Part:BBa_K3385068

T1G3R_Spilot_glaA

About the part: Insert of glaA expression cassette for A. niger. It is made to be integrated in albA, which will lead to white/yellow conidia upon correct integration in the genome. The insert is inserted in pSB1C3 via USER cloning to create the integration plasmid for A. niger. The signal peptide was changed to SigPilot using USER cloning and primer extension. The integration platform is made from part BBa_K3046030 pGIA2P1 designed by the DTU iGEM 2019 team.

Functionality: This signal peptide was investigated in Deeploc [1] to predict the localization of the protein which it is inserted upstream of.

Localization predicted by Deeploc.

Result: A tissue PCR was performed to show, that the synthetic signal peptide can be successfully integrated in the A. niger genome.

Tissue PCR showing the integration of the synthetic signal peptides SigPower, SigPeanut and SigPilot into the A. niger genome.

Tissue PCR showing the integration of the synthetic signal peptides SigPilot, SigPineapple and SigPuppy into the A. niger genome.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 1858
  Illegal EcoRI site found at 5765
  Illegal EcoRI site found at 7193
  Illegal XbaI site found at 1623
  Illegal XbaI site found at 5796
  Illegal SpeI site found at 1097
  Illegal PstI site found at 893
  Illegal PstI site found at 4027
  Illegal PstI site found at 6603
  Illegal PstI site found at 8337
  Illegal PstI site found at 8938
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 1858
  Illegal EcoRI site found at 5765
  Illegal EcoRI site found at 7193
  Illegal NheI site found at 4904
  Illegal SpeI site found at 1097
  Illegal PstI site found at 893
  Illegal PstI site found at 4027
  Illegal PstI site found at 6603
  Illegal PstI site found at 8337
  Illegal PstI site found at 8938
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 1858
  Illegal EcoRI site found at 5765
  Illegal EcoRI site found at 7193
  Illegal BglII site found at 3882
  Illegal BglII site found at 5404
  Illegal BglII site found at 8435
  Illegal BamHI site found at 1729
  Illegal BamHI site found at 2157
  Illegal BamHI site found at 4862
  Illegal BamHI site found at 5664
  Illegal BamHI site found at 5790
  Illegal BamHI site found at 6341
  Illegal XhoI site found at 548
  Illegal XhoI site found at 735
  Illegal XhoI site found at 963
  Illegal XhoI site found at 6201
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 1858
  Illegal EcoRI site found at 5765
  Illegal EcoRI site found at 7193
  Illegal XbaI site found at 1623
  Illegal XbaI site found at 5796
  Illegal SpeI site found at 1097
  Illegal PstI site found at 893
  Illegal PstI site found at 4027
  Illegal PstI site found at 6603
  Illegal PstI site found at 8337
  Illegal PstI site found at 8938
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 1858
  Illegal EcoRI site found at 5765
  Illegal EcoRI site found at 7193
  Illegal XbaI site found at 1623
  Illegal XbaI site found at 5796
  Illegal SpeI site found at 1097
  Illegal PstI site found at 893
  Illegal PstI site found at 4027
  Illegal PstI site found at 6603
  Illegal PstI site found at 8337
  Illegal PstI site found at 8938
  Illegal NgoMIV site found at 2566
  Illegal NgoMIV site found at 5122
  Illegal NgoMIV site found at 8325
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 3410
  Illegal BsaI site found at 3548
  Illegal BsaI site found at 4237
  Illegal BsaI.rc site found at 4730
  Illegal SapI site found at 3097
  Illegal SapI site found at 8826
  Illegal SapI site found at 9135
  

References:
[1] Jose Juan Almagro Armenteros, Casper Kaae Sønderby, Søren Kaae Sønderby, Henrik Nielsen, Ole Winther; DeepLoc: prediction of protein subcellular localization using deep learning, Bioinformatics, btx431