Eco-friendly E. coli

Introduction

Our Bee. coli can express antimicrobial peptides such as defensin and abaecin to fight against Nosema cerenae. However, it is also possible that our Bee. coli can contaminate the natural environment and cause death to other species. Therefore, a light-induced lysis system was created to ensure our Bee. coli only lives inside of the bee.

Background

Blue Light Sensing Device

We chose to use [http://parts.igem.org/Part:BBa_K592016 K592016] and [http://parts.igem.org/Part:BBa_K592006 K592006] as our light sensing device. [http://parts.igem.org/Part:BBa_K592016 K592016] consists two parts: YF1 and FixJ. YF1 IS a blue-light sensor protein. It works in conjunction with its response regulator, FixJ. [http://parts.igem.org/Part:BBa_K592006 K592006] is a light-sensing promoter, which can express downstream gene when unphosphorylated FixJ presents. When exposed to blue-light, YF1 will be activated and phosphorylate FixJ. Phosphorylated FixJ cannot activate the light-sensing promoter [http://parts.igem.org/Part:BBa_K592006 K592006], and the expression of downstream gene will be repress. This picture can simply show the logic of this light-sensing device.

Blue sensing device

Self-destruct Device

The self-destruct device is composed of blue-light sensing promoter [http://parts.igem.org/Part:BBa_K592006 K592006], the blue-light sensing protein [http://parts.igem.org/Part:BBa_K592016 K592016], and a set of genes from [http://en.wikipedia.org/wiki/Toxin-antitoxin_system toxin-antitoxin system]. The toxin gene will be constitutively express. Antitoxin gene will be placed after [http://parts.igem.org/Part:BBa_K592006 K592006].

Self-destruct device

Circuit design and Experimental Method

Circuit Design

[http://parts.igem.org/Part:BBa_K592016 K592016] is cloned after constitutive promoter [http://parts.igem.org/Part:BBa_J23102 J23102], so the proteins YF1 and FixJ were continuing produced. When exposed to blue light, the inactive YF1 and FixJ will be change to their active form and induce the downstream gene of promoter [http://parts.igem.org/Part:BBa_K592006 K592006]. In our circuit, the lysis protein [http://parts.igem.org/Part:BBa_K896999 K896999] will be produced and kill our Bee. coli which escaped from the midgut of a bee.

Circuit Device

Experimental Method

First, we substitute lysis protein [http://parts.igem.org/Part:BBa_K896999 K896999] with green fluorescent protein [http://parts.igem.org/Part:BBa_E0040 E0040]. By this way, we can test the efficiency of the circuit.

Testing Circuit

Next, by comparing the number of colonies of the plate that is exposed to light and the plate that is blocked from light after 16 hours of incubation, we can characterize the functions of our device.

Left: Light is block by aluminum foil.

Right: Plate is exposed to light.