Plasmid

Part:BBa_K4103000

Designed by: Ahmed Basil Hussain   Group: iGEM21_Open_Science_Global   (2021-10-21)


pHT43_PfuSso7d

This plasmid is a shuttle vector from E. coli to B. subtilis that enables genomic integration into B. subtilis and the expression, purification, and proper secretion (with the aid of our library secretion plasmids - parts BBa_K4103046-BBa_K4103051).

As a derivative of one of the first discovered natural B. subtilis vector plasmids, pBS72

This plasmid backbone (without the transcriptional unit (from the promoter to the terminator) can also be used in the production of Pichia constructs, using the relevant Pichia parts from the Open Yeast Collection, as shown below:


OYC-bridge-AGAC-CGAA BBF10K_000476 -> this is in case there is no 3' homology arm OYC-bridge-AGAC-GCAA BBF10K_000477 -> this is in case there is no 3' homology nor packaging (conjugation/transfer) OYC-bridge-CGAA-GCAA BBF10K_000478 -> this is in case there's no packaging OYC-eforRed-dropout-cassette BBF10K_000482 repA -> see design page for more information. oriT ecoli facilitator: https://cgsc.biology.yale.edu/Strain.php?ID=122765 E. coli conjugation helper strain with the RK2 plasmid. Mix it with Bsub and other E. coli containing shuttle vectors with at least one of our oriT's, and the C600 cells will conjugate our vector from E. coli to B. subtilis for us. (plus pichia parts from OYC - just link to freegenes)</p>

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 3028
    Illegal XbaI site found at 3037
    Illegal SpeI site found at 8672
    Illegal PstI site found at 8681
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 3028
    Illegal NheI site found at 8514
    Illegal SpeI site found at 8672
    Illegal PstI site found at 8681
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 3028
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 3028
    Illegal XbaI site found at 3037
    Illegal SpeI site found at 8672
    Illegal PstI site found at 8681
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 3028
    Illegal XbaI site found at 3037
    Illegal SpeI site found at 8672
    Illegal PstI site found at 8681
    Illegal AgeI site found at 7630
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 2076


[edit]
Categories
Parameters
None