Stolen moments (Vol 2)
From counting birds at Christmas to cataloguing bacteria in the human microbiome, Citizen Science is a template of scientific participation that can be co-opted by teachers to engage large numbers of students in research training.
I’m in the process of designing a brand new lab course for Semester 2, and this series of blog entries will be my attempt to “think out loud” and steal some time away from the pressure of looming deadlines... Last time we talked about recombinant DNA technology and how molecular cloning of specific genes can be adapted for citizen science projects. Following on from cloning novel plasmids that encode genes of interest, students can work with their own plasmids, or characterise existing plasmid libraries generated by your research labs to express and purify novel proteins of interest.
Inquiry Idea 1: Optimising Variables
Just like molecular cloning, in protein expression experiments if one step fails, everything fails. Giving students more autonomy in experimental design does not have to be impossibly difficult, you can limit the variables that students can change in each experiment and provide a scaffolded inquiry-driven learning activity. It helps that each protocol within protein chemistry has many variables that can be optimized, including incubation times, reagent concentrations, and choice of buffer solutions, and students can be given a high-degree of autonomy to survey the literature in search of optimal experimental conditions for their protein of interest.
Inquiry Idea 2: Choosing Target Proteins
Students can be allocated a protein of interest to work on, or choose from a subset of closely-related novel proteins. The experimental workflow involves transformation of plasmids into cells, most commonly using bacteria expression systems, induction of protein over-expression, and detecting of protein levels through SDS-PAGE and Western-Blotting. Even though this is largely a linear sequence of experimental protocols, giving students autonomy in choosing the target protein to work on empowers them to adopt inquiry-based learning. You can of course pair this with Inquiry Idea 1 and ask students to optimise variables at each step along the way
Inquiry Idea 3: Protein Purification + Functional Experiments
After this point it will depend on the features of the recombinant plasmid and the function of the gene product of interest. You may ask students to purify the protein using a variety of different techniques - gel-filtration, ion-exchange and/or affinity chromatography, and/or run functional experiments to measure protein activity. Of course there are almost an unlimited number of ways of defining “protein activity”, but focusing on protein families with known enzymatic activity is a pragmatic approach. You can run colorimetric assays where the enzyme activity results in a colour change which can be read by a plate reader, and very quickly analyse many different conditions with different protein products.
Inquiry Idea 4: Site-directed Mutagenesis
Site-directed mutagenesis experiments involve systematically mutating individual protein residues one by one to see how and when the protein function changes in response to these mutations. These mutant plasmid libraries are very useful to research labs but are otherwise considered a routine mechanistic exercise for researchers, so empowering undergraduate students to do this work at scale makes sense for everyone. Remember the three components of Citizen Science – authenticity, scalability, and communicability, and this work is authentic to the work researchers do in their labs. Students can either be involved in generating the mutant plasmid libraries, or be given an existing mutant plasmid library to work with. There could potentially be hundreds of mutants for a single protein, so even in a class with hundreds of students you can give each student or pairs of students their own mutant to work on. The mutant protein is then expressed, purified, and its enzyme activity measured, and the data across the whole class is pooled to figure out which amino acid residues are directly involved in the enzyme activity of the wild-type protein. Hopefully you can see by now that the iterative nature of these protocols, needing to be repeated for each mutant, inherently makes these lab activities scalable – a great fit for large class sizes, and ticking the “scalability” box for Citizen Science.
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That’s enough procrastination (and catharsis) for the moment… more details in the video below, and we’ll talk about genomics + bioinformatics tools in teaching next time.
Jack.
