FAVIS: Fast and Versatile protocol for metabarcoding of bulk Insect Samples from large-scale diversity monitoring projects v2

Abstract

Here we describe a non-destructive metabarcoding protocol that is optimized for high-throughput processing of Malaise trap samples and other bulk insect samples. The protocol details the process from obtaining bulk samples up to submitting them for sequencing. It is divided into four sections: 1) Laboratory workspace preparation; 2) Sample processing - decanting ethanol, measuring the wet-weight biomass and the concentration of the preservative ethanol, performing non-destructive lysis and preserving the insect material for future work; 3) DNA extraction and purification; and 4) Library preparation and sequencing. The protocol is cost-effective and relies on readily available reagents and materials. For the steps that require expensive infrastructure – such as the DNA purification robots or sequencing center services – we suggest alternative low-cost solutions when possible. The use of this protocol yields a comprehensive description of the number of species present in a given sample, their relative read abundances and the overall insect biomass. To date, we have successfully applied the protocol to more than 7000 Malaise trap samples obtained from Sweden and Madagascar. The protocol allows one lab technician to process 180 samples (2x96-well plates when we include all negative and positive controls), from bulk insect catches to ready-to-sequence libraries, in 7 working days. In other words, samples collected over 1 week from 565 Malaise traps can be processed in one month, allowing the timely delivery of the results. Thelast stepin this version contains a supplemental video with extra context and tips, as part of the protocols.io Spotlight series, featuring conversations with protocol authors.