Applied Molecular Biology

Some recent cool stuff we've* been doing is to use nanopore sequencing to engage students at the undergraduate level. Recent projects involve:

Applied Molecular Biology, third year undergrad course (203340) at Massey University: students write grant proposals to sequence the DNA from a biological sample(s) of their choice (check out the 2019 projects, 2018 projects, and 2017 projects). Students peer review the grant proposals and vote for the winning project. They then head out, collect the sample, isolate DNA as a group, and use the Oxford Nanopore MinION (or in 2019-2022 the flongle) - a portable DNA sequencer to investigate the winning sample(s) at the molecular level.

• • The 2019 winning projects (we chose two, they were both so great!) were to use 16S long read sequencing to investigate the microbial communities involved in

    1. Fermenting coffee beans from Ripe Coffee in Wellington (nicest people ever!!!) Did you know that green/raw coffee beans undergo a fermentation process before roasting???! I actually didn't... and I love coffee and microbes!

    2. Investigating what microbes are in Lucas Creek, a local creek next to campus at various locations along the creek.

  • Methods: We used the Qiagen powersoil kit and the 16S nanopore kit

  • Results: The most amazing thing about this project is how well it worked. We got >400,000 sequence reads from a 1,500bp region of the 16s rRNA gene using long read nanopore sequencing using a flongle (a mini DNA sequencing flow cell that runs around $90 USD (!!)) We ran 7 samples on this flow cell at once by barcoding each sample, so it cost around ~$20 a sample when you include the reagents. We ran Kraken2 to identify what species were present then visualised everything using heat maps (using the clickable web interface Pavian (paper). The details of the lab instructions are here for those who are interested. The data we generated is here.

• • • Interestingly, the two samples of coffee that we sequenced were from very different origins (Indonesia and Tanzania), yet had very similar microbial communties (namely from the genus Caballeronia), which are known plant endosymbionts.

    • The Lucas Creek samples had a far different profile, with the major genus dominated by Limnohabitans (a bacteria that lives in freshwater lakes and streams, as expected).

    • Anyway, a data table is here of the taxonomy results for those who are *really* interested.

• • Most of this work was funded with a Massey University Teaching Support award, which I'm not sure is still available, as the link is not working?