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Team:USTC CHINA/Parts
From 2013.igem.org
TD1, Transdermal peptide
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
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
BBa_K1074001
PHT43 is a E.coli-B.subtilis shuttle vector allowing high-level expression of secreted proteins in B.subtilis.
HBsAg
BBa_K1074002
HBsAg is the surface antigen of the hepatitis B virus (HBV). It indicates current hepatitis B infection.
Ag85b
BBa_K1074003
The antigen 85 proteins (FbpA, FbpB, FbpC) are responsible for the high affinity of mycobacteria for fibronectin, a large adhesive glycoprotein, which facilitates the attachment of M.tuberculosis to murine alveolar macrophages (AMs).
Protective Antigen Domain 4
BBa_K1074004
Protective antigen (PA) is the central component of the three-part protein toxin secreted by Bacillus anthracis, the organism responsible for anthrax.
LTB
BBa_K1074005
This eltB gene encodes for the Heat-labile enterotox(LT) in certain virulent strains of E.coli.However,in vitro,like Chinese Hamster Ovarylera toxin B subunit (CTB), it is an efficient mucosal adjuvant and carrier molecule for the generation of mucosal antibody responses and induction of systemic T-cell tolerance to linked antigens. So we use it to enhance the immune response to our transdermal vaccine.
Pctc+RBS+amilCP
BBa_K1074007
Promoter 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.
SdpA,SdpB,SpbC
BBa_K1074009
Killing factor of Bacillus subtilis ,SpbC Induces the lysis of other B.subtilis cells that have not entered the sporulation pathway, providing a source of nutrients to support sporulation. Its maturation is dependent on SdpA and SdpB. Also functions as a ligand, being able to bind to SdpI thereby triggering a signal transduction cascade that protects the cell against the toxic effects of its own SdpC. We use this killing factor to build a kill switch for Bacillus subtilis.
Pgrac+RBS+SDP
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.
PsdpRI+RBS+SDP
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.
Pgrac promoter
BBa_K1074012
Prgac promoter(consisting of the groE promoter,the lacO operator and the gsiBSD sequence) allow induction by addition of ITPG,While the background level of expression of these expression cassettes is very low in the absence of the inducer, an induction factor of about 1,300 was measured using the bgaB reporter gene (Phan et al.,2005). The amount of recombinant protein produced after addition of IPTG may represent 10 and 13%, respectively, of the total cellular protein (demonstrated when fusing the htpG and pbpE genes to the groE promoter; Phan et al., 2005).
promoter sdpR/I
BBa_K1074013
This part is a sigma factor A-dependent promotor of the gene sdpR derived from the B.subtilis ,and it contains an operon for the signaling pathway of sdpc,a bacterial toxin of B.subtilis. It's activated as the toxin sdpC existed,while the natural pructrue of the promotor, sdpR/I ,confers immunity to the SdpC toxin.
SamyQ
BBa_K1074014
SamyQ is the signal peptide of the amyQ gene encoding an α-amylase in Bacillus subtilis WB800N. Fuse it with the recombinant proteins to obtain the secretion of recombinant proteins in Bacillus subtilis.
RBS for Pgrac
BBa_K1074015
An efficient RBS for promoter Pgrac(BBa_K1074012) in Bacillus subtilis. We used it in all of our gene circuits containing promoter Pgrac.
RBS for Pctc
BBa_K1074016
This is an efficient RBS for promoter Pctc(BBa_K143010) in Bacillus subtilis. It's also an universal RBS for promoters in Bacillus subtilis. We also used it with the promoter PsdpRI(BBa_K1074013) in our project.
