Team:Chalmers-Gothenburg/Wetlab
Overview
This page contains everything you need to know about our laboratory work! Use the links below or the drop-down menu in the head bar to navigate between different sections. We recommend that you start by reading the design page to get an overview, then check out the results section and finish by looking at our measurements page. While reading, if there is anything you want to know more details about, go to Notebook and find it in the weekly notebook links!
Design
Gain a deeper insight into how we designed the experimental part of our project! Read about the wet lab process step-by-step; from identifying enzymes that degrade Elastane to designing our constructs and experimental course of action. You will also get a more detailed description of adjustments made according to input we received from stakeholders and experts in the industry. Finally, we present future directions and considerations as we hope that future iGEM teams will keep building on our idea.
Results
When you have read the design page you might wonder, how did it all go? Wonder no more! This page will take you through the results in a chronological matter, from gel electrophoresis pictures in the gene amplification phase to Western Blot membranes from the protein expression validation phase. We show successful his-tag introduction using PCR for all our nine enzymes and positive colony screening for several of the enzymes. Finally, we present western blot results that confirm the expression of BphC and discuss an aggregation problem that was identified.
Measurements
In this page we present the activity measurements we attempted of our expressed BphC enzyme. This was done by tracking the decrease in absorbance at 434 nm, the wavelength at which 2,3-dihydroxybiphenyl has been shown to absorb light. While no conclusions could be drawn based on the measurement, we calculate what results would be expected based on Beer’s law and present a rigorous experimental design where substrate and enzyme amount are varied.
Notebook
This page breaks down the entire project into actual experiments and laboratory procedures. What protocols did we follow when performing western blot? Which primers were used when introducing the his-tags? How many negative screening results did we go through before finding a positive colony? Check out our notes compiled on a weekly basis, as well as protocols and a complete list of the primers used.
