Part:BBa_K2938006

Cry4Ba + HA tag

Cry4Ba Bti toxin with HA-tag attached in the N- terminal in order to be able to detect the translated protein in western blot (using antibodies against the tag)

Description

Pesticidal crystal protein Cry4Ba is a 128 kDa protein which can be isolated from Bacillus thuringiensis subsp. israelensis (Bti). The pBtoxis is a toxin-encoding plasmid that its origin is the Bti and it encodes to four Cry toxins (Cry4Aa, Cry4Ba, Cry10Aa and Cry11Aa) and three Cyt toxins (Cyt1Aa, Cyt2Ba and Cyt1Ca)[1], which are specifically toxic against Aedes, Anopheles and Culex mosquitoes which are vectors to many diseases caused by parasites and viruses, such as Malaria, Zika and Dengue. Each of the toxins hold a low level of larvicidal effect but their combination has a synergetic effect which is stronger than the linear combination we expect. The Cyt1Aa toxin effect in the synergetic effect is mainly the prevention of resistance in the mosquito to the Cry toxins. The bigger Cry toxins, Cry4Aa and Cry4Ba have a conserved C-terminal end which helps the toxin fold correctly and form crystal structures.

Cry4Ba is toxic specifically against Aedes Aegypti mosquitoes’ larvae (compared to Cry4Aa) [2]

Amplification

PCR amplification of Cry4Ba by PCR.

Usage and Biology

The Cry4Ba is a toxin with larvicidal effect on mosquitos’ larvae, and HA-tag is used to detect proteins in western blot using antibodies specific to the tag. Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal XbaI site found at 1708
  Illegal SpeI site found at 2397
  Illegal PstI site found at 212
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 96
  Illegal SpeI site found at 2397
  Illegal PstI site found at 212
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal XbaI site found at 1708
  Illegal SpeI site found at 2397
  Illegal PstI site found at 212
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal XbaI site found at 1708
  Illegal SpeI site found at 2397
  Illegal PstI site found at 212
  Illegal NgoMIV site found at 2520
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 502