Difference between revisions of "Part:BBa K4724083:Design"

 
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===Design Notes===
 
===Design Notes===
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The DsbA gene sequence was inserted into the vector pET-22b (+) - LSPET-12 and a recombinant plasmid was constructed using the Gibson assembly method.
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In the initial design, since the signaling peptide is a small protein with a structural length of 20-30 amino acids, its coding sequence is approximately 60-90 bp. Therefore, we aimed to design primers to amplify the coding sequence of the signaling peptide and attach it to the coding sequence of the LSPET enzyme through PCR. We used previously designed reverse PCR primers between the RBS and ISPET enzyme sequences and added the signaling peptide sequence to the primers. By employing homologous recombination, we constructed the plasmid and ensured that there is a 20 bp homologous region of the signaling peptide sequence on both the forward and reverse primers.
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===Source===
 
===Source===
  
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An enzyme identified in the laboratory of our research group that can identify PET.
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Laboratory design in our research group.
  
 
===References===
 
===References===

Latest revision as of 19:41, 10 October 2023


DsbA-LSPETase


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 1186
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 322
    Illegal NgoMIV site found at 442
    Illegal AgeI site found at 529
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

In the initial design, since the signaling peptide is a small protein with a structural length of 20-30 amino acids, its coding sequence is approximately 60-90 bp. Therefore, we aimed to design primers to amplify the coding sequence of the signaling peptide and attach it to the coding sequence of the LSPET enzyme through PCR. We used previously designed reverse PCR primers between the RBS and ISPET enzyme sequences and added the signaling peptide sequence to the primers. By employing homologous recombination, we constructed the plasmid and ensured that there is a 20 bp homologous region of the signaling peptide sequence on both the forward and reverse primers.

Source

Laboratory design in our research group.

References