BONCAT-FACS-seq reveals the active fraction of a biocrust community undergoing a wet-up event

We apply Bioorthogonal Non-Canonical Amino Acid Tagging paired with Fluorescence-Activated Cell Sorting and Metagenomic sequencing (BONCAT-FACS-Seq) to laboratory microcosms of biocrusts after a simulated rain event with the goal of better understanding the diversity and potential functional capabilities of the active and inactive microorganisms in the biocrust community. Three biological replicates of intact biocrust cores were destructively harvested at two time points following a simulated rain event. After harvest, the microcosm soil was subjected to BONCAT click-chemistry in order to fluorescently label 'translationally active' cells. Cells were extracted from the soil, stained with SYBR Green, and sorted via FACS to receive a 'total cells' pool. The resultant total cells pool was subsequently sorted based on the BONCAT fluorescence signal to receive 'active' and 'inactive' cell fractions. All cell extracts were sequenced for shotgun metagenomics. Additionally, subsamples of the starting dry biocrust was also sequenced for shotgun metagenomics.