Difference between revisions of "Part:BBa K1104203"

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__NOTOC__
 
__NOTOC__
 
<partinfo>BBa_K1104203 short</partinfo>
 
<partinfo>BBa_K1104203 short</partinfo>
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==Introduction==
 
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We improve the function of a BioBrick Part: ahpC promoter([https://parts.igem.org/Part:BBa_K362001 Part:K362001])designed by [http://2010.igem.org/Team:KIT-Kyoto/Parts 2010 KIT-Tokyo team] into 6 versions.On PartRegistry, the complex part(according to [http://ecocyc.org/ECOLI/new-image?object=EG11384 Ecocyc]) composition contains hybrid promoters, shared TFBS (Transcription Factor Binding Site), and reverse promoter DsbG. In this part AhpCp1000, we succesfully mutated the Pst1 site (ctgcag->ctacag) of ahpC promoter([https://parts.igem.org/Part:BBa_K362001 Part:K362001]).
 
  
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[[File: NYMU_A1000x.png|thumb|600px|center|'''Mutation of ahpC promoter([https://parts.igem.org/Part:BBa_K362001 Part:K362001])''']]
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AhpCp1000 is a ROS-induced promoter controlled by OxyR (transcription factor) which is activated by ROS (Reactive Oxygen Species).
  
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ahpC promoter, as well as its improvement, can be activated by OxyR([https://parts.igem.org/Part:BBa_K1104200 Part:BBa_K1104200]).
+
AhpCp1000 is composed of part of dsbG coding sequence, AhpCp2 ([https://parts.igem.org/Part:BBa_K1104205 Part:BBa_K1104205]), reverse promoter DsbGp ([https://parts.igem.org/Part:BBa_K1104208 Part:BBa_K1104208]),and AhpCp1 ([https://parts.igem.org/Part:BBa_K1104207 Part:BBa_K1104207]). There are also two dual-TFBSs (Transcription Factor Binding Site) for OxyR binding between AhpCp2 ([https://parts.igem.org/Part:BBa_K1104205 Part:BBa_K1104205]), DsbGp ([https://parts.igem.org/Part:BBa_K1104208 Part:BBa_K1104208])and DsbGp ([https://parts.igem.org/Part:BBa_K1104208 Part:BBa_K1104208]),  AhpCp1 ([https://parts.igem.org/Part:BBa_K1104207 Part:BBa_K1104207]).
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[[File: NYMU_OxyRactive.png|frame|center|'''Illustration of OxyR Activation''']]
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[[File: NYMU_A1000x.png|thumb|400px|center|'''Mutation of ahpC promoter([https://parts.igem.org/Part:BBa_K362001 Part:K362001])''']]
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OxyR, a positive regulator of hydrogen peroxide-inducible genes in ''E. coli''.
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OxyR is activator of AhpCp1000 promoter. More details about OxyR can be found on the PartRegistry page: [https://parts.igem.org/Part:BBa_K1104200 Part:BBa_K1104200].
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The OxyR transcription factor regulates the H<sub>2</sub>O<sub>2</sub> response of ''E. coli''. H<sub>2</sub>O<sub>2</sub> oxidizes OxyR Cys199 to an R-SOH that reacts with Cys208 in an intramolecular disulphide bond.
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[[File: NYMU_ORD.png|thumb|300px|center|'''Structure of the OxyR Regulatory Domain'''<br />(A)Reduced Form: Inactive<br />(B)Oxidized Form: Activated by ROS<br /> ]]
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==Improvement==
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:::::<sub>Reference: D'Autréaux, B., & Toledano, M. B. (October 01, 2007). ROS as signalling molecules: mechanisms that generate specificity in ROS homeostasis. Nature Reviews Molecular Cell Biology, 8, 10, 813-824.</sub>
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 +
We improved a BioBrick Part: ahpC promoter ([https://parts.igem.org/Part:BBa_K362001 Part:K362001]) designed by [http://2010.igem.org/Team:KIT-Kyoto/Parts 2010 KIT-Tokyo team]. ahpC promoter, as well as its improvement, can be activated by OxyR ([https://parts.igem.org/Part:BBa_K1104200 Part:BBa_K1104200]). On PartRegistry, the complex part(according to [http://ecocyc.org/ECOLI/new-image?object=EG11384 Ecocyc]) composition contains hybrid promoters, shared TFBS (Transcription Factor Binding Site), and reverse promoter DsbG. In this part AhpCp1000, we succesfully mutated the PstI cutting site (ctgcag->ctacag) of ahpC promoter ([https://parts.igem.org/Part:BBa_K362001 Part:K362001]).
 +
===How ahpC ([https://parts.igem.org/Part:BBa_K362001 Part:BBa_K362001]) is improved?===
 +
'''In this part,the PstI cutting site in ahpC (K362001) is mutated at one point.'''
 +
 
 +
We annotated it thouroughly based on data from ([http://ecocyc.org/ECOLI/new-image?object=EG11384 Ecocyc]), and found that it contains DsbG coding sequence, AhpCp2 ([https://parts.igem.org/Part:BBa_K1104205 Part:BBa_K1104205]), reverse promoter DsbGp ([https://parts.igem.org/Part:BBa_K1104208 Part:BBa_K1104208]),and AhpCp1 ([https://parts.igem.org/Part:BBa_K1104207 Part:BBa_K1104207]), and a PstI cutting site. Thus we improved the promoter by first mutating the PstI cutting site in ahpCp  ([https://parts.igem.org/Part:BBa_K362001 Part:BBa_K362001]) and make AhpCp1000.
 +
 
 +
Here is the overview about the other ahpC promoter ([https://parts.igem.org/Part:BBa_K362001 Part:BBa_K362001]) improvements:
 +
{| border="1" cellpadding="5" cellspacing="0"
 +
| AhpCp1000 ([https://parts.igem.org/Part:BBa_K1104203 Part:BBa_K1104203])
 +
| The PstI cutting site is mutated.
 +
|-
 +
| AhpCp2D1 ([https://parts.igem.org/Part:BBa_K1104204 Part:BBa_K1104204])
 +
| After mutating the PstI cutting site, the truncated coding sequence from the DsbG promoter sequence is removed.
 +
|-
 +
| AhpCp2 ([https://parts.igem.org/Part:BBa_K1104205 Part:BBa_K1104205])
 +
| Only one promoter(AhpCp2) and its TFBS.
 +
|-
 +
| AhpCpD1 ([https://parts.igem.org/Part:BBa_K1104206 Part:BBa_K1104206])
 +
| Bidirectional promoter: AhpCp1 and DsbGp(reverse promoter), and their shared TFBS.
 +
|-  
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| AhpCp1 ([https://parts.igem.org/Part:BBa_K1104207 Part:BBa_K1104207])
 +
| Only one promoter(AhpCp1) and its TFBS.
 +
|}
  
 
===Usage and Biology===
 
===Usage and Biology===
 +
We designed circuit fighting against ''Nosema ceranae''. After Nosema ceranae infected midgut cells of bees, and Bee. coli should sense the pathogen first before the following circuit(fighting against Nosema ceranae)is triggered, and substance such as [https://parts.igem.org/Part:BBa_K1104300 Defensin (Part:BBa_K1104300)], [https://parts.igem.org/Part:BBa_K1104301  Abaesin(Part:BBa_K1104301)] (more details on [http://2013.igem.org/Team:NYMU-Taipei/Project/Inhibition/Killing Killing Nosema] page) in the following circuit will express.
  
 +
To enhance the strength , we added a device (more details on [http://2013.igem.org/Team:NYMU-Taipei/Project/Inhibition/Sensor Sensing Nosema] page).
 +
[[File: NYMU_O12.png|thumb|500px|center|'''Strenthening device''']]
  
 
===Related Parts===
 
===Related Parts===
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{| border="1" cellpadding="5" cellspacing="0"
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*ahpC([https://parts.igem.org/Part:BBa_K362001 Part:BBa_K362001)]
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| colspan=6 align="center"| ahpC([https://parts.igem.org/Part:BBa_K362001 Part:BBa_K362001)]
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**AhpCp1000([https://parts.igem.org/Part:BBa_K1104203 Part:BBa_K1104203])
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|-
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***[https://parts.igem.org/Part:BBa_K1104242 Part:BBa_K1104243]: AhpCp1000+[https://parts.igem.org/Part:BBa_E0840 E0840]
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| AhpCp1000 ([https://parts.igem.org/Part:BBa_K1104203 Part:BBa_K1104203])
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**AhpCp2D1([https://parts.igem.org/Part:BBa_K1104204 Part:BBa_K1104204])
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| AhpCp2D1 ([https://parts.igem.org/Part:BBa_K1104204 Part:BBa_K1104204])
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***[https://parts.igem.org/Part:BBa_K1104244 Part:BBa_K1104244]: AhpCp2D1+[https://parts.igem.org/Part:BBa_E0840 E0840]
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| AhpCp2 ([https://parts.igem.org/Part:BBa_K1104205 Part:BBa_K1104205])
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**AhpCp2([https://parts.igem.org/Part:BBa_K1104205 Part:BBa_K1104205])
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| AhpCpD1 ([https://parts.igem.org/Part:BBa_K1104206 Part:BBa_K1104206])
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***[https://parts.igem.org/Part:BBa_K1104245 Part:BBa_K1104245]: AhpCp2+[https://parts.igem.org/Part:BBa_E0840 E0840]
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| AhpCp1 ([https://parts.igem.org/Part:BBa_K1104207 Part:BBa_K1104207])
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**AhpCpD1([https://parts.igem.org/Part:BBa_K1104206 Part:BBa_K1104206])
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| DsbGp ([https://parts.igem.org/Part:BBa_K1104208 Part:BBa_K1104208])
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***[https://parts.igem.org/Part:BBa_K1104246 Part:BBa_K1104246]: AhpCpD1+[https://parts.igem.org/Part:BBa_E0840 E0840]
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|-
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**AhpCp1([https://parts.igem.org/Part:BBa_K1104207 Part:BBa_K1104207])
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| AhpCp1000+[https://parts.igem.org/Part:BBa_E0840 E0840] ([https://parts.igem.org/Part:BBa_K1104243 Part:BBa_K1104243])
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***[https://parts.igem.org/Part:BBa_K1104247 Part:BBa_K1104247]: AhpCp1+[https://parts.igem.org/Part:BBa_E0840 E0840]
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| AhpCp2D1+[https://parts.igem.org/Part:BBa_E0840 E0840] ([https://parts.igem.org/Part:BBa_K1104244 Part:BBa_K1104244])  
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**DsbGp([https://parts.igem.org/Part:BBa_K1104208 Part:BBa_K1104208])
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| AhpCp2+[https://parts.igem.org/Part:BBa_E0840 E0840] ([https://parts.igem.org/Part:BBa_K1104245 Part:BBa_K1104245])  
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***[https://parts.igem.org/Part:BBa_K1104248 Part:BBa_K1104248]: DsbGp+[https://parts.igem.org/Part:BBa_E0840 E0840]
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| AhpCpD1+[https://parts.igem.org/Part:BBa_E0840 E0840] ([https://parts.igem.org/Part:BBa_K1104246 Part:BBa_K1104246])  
 +
| AhpCp1+[https://parts.igem.org/Part:BBa_E0840 E0840] ([https://parts.igem.org/Part:BBa_K1104247 Part:BBa_K1104247])
 +
| --
 +
|}
  
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Latest revision as of 14:05, 21 October 2013

AhpCp1000

AhpCp1000 is a ROS-induced promoter controlled by OxyR (transcription factor) which is activated by ROS (Reactive Oxygen Species).

AhpCp1000 is composed of part of dsbG coding sequence, AhpCp2 (Part:BBa_K1104205), reverse promoter DsbGp (Part:BBa_K1104208),and AhpCp1 (Part:BBa_K1104207). There are also two dual-TFBSs (Transcription Factor Binding Site) for OxyR binding between AhpCp2 (Part:BBa_K1104205), DsbGp (Part:BBa_K1104208)and DsbGp (Part:BBa_K1104208), AhpCp1 (Part:BBa_K1104207).

Mutation of ahpC promoter(Part:K362001)

OxyR is activator of AhpCp1000 promoter. More details about OxyR can be found on the PartRegistry page: Part:BBa_K1104200.

Improvement

We improved a BioBrick Part: ahpC promoter (Part:K362001) designed by [http://2010.igem.org/Team:KIT-Kyoto/Parts 2010 KIT-Tokyo team]. ahpC promoter, as well as its improvement, can be activated by OxyR (Part:BBa_K1104200). On PartRegistry, the complex part(according to [http://ecocyc.org/ECOLI/new-image?object=EG11384 Ecocyc]) composition contains hybrid promoters, shared TFBS (Transcription Factor Binding Site), and reverse promoter DsbG. In this part AhpCp1000, we succesfully mutated the PstI cutting site (ctgcag->ctacag) of ahpC promoter (Part:K362001).

How ahpC (Part:BBa_K362001) is improved?

In this part,the PstI cutting site in ahpC (K362001) is mutated at one point.

We annotated it thouroughly based on data from ([http://ecocyc.org/ECOLI/new-image?object=EG11384 Ecocyc]), and found that it contains DsbG coding sequence, AhpCp2 (Part:BBa_K1104205), reverse promoter DsbGp (Part:BBa_K1104208),and AhpCp1 (Part:BBa_K1104207), and a PstI cutting site. Thus we improved the promoter by first mutating the PstI cutting site in ahpCp (Part:BBa_K362001) and make AhpCp1000.

Here is the overview about the other ahpC promoter (Part:BBa_K362001) improvements:

AhpCp1000 (Part:BBa_K1104203) The PstI cutting site is mutated.
AhpCp2D1 (Part:BBa_K1104204) After mutating the PstI cutting site, the truncated coding sequence from the DsbG promoter sequence is removed.
AhpCp2 (Part:BBa_K1104205) Only one promoter(AhpCp2) and its TFBS.
AhpCpD1 (Part:BBa_K1104206) Bidirectional promoter: AhpCp1 and DsbGp(reverse promoter), and their shared TFBS.
AhpCp1 (Part:BBa_K1104207) Only one promoter(AhpCp1) and its TFBS.

Usage and Biology

We designed circuit fighting against Nosema ceranae. After Nosema ceranae infected midgut cells of bees, and Bee. coli should sense the pathogen first before the following circuit(fighting against Nosema ceranae)is triggered, and substance such as Defensin (Part:BBa_K1104300), Abaesin(Part:BBa_K1104301) (more details on [http://2013.igem.org/Team:NYMU-Taipei/Project/Inhibition/Killing Killing Nosema] page) in the following circuit will express.

To enhance the strength , we added a device (more details on [http://2013.igem.org/Team:NYMU-Taipei/Project/Inhibition/Sensor Sensing Nosema] page).

Strenthening device

Related Parts

ahpC(Part:BBa_K362001)
AhpCp1000 (Part:BBa_K1104203) AhpCp2D1 (Part:BBa_K1104204) AhpCp2 (Part:BBa_K1104205) AhpCpD1 (Part:BBa_K1104206) AhpCp1 (Part:BBa_K1104207) DsbGp (Part:BBa_K1104208)
AhpCp1000+E0840 (Part:BBa_K1104243) AhpCp2D1+E0840 (Part:BBa_K1104244) AhpCp2+E0840 (Part:BBa_K1104245) AhpCpD1+E0840 (Part:BBa_K1104246) AhpCp1+E0840 (Part:BBa_K1104247) --

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]