Synthesis

Overview

The overarching goal for the synthesis component of the project was to produce poly-3-hydroxybutyrate (PHB) by utilizing the nutrients present in human waste. In order to accomplish this, we:

1. analyzed human waste and chose organic compounds to use as feedstocks for our bacteria, and
2. researched and optimized efficient pathways to turn relevant components of human waste (acetyl-CoA and volatile fatty acids) into PHB

Analysis of human waste

Choice of pathways

Chassis and vector

Our gene construct was placed under an inducible promoter (lacZ, lacY). The construct was inserted into pET29(b)+ vector, which contains the promoter and kan resistant gene. The chassis used for our experiments was E. coli (BL21) because of its better ability in expression of proteins (source). (Add more reasons for why e. Coli was used) The operon was induced using IPTG at different concentrations.