We are happy. Happy about our successful first step on the way of teasing secrets out of freshly caught deep-sea isopods from 3000 m depth. Which means, we have extracted DNA and after the first successful runs prepared extractions all day long, highly motivated. How we got there:
After META, our epibenthic sledge, had brought the samples for us on deck, we divided the sample immediately by weight and live-sorted one-half in the cooling container at 0°C and preserved the other half in precooled alcohol at –20°C. While live sorting we got a first impression of the creatures that awaited us: bristle worms (Polychaeta), amphipods, several forams and, among many other taxa, our target group, the isopods. (By then we were exhausted but happy!)
The individuals picked in the cooling container were fixed in RNAlater (a special solution to store DNA) or shock frozen at –80°C. These animals we want to work up later to compare the suitability of both fixation methods for genetic studies.
After that we had to wait 48 hours before we could sort the alcohol-fixed samples. A test of our patience, as we were very curious what would be hidden in the deep-sea mud... During the sorting some 200 isopods of many shapes and forms were discovered. Each individual is photographed and catalogued. This is our first contribution to the “Barcode of Life” project “Barcoding CeDAMar Isopoda”. Our work focuses, aside from the barcoding aspect, on certain families whose relationships we want to reveal with the help of genetic methods. And of those families, we already found several individuals in the samples!
In a second step, we steal three to four legs from each isopod (out of 14 legs or 7 pairs of legs which isopods possess.) This is not a trivial task, considering how minute isopods are ( 1-10 mm)... The biggest challenge is to transfer the tiny extremities into Eppendorf vials. On a single day we worked up some 50 isopods, and the next batch of extractions is already sitting in a 56°C water bath for lysis. This was a successful day in every meaning of the word.
Out of a total of five strategies to process samples, we finally found the one that is successful, which will be a base for analyses at home. Now that the DNA extraction with classical kits is working, the question is, how will the RNAlater and the frozen animals work? Is there a difference between extracts from animals processed right after the catch and those fixed in alcohol? There are more revelations on the horizon...
Saskia Brix, Senckenberg
Photos: S. Brix and M. Schüller, Ruhr-University of Bochum