Microbiome

Environmental DNA

16S ribosomal RNA

Bacteria

Evolutionary biology

Amplicon

Biology

Gene

Biodiversity

Deep sequencing

Metagenomics

Genome

Ecology

DNA sequencing

Computational biology

DNA extraction

Amplicon sequencing

Polymerase chain reaction

Genetics

Genome biology

Microbiome samples with low microbial biomass or severe DNA degradation remain challenging for amplicon-based or whole-metagenome sequencing approaches. Here, we introduce 2bRAD-M, a highly reduced and cost-effective strategy which only sequences ~ 1% of metagenome and can simultaneously produce species-level bacterial, archaeal, and fungal profiles. 2bRAD-M can accurately generate species-level taxonomic profiles for otherwise hard-to-sequence samples with merely 1 pg of total DNA, high host DNA contamination, or severely fragmented DNA from degraded samples. Tests of 2bRAD-M on various stool, skin, environmental, and clinical FFPE samples suggest a successful reconstruction of comprehensive, high-resolution microbial profiles.

Species-resolved sequencing of low-biomass or degraded microbiomes using 2bRAD-M