Team:Cooper Union/Biohack project

The Biohacker Kit

The premise of this project was to create a two plasmid system that was linked to create a bio-system whose inputs and outputs can be easily swapped. The purpose of this system is to create a means by which students can be easily introduced to synthetic biology (the design and re-design of biological systems for useful purposes) in their classrooms without the need for high-tech, expensive equipment.

How It Works

The system uses two different types of plasmids (pACYC184 as the input plasmid and pBR322 as the output plasmid). The input plasmid had the promoter gene along with a phage activator ligated into it and the reporter promoter had a phage promoter and a reporter gene ligated into it. When the promoter is activated, it will trigger the phage activator. The phage activator will then activate the phage promoter in the output plasmid, which will turn on the output gene. In this way, there could be a whole palate of input and output plasmids that can be mixed and matched at the students' will to easily create their own individual systems. The two plasmids have different antibiotic resistances so that one can tell when their programmed plasmids have been successfully transformed into cells, along with different origins of replication. Using two different antibiotic resistances will help ensure that the colonies have both plasmids co-transformed inside the E. coli cells.



The Biohacker Kit would be a box that schools could buy that contained all of the input and output constructs. The lab work involved would be to co-transform the input and output of the student's choice with E. coli cells, and plate them on the correct antibiotic plates. After overnight incubation, the students can see all of the colonies and then activate the promoter to kick-start the system. Then, depending on the promoter, over the next day or two the system will begin to express the reporter gene! Students will also have the ability to monitor the system to see how the gene expression varies with time.

Results

We used this part to design a composite part, BBa_K1354000, with this promoter and a phage activator, BBa_I746352. This phage activator turned on a phage promoter with a GFP reporter composite part, BBa_I746321. The two composite parts were in separate plasmids. When shocked with UV, the UV promoter was turned on, which activated the phage activator. This then turned the phage promoter on which activated the GFP reporter. The fluorescence readings are shown below.

Future Work