Part:BBa_K3606814

mcbBCD

This part is an antibiotic coding gene cluster with proteins for MccB17 maturation.

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.

Results:

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

Comparing the 5th and 6th lanes, after IPTG induction, there is a clear band, which is the product of McbG expression.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 2062
  Illegal PstI site found at 2095
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal PstI site found at 2062
  Illegal PstI site found at 2095
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 1969
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 2062
  Illegal PstI site found at 2095
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 2062
  Illegal PstI site found at 2095
  Illegal NgoMIV site found at 1724
  Illegal AgeI site found at 1896
  
• 1000
  COMPATIBLE WITH RFC[1000]