Part:BBa_K5246036

Rosetta(DE3)pLysS ΔWecA

Usage and Biology

WecA is Undecaprenyl-phosphate α-N-acetylglucosaminyl transferase initiating the biosynthesis of enterobacterial common antigen (ECA) and O-antigen PS by catalyzing the transfer of N-acetylglucosamine (GlcNAc)-1-phosphate onto undecaprenyl phosphate to form Und-P-P-GlcNAc. WecA is the first glycosyltransferase, and deleting the wecA gene impairs the ECA synthesis pathway in E.coli, therefore lipopolysaccharides containing it are not produced. We wanted to disable the ECA pathway to reduce the metabolic burden on the cell. We aimed to optimize polysaccharide production by creating a more efficient strain. Rosetta(DE3)pLysS strain is designed to overcome the limitations of E. coli's codon usage by providing additional tRNAs for certain codons found in eukaryotic proteins. This allows for better expression of these proteins. In Rosetta(DE3)pLysS, the rare tRNA genes and the T7 lysozyme gene are on the same plasmid.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal XbaI site found at 128
  Illegal XbaI site found at 1350
  Illegal PstI site found at 106
  Illegal PstI site found at 693
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal PstI site found at 106
  Illegal PstI site found at 693
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 480
  Illegal BamHI site found at 94
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal XbaI site found at 128
  Illegal XbaI site found at 1350
  Illegal PstI site found at 106
  Illegal PstI site found at 693
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal XbaI site found at 128
  Illegal XbaI site found at 1350
  Illegal PstI site found at 106
  Illegal PstI site found at 693
  Illegal NgoMIV site found at 1144
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI.rc site found at 993
  Illegal SapI.rc site found at 1203
  

Functional Parameters

genotype

F- ompT hsdSB(rB- mB-) gal dcm (DE3) pLysSRARE (CamR)(KanR) ΔwecA