Difference between revisions of "Part:BBa K1088013"
 
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<partinfo>BBa_K1088013 short</partinfo>
 
<partinfo>BBa_K1088013 short</partinfo>
  
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This part consist of the ''dxs'' gene derived from ''B. subtilis'' under the control of the lac promoter and has a strong RBS.
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This part consist of the ''dxs'' gene derived from ''B. subtilis'' under the control of the ''lac'' promoter and has a strong RBS.
  
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To repress expression from the lac promoter, the ''lacI:LVA'' gene (under a constitutive promoter, with a strong RBS and a efficient terminator) was placed counter-clockwise to the ''dxs'' gene. The repression can be relieved with addition of IPTG, which binds and inhibits the function of LacI:LVA.  
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To repress expression from the ''lac'' promoter, the ''lacI:LVA'' gene (under a constitutive promoter with a strong RBS and a efficient terminator) was placed upstream to the ''dxs'' gene. The repression can be relieved with addition of IPTG, which binds and inhibits the function of LacI:LVA.  
  
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The levels of expression was meassured with a similar Brick with linker-GFP fused to Dxs (BBa_1088009) in ''E. coli'' K-12 MG1655 using fluorescence activated cell sorting (FACS). The experiment proved that the reporter fusion was only very weakly expressed when the strain was grown without IPTG in the media, and that the reporter fusion was expressed at much higher levels after addition of IPTG. Though, in the same experiment we compared the LVA-tagged LacI with the natural and found that the natural LacI responded better to the IPTG addition. See experience for more details.
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The levels of protein expression were measured with a similar brick with linker-GFP fused C-terminal to Dxs ([https://parts.igem.org/Part:BBa_1088009 BBa_1088009]) in ''E. coli'' K-12 MG1655 using fluorescence activated cell sorting (FACS). The experiment proved that the reporter fusion wasn't expressed when the strain was grown without IPTG in the media, and that the expression was at its maximum approximately 90 min after induction with 1 mM IPTG. See [https://parts.igem.org/Part:BBa_1088009 BBa_1088009] for more details.
  
 
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<!-- Add more about the biology of this part here

Latest revision as of 18:47, 28 October 2013

B. subtilis dxs (lac promoter with lac inhibitor: IPTG inducible)

This part consist of the dxs gene derived from B. subtilis under the control of the lac promoter and has a strong RBS.

To repress expression from the lac promoter, the lacI:LVA gene (under a constitutive promoter with a strong RBS and a efficient terminator) was placed upstream to the dxs gene. The repression can be relieved with addition of IPTG, which binds and inhibits the function of LacI:LVA.

The levels of protein expression were measured with a similar brick with linker-GFP fused C-terminal to Dxs (BBa_1088009) in E. coli K-12 MG1655 using fluorescence activated cell sorting (FACS). The experiment proved that the reporter fusion wasn't expressed when the strain was grown without IPTG in the media, and that the expression was at its maximum approximately 90 min after induction with 1 mM IPTG. See BBa_1088009 for more details.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 2505
    Illegal EcoRI site found at 3162
    Illegal PstI site found at 2563
    Illegal PstI site found at 3009
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 2505
    Illegal EcoRI site found at 3162
    Illegal NheI site found at 7
    Illegal NheI site found at 30
    Illegal PstI site found at 2563
    Illegal PstI site found at 3009
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 2505
    Illegal EcoRI site found at 3162
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 2505
    Illegal EcoRI site found at 3162
    Illegal PstI site found at 2563
    Illegal PstI site found at 3009
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 2505
    Illegal EcoRI site found at 3162
    Illegal PstI site found at 2563
    Illegal PstI site found at 3009
    Illegal NgoMIV site found at 2462
    Illegal AgeI site found at 2355
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 2304
    Illegal SapI.rc site found at 3003