Part:BBa_K4999000

pBAD driven T4 lysis cassette

We constructed an arabinose inducible promoter driven lysis cassette. We cloned the T4 lysis cassette which includes i) T4 endolysin gene ii) T4 holin gene in a polycistronic CDS driven by a pBAD promoter. The plasmid also contains two supporting transcriptional units expressing i) araC gene involved in arabinose induction ii) an anti-holin gene that restricts lysis due to leaky expression. We used Gibson assembly to make the final construct. We screened for correct clones by i) confirm lysis upon arabinose induction ii) whole plasmid sequencing the construct. This plasmid contains all the genetic elements necessary for performing controlled lysis. More information about the characterization can be found on our contribution page: https://2023.igem.wiki/harvard/contribution

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 1205
  Illegal NheI site found at 2560
  Illegal NheI site found at 2583
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 1144
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 979
  Illegal AgeI site found at 2165
  Illegal AgeI site found at 2235
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI site found at 961
  Illegal SapI site found at 2816