Single cell sequencing of environmental cells (emCells)

Dr Haylea Miller1, Ms Miwa  Takahashi1, Dr Henry  Hui2, Associate Professor Kathy Fuller2, Professor Simon  Jarman3, Dr Olly  Berry1

1Csiro, Perth, Australia, 2Translational Cancer Pathology Laboratory, UWA, Perth, Australia, 3University of Western Australia, Perth, Australia

Abstract:

Environmental DNA (eDNA) metabarcoding has radically altered ecological research, because it permits rapid, accurate species detection from environmental samples, without capture or observation. However, because eDNA analysis targets few, short DNA fragments it has limited capacity to derive much needed but more sophisticated information, such as abundance. Here we describe the isolation and sequencing of single fish cells isolated from a 3 million litre mesocosm. We term these cells environmental metazoan cells (emCells), which are whole cells shed from macro-organisms into aquatic environments. emCells derived from dissociated tissues and collected from the mesocosm were isolated using flow cytometry with fluorescent activated cell sorting (FACS) using a custom molecular probe targeting bony fishes and a novel multi-factor gating strategy. We verified the presence of intact cells by amplifying and sequencing both mitochondrial and nuclear DNA from individual and pools of cells (1-1000 cells). Optimal sequencing success was achieved with pools of 5 cells (68.8%), however 14.6% of single cells yielded mtDNA sequences and 10.4% yielded nuclear sequences, while 12.5% yielded both mtDNA and nuclear sequences. Isolation and analysis of emCells promises to take eDNA analysis beyond the presence/absence of taxa. Coupling our new approach with genetic mark-recapture methods has the potential to estimate fish abundance from a bucket of water.


Biography:

Biography to come

Date

Mar 21 - 23 2022
Expired!