Coding

Part:BBa_K1104200:Experience

Designed by: Ting-Yun Chiang   Group: iGEM13_NYMU-Taipei   (2013-09-11)
Revision as of 14:14, 27 October 2013 by Windy6632 (Talk | contribs) (→‎SDS-PAGE)

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SDS-PAGE

We conduct an SDS Polyacrylamide gel electrophoresis for OxyR expression.

NYMU SDS.png

[[Image:NYMU_O1.png|frame|center|Device1(Part:BBa_K1104250) Setting control group contains only E.coli MG1655 and experimental group with K1104250 added to AhpCp2D1(K1104204), the electrophoresis result shows OxyR (Molecular Weight of Polypeptide: 34.276 kD) can really be expressed.

Applications of BBa_K1104200

OxyR-included circuit(Device1+Device2)

In order to enhance the strength and sensitivity of ROS-sensing promoters, we make E. coli continuously produce inactivated OxyR, so that ROS can affect promoters more easily by adding a device: Device1(Part:BBa_K1104250).

Device1 in order to enhance the effect of ROS on E. coli is composed of :

  1. a constitutive promoter(Part:BBa_J23102)
  2. RBS(Part:BBa_B0034)
  3. activator OxyR(Part:BBa_K1104200)
  4. double terminator [https://parts.igem.org/Part:BBa_B0015 Part:BBa_B0015
Device1(Part:BBa_K1104250)
a constitutive promoter(Part:BBa_J23102)+ RBS(Part:BBa_B0034)+ activator OxyR(Part:BBa_K1104200)+ double terminator Part:BBa_B0015
Device2
a sensor (OxyR-induced promoter, including TrxCp, HemHp, sufA, AhpCp1000, AhpCp2D1, AhpCp2, AhpCpD1, AhpCp1, DsbGp) + reporter (Part:BBa_E0840)

Putting Device1 BBa_K1104250 upstream to promoter AhpCp2D1(Part:BBa_K1104204) remarkably enhanced the strength of promoter AhpCp2D1(Part:BBa_K1104204) as our experiment results shows.

We designed circuit fighting against Nosema ceranae. 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

Not only for sensing the midgut oxygen stress caused by Nosemasis, in our project, there are some previous iGEM team who used OxyR-induced promoter, such as [http://2010.igem.org/Team:KIT-Kyoto/Parts 2010 KIT-Tokyo team] using it for the ink of E.coli Pen colorful. And those OxyR-induced promoters relative to the projects.

Since ROS is a natural byproduct of the metabolism and have important roles in cell signaling and homeostasis, the improvement of ROS-induced promoter is useful for sensing some specific conditions. ROS is found to involve in cancer, aging and apoptosis. Thus ROS-induced promoter in the future application can be apply to diagnosis and therapy of certain kinds of physical phenomena or disease.

Devasagaya, TPA; Tilak JC, Boloor KK, Sane Ketaki S, Ghaskadbi Saroj S, Lele RD (October 2004). "Free Radicals and Antioxidants in Human Health: Current Status and Future Prospects". Journal of Association of Physicians of India (JAPI) 52: 796.

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