Difference between revisions of "Part:BBa K2036027"
Line 4: Line 4:
  
  
−
Cro represses pRM.CII can be rapidly degraded by FtsH.CIII acts as an inhibitor of FtsH.PRE promoter of bacteriophage λ is active only in the presence of CII protein.This whole part can reducing noise and converting pulse signal into robust persistent signal.It serves as a tool kit.
+
This is the whole characterization circuit of Signal Filter prokaryote version([File:http://2016.igem.org/Team:HUST-China Details see to HUST-China 2016 wiki]) as Fig1 shows.
−
 
+
[[File:T--HUST-China--Experiments-Fig14-1.png|thumb|500px|center|Fig1:Prokaryote version of Signal Filter characterization circuit]]
 +
<br>Here is how we conduct our characterization: We employ T7 and ptrp as our input sensors and BL21 as host strain. When induced by IPTG,Cro and CII are expressed under T7 promoter and CII will give a positive feedback to enhance Cro's expression by promoting pRE. Accumulated Cro will stably bind to the bingding site within pRM thus repress GFP and at the same time RFP expression will not be interrupted (see to Fig2) 
 +
[[File:T--HUST-China--Experiments-Fig14-2.png|thumb|500px|center|Fig2:Prokaryote version of Signal Filter characterization plasmid]]
 +
<br>When IAA is added, CI will be expressed to further block pR. And with CII's degradation by Ftsh, GFP expression will gradually comes up to a stable state. Meanwhile, RFP level will decrease by protease degradation system through LVAssrA tag. (see to Fig3)
 +
[[File:T--HUST-China--Experiments-Fig14-3.png|thumb|500px|center|Fig3:Prokaryote version of Signal Filter characterization plasmid]]
  
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Revision as of 08:06, 29 September 2016


pRE-RBS-Cro-RBS-CII-TT-ptrp-RBS-CI-TT-pR-RBS-CIII-RBS-RFP-LAAssrAtag-TT-pRM-RBS-GFP-LVAssrAtag


This is the whole characterization circuit of Signal Filter prokaryote version([File:http://2016.igem.org/Team:HUST-China Details see to HUST-China 2016 wiki]) as Fig1 shows.

Fig1:Prokaryote version of Signal Filter characterization circuit


Here is how we conduct our characterization: We employ T7 and ptrp as our input sensors and BL21 as host strain. When induced by IPTG,Cro and CII are expressed under T7 promoter and CII will give a positive feedback to enhance Cro's expression by promoting pRE. Accumulated Cro will stably bind to the bingding site within pRM thus repress GFP and at the same time RFP expression will not be interrupted (see to Fig2)

Fig2:Prokaryote version of Signal Filter characterization plasmid


When IAA is added, CI will be expressed to further block pR. And with CII's degradation by Ftsh, GFP expression will gradually comes up to a stable state. Meanwhile, RFP level will decrease by protease degradation system through LVAssrA tag. (see to Fig3)

Fig3:Prokaryote version of Signal Filter characterization plasmid

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal SpeI site found at 717
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal SpeI site found at 717
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal SpeI site found at 717
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal SpeI site found at 717
    Illegal AgeI site found at 2533
    Illegal AgeI site found at 2645
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 3561