Team:William and Mary

NOISE - W&M iGEM

NOISE

Characterization of promoter-driven transcriptional noise in E. coli

What is noise?

As synthetic biologists begin to construct increasingly complex gene regulatory networks, the need for accurate quantitative characterization of regulatory components becomes more pressing. Despite the extensive characterization of the average strength of the promoters available on the BioBrick registry, very few have information pertaining to the variability in their expression. Our project aims to characterize this variability, commonly referred to as stochasticity, or noise, in gene expression, for the most commonly used promoters in synthetic biology and provide additional tools for the regulation of these promoters.

Parts

In deciding which parts to submit to the iGEM Registry we focused on three main aspects.

First: ensuring our project is as reproducible and extensible as possible. To that end we have submitted all of new composite fluorescent protein parts that we constructed during the project. Second: Making genome integration as straightforward as possible for iGEM teams. In order to accomplish this goal we designed, tested, and validated a new integrator cassette that allows for simple genome integration using either 3A or Gibson Assembly. Third: Increasing the number of tools available for promoter-mediated regulation in synthetic biology. We created and validated an E. coli codon optimized dCas9 variant and a suite of gRNAs to target the most commonly used promoters in iGEM.

All Parts

A categorized list of all parts.

Part Collection

Guide RNAs for iGEMs top promoters.

Basic Part

Targeted repression with dCAS9.

Composite Part

An original integrator suite for the E. coli chromosome.

Measurement & Modeling

We measured noise in fluorescence data for dual-integrated sets of CFP and YFP under three promoters: BBa_R0010, BBa_R0011, and BBa_R0051. We also developed an analytic model of the impact of plasmid copy number fluctuations on transcriptional noise, which revealed that intrinsic noise cannot be accurately measured from reporters on the pSB1X3 plasmid series.

Data & Analysis

Plasmid Fluctuation Model

Human Practices

Our Human Practices effort was a multi-faceted outreach approach to science literacy, focusing specifically on spreading a basic understanding of synthetic biology to the general public. We collaborated with numerous organizations (logos below) to host nine educational Synthetic Biology workshops for the public (from first graders to adults!) and to implement our original, 24-activity Synthetic Biology curriculum into schools worldwide, to further sustain our efforts for years to come. Along the way, we worked with several schools and organizations. We've listed some of their logos below.