Part:BBa_K842013
fliC-GFP
When transcribed, RBS-fliC-GFP generates the flagellum protein bonded with a green fluorescent protein. This structure would then be exported to the growing flagella apparatus and used in the construction of the filament. When strongly expressed, this structure would directly result in the flagella filament's containing GFP, allowing for the flagella to be detectable. Unfortunately we were unable to promote the sequence enough to test whether this construct actually functions in this manner.
Cloning and Uses: Both the fliC were cloned via PCR from the E. coli strain DH5α and cloned into the vector pSB1C3. Due to the fact that fliC has multiple S and P restriction enzyme sites located within its DNA sequence, it was generated with XbaI sites on the forward and reverse primer. This is so as to allow for the placement of a gene promoter in the E-X sites upstream of the construct in an iGEM biobrick vector. It is important to note though that the current structure consists of the following so as to avoid the accidental loss of a gene during the cloning procedure.
Used for reconstitution of flagella apparatus in E. coli B strains.
EcoRI-XbaI-RBS-fliC-XbaI-GFP-SpeI-PstI
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal SpeI site found at 547
Illegal PstI site found at 872
Illegal PstI site found at 919
Illegal PstI site found at 1486 - 12INCOMPATIBLE WITH RFC[12]Illegal SpeI site found at 547
Illegal PstI site found at 872
Illegal PstI site found at 919
Illegal PstI site found at 1486 - 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 1224
- 23INCOMPATIBLE WITH RFC[23]Illegal SpeI site found at 547
Illegal PstI site found at 872
Illegal PstI site found at 919
Illegal PstI site found at 1486 - 25INCOMPATIBLE WITH RFC[25]Illegal SpeI site found at 547
Illegal PstI site found at 872
Illegal PstI site found at 919
Illegal PstI site found at 1486
Illegal AgeI site found at 301
Illegal AgeI site found at 709 - 1000COMPATIBLE WITH RFC[1000]
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