our protein and constructed a plasmid containing RcfB gene to initiate Prcf activity.
  
 
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1.The DNA level
 
1.The DNA level
  
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We selected the DYHSG-RR double plasmid system for verification, transformed PUCYH and PET-RR into DH5α, and extracted the plasmid from the plasmid-transformed DYHSG-RR, as shown in Figure 29. The sample band is between 5000-7500bp, which is in line with the purpose. The band size is around 6700bp. The size of the PCR band is about 1000bp, and the restriction band is about 2300bp, both of which are in line with the current band size.
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We selected the DYHSG-RR double plasmid system for verification, transformed PUCYH and PET-RR into DH5α, and extracted the plasmid from the plasmid-transformed DYHSG-RR, as shown in Figure 1. The sample band is between 5000-7500bp, which is in line with the purpose. The band size is around 6700bp. The size of the PCR band is about 1000bp, and the restriction band is about 2300bp, both of which are in line with the current band size.
 
[[File:T--SZU-China--resultfigure28.png|500px|center]]
 
[[File:T--SZU-China--resultfigure28.png|500px|center]]
 
<center>Fig.1 The transformed DYHSG-RR plasmid, PCR, restriction gel electrophoresis diagram.</center>
 
<center>Fig.1 The transformed DYHSG-RR plasmid, PCR, restriction gel electrophoresis diagram.</center>
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2.Functional representation
 
2.Functional representation
  
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[[File:T--SZU-China--ENS8.png|500px|center]]
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[[File:T--SZU-China-Engineering Success4.png|500px|center]]
 
<center>Fig.2 SOD enzyme activity of constitutive promoter and acid-induced promoter expression system under different PH conditions.</center>
 
<center>Fig.2 SOD enzyme activity of constitutive promoter and acid-induced promoter expression system under different PH conditions.</center>

Latest revision as of 20:09, 21 October 2021


YH-SOD

The overproduction of Superoxide dismutase (SOD) and reactive oxygen species (ROS) is considered to be one of the pathogenesis of IBD. The use of SOD can significantly reduce the peroxidation reaction in the inflamed colon. Studies have shown that the decreased expression of Cu/Zn-SOD has been observed in IBD patients. SOD can not be directly treated, so carrier transport is needed. Therefore, we selected for heteroexpression of superoxide dismutase (SOD) gene from Lactobacillus casei Lc18.We used the Prcf promoter to express our protein and constructed a plasmid containing RcfB gene to initiate Prcf activity.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal prefix found in sequence at 9
    Illegal EcoRI site found at 534
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 9
    Illegal EcoRI site found at 534
    Illegal NheI site found at 147
    Illegal NheI site found at 170
    Illegal NotI site found at 15
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 9
    Illegal EcoRI site found at 534
    Illegal BglII site found at 577
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal prefix found in sequence at 9
    Illegal EcoRI site found at 534
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal prefix found in sequence at 9
    Illegal EcoRI site found at 534
    Illegal XbaI site found at 24
    Illegal NgoMIV site found at 738
  • 1000
    COMPATIBLE WITH RFC[1000]


Data:SZU-China 2021 TEAM

We placed the engineered bacteria in media with different pH and cultured them to the same concentration, and then tested their intracellular superoxide dismutase activity.We can verify the difference in the activity of the SOD enzyme expressed between the constitutive and acid-inducible expression systems through this experiment and verify the difference in the expression efficiency of the double-plasmid acid-inducible expression system itself under different pH environments.

1.The DNA level

We selected the DYHSG-RR double plasmid system for verification, transformed PUCYH and PET-RR into DH5α, and extracted the plasmid from the plasmid-transformed DYHSG-RR, as shown in Figure 1. The sample band is between 5000-7500bp, which is in line with the purpose. The band size is around 6700bp. The size of the PCR band is about 1000bp, and the restriction band is about 2300bp, both of which are in line with the current band size.

T--SZU-China--resultfigure28.png
Fig.1 The transformed DYHSG-RR plasmid, PCR, restriction gel electrophoresis diagram.


2.Functional representation


T--SZU-China-Engineering Success4.png
Fig.2 SOD enzyme activity of constitutive promoter and acid-induced promoter expression system under different PH conditions.