Team:Tokyo Tech/Project/wolf coli


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iGEM Tokyo Tech 2010 "E.coli with Humanity"

4 Wolf coli. Overview



Figure3-1. Introduction of “Wolf coli.”

Regulation of Artificial Cooperation System (ACS)lead to alternative behavioral of E. coli.Our team challenged to design “Wolfcoli”, an E coli. that its behavior changes due to the intensity of lightinspired by famous legend of wolfman. When there is intermediate light intensity, two cells doesn’t help each other at only full moon night .  From these behaviors, we call this “Wolfcoli” (Fig. 3-1).

Figure3-2. Overview of “Wolfcoli” system

In order to create the “Wolf coli.”, we introduced " red-light-dependent gene expression network"[1] and "band-detect network"[2] and combined these networks with the Artificial Cooperation System (Fig. 3-2). We conducted experiment on series of OmpC promoter (PompC), and LacIM1 which are crucial parts of our networks.


Figure3-3. The induction of new PompC series in high osmolarity medium at hours. This work is done by Thiprampai THAMAMONGOOD and Taichi NAKAMURA
Figure3-4.Repression efficiency of LacIM1 (BBa_K395401) / LacIWT (BBa_K395402) exposed to arabinose and IPTG. This work is done by Mitsuhiko ODERA

We succeeded in designing 2 new osmoregulative promoters, PompC(CB) (BBa_K395302) and PompC(CS1) (BBa_K395303), which can be utilized in the red-light-dependent gene expression network (Fig. 3-3).

We characterized lacIM1 (BBa_K082026), a mutation of LacI (BBa_?????),which is a key component in the band-detect network. In fact, this part was registered by USTC(2008) [3], however it was not well characterized in the BioBrick registry. We found that product of lacIM1 shows weaker repression to Plac than its wild type. (Fig. 3-4)

Wolf coli. and Artificial Cooperative System

The Aftificial Cooperation System was designed to be switching off during the “full moon night” which means sympathetic coli. would be transformed into wolf coli. by induction of the “full moon light”. During this period, communication between 2 types of cells is inhibited. Therefore, both types of cells are competitors and they are unable to recognize each other by quorum sensing.

Tokyotech Cooperative activity.jpg

Artificial Cooperation system turns on during the daytime and the crescent-moon night which means the systems is activated by either weak or strong light intensity. In contrast, the system is off during the full moon night due the inhibition of intermediate light intensity.

POmpC in red-light-dependent gene expression network

PompC(WT) and the two-component system

OmpC promoter, a key in light sensing system, plays a crucial role in initiating the transformation of sympathetic coli. into Werewolf coli. in light sensitive system. To accomplish band-detect circuit in light sensitive system, varieties of OmpC promoters were designed and characterized in order to find an appropriate strength of the promoter particularly which can be activated by the “full moon light”.

LacIMI in band-detect network

PompC(WT) and the two-component system

This system is accomplished by integrating the red-light-dependent gene expression network and the band-detect network (Fig. 3-2).