Difference between revisions of "Part:BBa K4244052"
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This part contains a rhamnose inducible system upstream the Type-IC CRISPR array and Cas genes. The spacer in the CRISPR array is a random sequence that can be easily exchanged by restriction-ligation with BsaI. The presence of rhamnose induces the expression of the CRISPR-Cas system, leading to DNA cleavage. The effect of different rhamnose concentrations has been tested in Escherichia coli Nissle 1917, and it has been concluded that concentrations higher than 3 mM of rhamnose produce a high genotoxic effect in the cells, as shown in the graph.
 
This part contains a rhamnose inducible system upstream the Type-IC CRISPR array and Cas genes. The spacer in the CRISPR array is a random sequence that can be easily exchanged by restriction-ligation with BsaI. The presence of rhamnose induces the expression of the CRISPR-Cas system, leading to DNA cleavage. The effect of different rhamnose concentrations has been tested in Escherichia coli Nissle 1917, and it has been concluded that concentrations higher than 3 mM of rhamnose produce a high genotoxic effect in the cells, as shown in the graph.
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[[File:BBa K4244052-killswitch.svg|thumb|500px|center|<font size="1"> Plate reader experiment conducted during 24h. Escherichia coli Nissle 1917 containing a plasmid with this part was tested in M9 medium with 50 mM of glucose and different rhamnose concentrations (0, 0.1, 0.5, 1, 3 and 5 mM). The positive control corresponds to Escherichia coli Nissle WT in M9 medium supplemented with 50 mM of glucose. By inducing the kill switch with high concentrations of rhamnose (3 and 5 mM), we managed to significantly impair the growth of EcN, which was effectively inhibited for 15 h.</font>]]
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<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Revision as of 15:49, 10 October 2022


prhaBAD type-IC CRISPR-Cas

This part contains a rhamnose inducible system upstream the Type-IC CRISPR array and Cas genes. The spacer in the CRISPR array is a random sequence that can be easily exchanged by restriction-ligation with BsaI. The presence of rhamnose induces the expression of the CRISPR-Cas system, leading to DNA cleavage. The effect of different rhamnose concentrations has been tested in Escherichia coli Nissle 1917, and it has been concluded that concentrations higher than 3 mM of rhamnose produce a high genotoxic effect in the cells, as shown in the graph.


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Plate reader experiment conducted during 24h. Escherichia coli Nissle 1917 containing a plasmid with this part was tested in M9 medium with 50 mM of glucose and different rhamnose concentrations (0, 0.1, 0.5, 1, 3 and 5 mM). The positive control corresponds to Escherichia coli Nissle WT in M9 medium supplemented with 50 mM of glucose. By inducing the kill switch with high concentrations of rhamnose (3 and 5 mM), we managed to significantly impair the growth of EcN, which was effectively inhibited for 15 h.


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 33
    Illegal EcoRI site found at 202
    Illegal EcoRI site found at 2523
    Illegal XbaI site found at 764
    Illegal PstI site found at 196
    Illegal PstI site found at 1246
    Illegal PstI site found at 1476
    Illegal PstI site found at 2268
    Illegal PstI site found at 3541
    Illegal PstI site found at 3961
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 33
    Illegal EcoRI site found at 202
    Illegal EcoRI site found at 2523
    Illegal NheI site found at 240
    Illegal NheI site found at 518
    Illegal NheI site found at 4443
    Illegal PstI site found at 196
    Illegal PstI site found at 1246
    Illegal PstI site found at 1476
    Illegal PstI site found at 2268
    Illegal PstI site found at 3541
    Illegal PstI site found at 3961
    Illegal NotI site found at 1886
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 33
    Illegal EcoRI site found at 202
    Illegal EcoRI site found at 2523
    Illegal BglII site found at 3512
    Illegal XhoI site found at 208
    Illegal XhoI site found at 2630
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 33
    Illegal EcoRI site found at 202
    Illegal EcoRI site found at 2523
    Illegal XbaI site found at 764
    Illegal PstI site found at 196
    Illegal PstI site found at 1246
    Illegal PstI site found at 1476
    Illegal PstI site found at 2268
    Illegal PstI site found at 3541
    Illegal PstI site found at 3961
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 33
    Illegal EcoRI site found at 202
    Illegal EcoRI site found at 2523
    Illegal XbaI site found at 764
    Illegal PstI site found at 196
    Illegal PstI site found at 1246
    Illegal PstI site found at 1476
    Illegal PstI site found at 2268
    Illegal PstI site found at 3541
    Illegal PstI site found at 3961
    Illegal NgoMIV site found at 921
    Illegal NgoMIV site found at 1171
    Illegal NgoMIV site found at 1314
    Illegal NgoMIV site found at 2113
    Illegal NgoMIV site found at 2650
    Illegal NgoMIV site found at 3154
    Illegal NgoMIV site found at 4308
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 150
    Illegal BsaI.rc site found at 132