From Sample to Species Identity

The roads to El Tatio remained closed through yesterday and today, which unfortunately meant that we could not see the geysers or silica sinters. The heavy rains and snow were part of the worst storm in the region in over 60 years. I suppose we should be glad we witnessed such a rare event. Luckily for us, the Atacama Desert is a beautiful place that’s used as an analog for Mars and to test landers, rovers and other Mars-bound instruments. Between the extreme aridity (just an average of 15 mm of rain per year), red rocks, and spectacular sand dunes, it is not difficult to imagine that you’re standing on a martian surface! So, we decided to try out our DNA extraction, sequencing and analysis in this remote setting.

We picked two sites, one yesterday and one today, to test remote field sequencing with offline bioinformatics. Our first site was in cactus-lined hills between Machuca and Guatín. The presence of vegetation and a small stream below suggested that there might be enough biomass in this area for our very first test. Our second site had no vegetation or water, beautiful large sand dunes, and veins of gypsum throughout. The veins looked just like those on images taken by the Curiosity rover!

For the in situ extractions, we collected ~200 mg sandy material into a 2ml tube containing DNA/RNA shield and bashing beads, bead-beat the sand using the TerraLyzer, and then recovered the DNA in the lysate using a spin column. Next, we made a Nanopore rapid sequencing library (RAD002), loaded it onto the MinION and sequenced without live basecalling or “pinging” ONT. After 60 (yesterday) or 90 (today) minutes of sequencing, we paused sequencing, basecalled the reads, and classified the resulting sequences using Centrifuge software with archaeal and bacterial reference genomes. We then restarted sequencing for later analyses. We’ll bring some the DNA back with us to look at the fragment length distribution and DNA purity (A260/280 ratio). To check for contamination, we extracted negative controls, and will sequence them once we return. In addition to being completely offline, we used portable power – a powerbank to run the miniPCR for incubations at 30°C, 75°C and 22°C (“RT”) and the laptop’s battery for laptop and MinION.

A brief look at the taxonomic assignments indicated that we sequenced different communities at the two sites. There were >100 distinct genera at the first site and ~200 at the second. While some assignments overlapped, the distribution and identities were different from each other (and Thursday’s community standard). Looking forward to comparing these field-based assignments with longer lab runs and more in-depth analyses!

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