Difference between revisions of "Part:BBa K2244011"
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<partinfo>BBa_K2244011 short</partinfo>
 
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The device is a functional plasmid containing a light repressor LEV1 ([https://parts.igem.org/Part:BBa_K2244005 BBa_K2244005]), which undergoes conformational change upon light irradiation and dimerized to bind regulatory sequence on ColE promoter ([https://parts.igem.org/Part:BBa_K2244006 BBa_K2244006]), thus inhibits target gene expression. While in darkness, dimerization does not occur thus gene expression proceeds. This device is a light repressing system with high induction efficiency and low leakage. mCherry (([https://parts.igem.org/Part:BBa_K2244008 BBa_K2244008])) is reporter gene used in this system to prove its function as well as help to characterize it. mCherry gene can be replaced by any gene sequence in light-regulated studies
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The device we constructed this year is one of the functional plasmids in the light-regulate gene expression system. With torA-opdA which is constructed in the other device. it’s can be successful expressed by using light regulate system.that can encoding organophosphorus hydrolase(OPH) to degrade the pesitcide residuce. 
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To enable secretion of OPH (gene product of opdA) to the periplasm of E. coli for the development of live cell biocatalysts, the TorA signal peptide followed by four amino acid residues of the mature TorA protein is fused directly to the N-terminal of OPH domain. TorA signal peptide contains a twin-arginine motif of ‘SRRxFLA’, and a recognition site for type I signal peptidases.
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===Biology===
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ColE promoter([https://parts.igem.org/Part:BBa_K2244006 BBa_K2244006]) sequence is derived from the promoter region of colicin E gene located in the ColE1 plasmid of E.coli. ColE promoter contains a ‘SOS’ operator region that allows the binding of LexA protein to repress transcription.opdA gene was obtained from the chromosome of Agrobacterium radiobacter. The enzyme opdA encodes is a transposable phosphotriesterase. In addition, opdA-like gene is evolutionary conserved in many bacteria species, i.e. Sphingobium fuliginis, Brevundimonas diminuta, and Mycobacterium genera, which indicates horizontal mobility.  
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TorA([https://parts.igem.org/Part:BBa_K2244008 BBa_K2244008]) is a twin-arginine signal peptide from E. coli. The twin-arginine system a bacterial protein export pathway with the remarkable ability to transport folded proteins across the cytoplasmic membrane. Proteins, with the N-terminal TorA bearing a consensus motif of SRRxFLK, are targeted to a membrane-embedded Tat translocase.  
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T1terminator is the most commonly used terminator. It seems to be reliable.
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===Functional Parameters===
 
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<partinfo>BBa_K2244011 parameters</partinfo>
 
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Revision as of 16:15, 25 October 2017

CloE promoter+TorA+opdA+T1 terminator


The device we constructed this year is one of the functional plasmids in the light-regulate gene expression system. With torA-opdA which is constructed in the other device. it’s can be successful expressed by using light regulate system.that can encoding organophosphorus hydrolase(OPH) to degrade the pesitcide residuce.

To enable secretion of OPH (gene product of opdA) to the periplasm of E. coli for the development of live cell biocatalysts, the TorA signal peptide followed by four amino acid residues of the mature TorA protein is fused directly to the N-terminal of OPH domain. TorA signal peptide contains a twin-arginine motif of ‘SRRxFLA’, and a recognition site for type I signal peptidases.

Biology

ColE promoter(BBa_K2244006) sequence is derived from the promoter region of colicin E gene located in the ColE1 plasmid of E.coli. ColE promoter contains a ‘SOS’ operator region that allows the binding of LexA protein to repress transcription.opdA gene was obtained from the chromosome of Agrobacterium radiobacter. The enzyme opdA encodes is a transposable phosphotriesterase. In addition, opdA-like gene is evolutionary conserved in many bacteria species, i.e. Sphingobium fuliginis, Brevundimonas diminuta, and Mycobacterium genera, which indicates horizontal mobility.

TorA(BBa_K2244008) is a twin-arginine signal peptide from E. coli. The twin-arginine system a bacterial protein export pathway with the remarkable ability to transport folded proteins across the cytoplasmic membrane. Proteins, with the N-terminal TorA bearing a consensus motif of SRRxFLK, are targeted to a membrane-embedded Tat translocase. T1terminator is the most commonly used terminator. It seems to be reliable.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 1286
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 383
    Illegal AgeI site found at 578
    Illegal AgeI site found at 917
  • 1000
    COMPATIBLE WITH RFC[1000]