Computational protocol: Prokaryotic diversity and biogeochemical characteristics of benthic microbial ecosystems at La Brava, a hypersaline lake at Salar de Atacama, Chile

Similar protocols

Protocol publication

[…] Diversity of the microbial community was assessed by sequence analysis the of the V4 hypervariable region of bacterial 16S rRNA using the QIIME software package v.1.7.0 []. Sequences were clustered into OTUs using UCLUST [] at the 97% similarity level using the most abundant sequence as the representative sequence for each OTU. A table was compiled with the number of sequences per OTU. Each representative OTU sequence was characterized taxonomically with the RDP classifier [] based on the Greengenes database version 12.10 [] using a bootstrap confidence of 50%. OTUs assigned to chloroplasts or mitochondria were removed from the analysis.Functional assignments were inferred based on literature search. From the taxonomic classification of OTUs, functional groups were attributed according to reported metabolisms for each known taxon [].From the OTU table, lists for each sample were obtained, and a Venn graph was generated using jvenn tool []. OTU tables were subsampled using 10 replicates for each sampling effort at increasing intervals of 100 sequences. Alpha diversity indices were calculated on each subsample of the rarefaction curve and on the complete OTU table (including all sequences) using QIIME. Alpha diversity metrics calculated included observed species, CHAO1, Shannon, Simpson, Equitability and Dominance indices. Results from bulk samples were compared using the principal coordinate analysis (PCoA) implemented in QIIME. Briefly, OTU tables were rarefied, and weighted unifrac distance matrices were built for each rarefied table. A jackknifed replicate PCoA plot was obtained from all these matrices. […]

Pipeline specifications

Software tools QIIME, UCLUST, RDP Classifier, jvenn, UniFrac
Databases Greengenes
Application 16S rRNA-seq analysis
Organisms Hemisus marmoratus
Chemicals Arsenic, Calcium, Calcium Carbonate, Carbon, Lithium, Magnesium, Superoxides