Part:BBa_K5366049

pACYCDuet-1-tagR-P-RS-sfGFP1

Sensor plasmids for deterrent protein and green fluorescent constructs

The recombinant plasmid pETDuet-1-tagR-P-RS-sfGFP1 was used as a template to obtain the target gene fragment (1936 bp) using Primer 4 and Primer 5. The resulting fragment was then ligated to the pACYCDuet fragment and introduced into E.coli Nissle 1917 (EcN). Colony PCR (2103 bp) was conducted on the transformed colonies using TB validation-for-0112 and T7T as primers. Following sequencing verification, the recombinant plasmid pACYCDuet-1-tagR-P-RS-sfGFP1 was successfully obtained. However, this biosensor did not perform as expected and exhibited stunted functionality.

Fig. 1 Construction maps of plasmids of pacycduet-1-tagr-p-rs-sfgfp1-cmr

Fig. 2 Construction maps of plasmids of pacycduet-1-tagr-p-rs-sfgfp1-ampr

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 1850
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 1850
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 1850
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 1850
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 1850
  Illegal AgeI site found at 1211
  Illegal AgeI site found at 1334
  
• 1000
  COMPATIBLE WITH RFC[1000]