Difference between revisions of "Part:BBa K3606812"
 
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<h2>Design:</h2>
 
<h2>Design:</h2>
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Since we encountered difficulties in the expression of McbABCDEFG at the beginning, we decided to check whether the McbA/B/C/D/E/F/G genes can be expressed and translated separately. We successfully cloned McbG into the pGEX plasmid and transcribed it into BL21. After 5 hours of induction of McbG expression with IPTG, the whole protein SDS-PAGE was used to detect whether the McbG product was produced.
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Since we encountered difficulties in the expression of McbABCDEFG at the beginning, we decided to check whether the McbA/B/C/D/E/F/G genes can be expressed and translated separately. We successfully cloned McbG into the pGEX plasmid and transcribed it into BL21. After 5 hours of induction of McbG expression with IPTG, the whole protein SDS-PAGE was used to detect whether the McbG product was produced, yet the result of colony PCR turns out as false positive.
  
 
<h2>Results:</h2>
 
<h2>Results:</h2>
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https://2020.igem.org/wiki/images/2/22/T--Fudan--img_McbA-E-G-BCD.jpeg
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Figure 1. SDS-PAGE result. lane 1: mcbA lane 2: mcbA+IPTG lane 3: mcbE lane 4: mcbE+IPTG  lane 5: mcbG  lane 6: mcbG+IPTG  lane 7: mcbBCD  lane 8: mcbBCD+IPTG lane 9: pGEX+ IPTG
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We've constructed the plasmid for mcbG expression, yet the results of colony PCR is false positive thus further experiment is needed for the fundamental validation of the antimicrobial peptide(mccb17) expression.
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https://2020.igem.org/wiki/images/f/f3/T--Fudan--Poster_Mcb-gels.jpg
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Figure2. gel map for mcbA, mcbC, mcbE and mcbF
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<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here
 
===Usage and Biology===
 
===Usage and Biology===

Latest revision as of 02:40, 5 December 2020


mcbG

Usage and Biology:

McbG is responsible for immunity against MccB17.

Design:

Since we encountered difficulties in the expression of McbABCDEFG at the beginning, we decided to check whether the McbA/B/C/D/E/F/G genes can be expressed and translated separately. We successfully cloned McbG into the pGEX plasmid and transcribed it into BL21. After 5 hours of induction of McbG expression with IPTG, the whole protein SDS-PAGE was used to detect whether the McbG product was produced, yet the result of colony PCR turns out as false positive.

Results:

We've constructed the plasmid for mcbG expression, yet the results of colony PCR is false positive thus further experiment is needed for the fundamental validation of the antimicrobial peptide(mccb17) expression. T--Fudan--Poster_Mcb-gels.jpg Figure2. gel map for mcbA, mcbC, mcbE and mcbF

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
    COMPATIBLE WITH RFC[1000]