Coding

Part:BBa_K2273110

Designed by: Nina Lautenschlaeger   Group: iGEM17_TU_Dresden   (2017-09-25)
Revision as of 14:59, 25 September 2017 by NinaL (Talk | contribs)


BlaI Repressor of the bla operon derived from Staphylococcus aureus

The blaI gene is a part used in the Beta-Lactam Biosensor project of [http://2017.igem.org/Team:TU_Dresden iGEM Team TU Dresden 2017 (EncaBcillus - It's a trap!)].

This part is a composite of the bla operon found in Staphylococcus aureus and encodes a Repressor Protein that binds to the PblaZ promoter to inhibit gene expression of the gene blaZ, coding for a beta-lactamase. If the microorganism is exposed to beta-lactam antibiotics, a receptor, named blaR1 [1], senses the compound and transducer a signal into the cytoplasm. Subsequently, the BlaI repressor Protein is degraded and frees the PblaZ promoter. Following, the bla operon is transcribed and confers resistance to the antibiotic.

This part has been codon optimized for expression in Bacillus subtilis using the online tool provided by IDT DNA. A Ribosome Binding Site (AGGAGG) specific for translation in Bacillus subtilis has been added upstream of the gene followed by a seven nucleotide spacer. Further the part features the RFC10 prefix and suffix:

Prefix with EcoRI, NotI, XbaI, RBS and spacer sequence GAATTCGCGGCCGCTTCTAGAAGGAGGTGTCAAA
Suffix with SpeI, NotI and PstI ACTAGTAGCGGCCGCTGCAGA

Sites of restriction enzymes generating compatible overhangs are indicated by sharing one color. (EcoRI and PstI are marked in blue, NotI in green, XbaI and SpeI in red

Beta-Lactam Biosensor

In this subproject, we developed a functional and complete heterologous beta-lactam biosensor in Bacillus subtilis. By the time these specified cells sense a compound of the beta-lactam family, they will respond by producing a measurable luminescence signal. We further investigated the detection spectrum of the biosensor by testing different beta-lactam antibiotics from various subclasses. For increased control and easy handling of the biosensor strain during a potential field application, we demonstrate that the encapsulation of the cells into Peptidosomes is quite advantageous.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 66


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