Part:BBa_K4348005
pBSChag
A plasmid backbone we developed that is intended for use with the hag promoter. It integrates into the flgM site of the B. subtilis genome. flgM binds the SigD transcription factor and inactivates it, thus blocking the P hag promoter, one of the strongest promoters discovered.
Introduction
The Bacillus BioBrick Box designed by LMU-Munich iGEM 2012 was used extensively in the McGill 2022 project. The box contains three B. subtilis integrative vectors targeted to the amyE, thrC, and lacA sites, as well as constitutive and inducible promoters with varying strengths. We decided to design a fourth integrative vector and an accompanying promoter as our parts contribution.
Biology
pBSChag is a modification of pBS1C of the BioBrick Box to integrate into the flgM site. Like pBS1C, pBSChag will have an ampicillin selection marker for E. coli and chloramphenicol for B. subtilis. The “hag” in the vector name indicates that it is designed to be used with Phag as the promoter, but others also work fine. The flgM gene is an inhibitor of Phag; pBSChag eliminates this inhibition by integrating and disrupting flgM.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal EcoRI site found at 5854
Illegal SpeI site found at 2
Illegal PstI site found at 16
Illegal NotI site found at 9
Illegal NotI site found at 5860 - 21INCOMPATIBLE WITH RFC[21]Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal EcoRI site found at 5854
Illegal XhoI site found at 3181 - 23INCOMPATIBLE WITH RFC[23]Illegal prefix found at 5854
Illegal suffix found at 2 - 25INCOMPATIBLE WITH RFC[25]Illegal prefix found at 5854
Plasmid lacks a suffix.
Illegal XbaI site found at 5869
Illegal SpeI site found at 2
Illegal PstI site found at 16
Illegal NgoMIV site found at 1128
Illegal NgoMIV site found at 1507
Illegal NgoMIV site found at 3145
Illegal NgoMIV site found at 4284 - 1000INCOMPATIBLE WITH RFC[1000]Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal BsaI site found at 2034
Illegal BsaI.rc site found at 4108
Illegal SapI.rc site found at 1675
Illegal SapI.rc site found at 5505
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