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Team:BostonU/Project Overview
From 2012.igem.org
Project Overview
- Building: Build Genetic Circuits with MoClo Parts
- Characterizing: Characterize Circuits using Flow Cytometry
- Sharing: Generate Data Sheet for MoClo Parts
Our project has two major goals: 1. To introduce MoClo as an alternative assembly method for use by iGEM teams and 2. To develop a standard protocol for the characterization of genetic circuits containing fluorescent proteins and share this protocol with the synthetic biology community.
In order to achieve these goals, there many steps must be taken. First, we must convert BioBrick Parts into MoClo Parts using PCR. Upon converting basic parts, our project will have three thrusts:
As we worked towards our first goal over the summer months, we ran into difficulties with some of our PCR and cloning reactions. This has unfortunately delayed our other goals, but we're still working hard towards generating a characterization workflow and a MoClo data sheet format for the Jamboree.
Abstract
Abandon All Hope, Ye Who PCR: MoClo and the Quest for Genetic Circuit Characterization
Monique De Freitas¹, Shawn Jin¹, Evan Appleton², Swati Carr², Sonya Iverson², Traci Haddock³, and Douglas Densmore§
¹iGEM Team Member, ²iGEM Team Mentor, ³iGEM Team Advisor, §Faculty Sponsor, Department of Electrical and Computer Engineering, Boston University, Boston, MA, USA
Our project aims to develop a standardized protocol for the characterization of genetic circuits using flow cytometry. We built a significant number of both simple and complex genetic circuits that were characterized using flow cytometry. These genetic circuits were built using an assembly technique called MoClo (developed by Weber et al, 2011), which involves a multi-way, one-pot digestion-ligation reaction, enabling faster and more efficient construction of genetic circuits. We converted a large subset of BioBrick Parts from the Registry into MoClo Parts using PCR and cloning strategies. We built and characterized various genetic circuits using MoClo Parts and compared them against their pre-existing BioBrick counterparts in order to compare the characterization results from the two assembly techniques. We also present a standard data sheet to be included in the Registry of Standard Biological Parts for each Part we characterized to easily share our data with the synthetic biology community.
