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Team:USTC CHINA/Partsat1
Parts
TD1, Transdermal peptide Main Page-BBa_K1074000 TD1 is a short synthetic peptide(ACSSSPSKHCG)identified by in vivo phage display, facilitated efficient transdermal protein delivery through intact skin. Studies suggested that the peptide creates a transient opening in the skin barrier to enable macromolecular material to reach systemic circulation
Pgrac+RBS+SamyQ+TD1+GFP Main Page-BBa_K1074006 This is the main circuit of our project to allow high expression of target protein(antigen,adjuvant).GFP can be substituted by various proteins via a modular PCR or standard cut/ligation method.
PHT43 PHT43 is a E.coli-B.subtilis shuttle vector allowing high-level expression of secreted proteins in B.subtilis. It is based on the strong σA-dependent promoter preceding the groE operon of B. subtilis which has been converted into an efficiently controllable (IPTGinducible)promoter by addition of the lac operator. An efficient Shine-Dalgarno (SD) sequence as well as a multiple cloning site (BamH I, Xba I, Aat II, Sma I) were also inserted. To obtain secretion of recombinant proteins, the coding region for the signal peptide of the amyQ gene encoding an α-amylase was fused to the SD sequence.
HBsAg HBsAg is the surface antigen of the hepatitis B virus (HBV). It indicates current hepatitis B infection. Sequence and Features
Composite parts
BBa_K1074007Promoter ctc(BBa_K143010) is a sigma factor B-dependent promoter in B. subtilis. Activated by endogenous sigma factor B under mild stress( nutrient stress response or physical stress response). amilCP(BBa_K592009), blue chromoprotein,naturally exhibits strong color when expressed. when bacterial grow to the post-log phase, nutrient and space stress arise,Promoter ctc should be activited. We assemble this gene circuit just to expect a visual signal when B.subtilis grow to the post-log phase. BBa_K1074010 Spbc in SDP(sdpA,sdpB,spbC)(BBa_K1074009)operon is a killing factor of Bacillus subtilis. We put the SDP operon under the strong Regulated promoter Pgrac(BBa_K1074012) to see whether overexpression of spbc can induce the suicide of engineered Bacillus subtilis. BBa_K1074011 We construct this gene circuit as our kill switch to kill the engineered Bacillus subtilis for the safety purpose.Killing is mediated by the exported toxic protein SpbC. Extracellular SpbC induces the synthesis of an immunity protein, SdpI, which protects toxin-producing cells from being killed. SdpI, a polytopic membrane protein, is encoded by a two-gene operon under sporulation control that contains the gene for an autorepressor, SdpR. The autorepressor binds to and blocks the promoter for the operon. Evidence indicates that SdpI is also a signal-transduction protein that responds to the SpbC toxin by sequestering the SdpR autorepressor at the membrane. Sequestration relieves repression and stimulates synthesis of immunity protein. The kill switch is based on a high-copy vector fused with promoter for operon sdpIR and coding sequence for protein SpbC. When SpbC toxins are sensed, they will be captured by Immunity Protein SdpI at the membrane, enabling SdpI to sequester SdpR. As a result, repression on promoter SdpIR is released and more SpbC will be produced. Trapped in this endless loop, the SpbC producing cells fails to cope with enormous toxin SpbC and doomed after eliminating their siblings. Eventually, the group of engineered Bacillus subtilis is destroyed instead of sporulating.
HbpR BBa_K1031300 is the coding sequence of HbpR with terminators. BBa_K1031301 and BBa_K1031302 are the RBS library of HbpR biosensor's reporter.
HcaR BBa_K1031410 is the coding sequence of HcaR with terminators. BBa_K1031441 BBa_K1031442 BBa_K1031443 and BBa_K1031444 are the Pc library of HbpR to tune its performance . BBa_K1031420 and BBa_K1031421 are the RBS library of HcaR biosensor's reporter.
CapR BBa_K1031100 is the coding sequence of CapR transcriptional factor. BBa_K1031111 and BBa_K1031112 are basic sensor construction with different CapR expression level. BBa_K1031113 and BBa_K1031114 are reporters with different RBS for the CapR biosensor.
HpaR BBa_K1031510 is the coding sequence of HpaR transcriptional factor. BBa_K1031520 and BBa_K1031521 are reporters with different RBS for the HpaR biosensor.
NahR BBa_K1031610 is the NahR biosensor circuit.
PaaX BBa_K1031710 is the coding sequence of PaaX with terminators. BBa_K1031742 BBa_K1031743 and BBa_K1031744 are the Pc library of PaaX.
XylS BBa_K1031911 is the coding sequence of XylS transcriptional factor with terminators. BBa_K1031912 BBa_K1031913 and BBa_K1031916 are the Pc library of XylS.
XylR BBa_K1031803 BBa_K1031804 are the RBS library of XylR biosensor's reporter.
Parts for Adaptors
NahF BBa_K1031620 is the coding sequence of XylS transcriptional factor with terminators. BBa_K1031621 BBa_K1031622 BBa_K1031623 BBa_K1031624 and BBa_K1031625are the Pc library of NahF.
Parts for Band-Pass Filter
BBa_K1031021 is the hybrid promoter for the reporter node, which can be activated as well as inhibited. BBa_K1031011 is the transcriptional factor that can activate its cognate promoter, which is used as the activate node in the band-pass filter. BBa_K1031013 BBa_K1031014 BBa_K1031015 BBa_K1031016 are RBS library to tune the expression of C1, under the control of pSal promoter, which is a crucial part for band-pass filter.
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