Hulselmans, G. J. | bioRxiv Science

bioRxiv2023

Spatial transcriptomics in adult Drosophila reveals new cell types in the brain and identifies subcellular mRNA patterns in muscles

Recently, we have achieved a significant milestone with the creation of the Fly Cell Atlas. This single-nuclei atlas encompasses the entire fly, covering the entire head and body, in addition to all major organs. This atlas catalogs many hundreds of cell types, of which we annotated 250. Thus, a large number of clusters remain to be fully characterized, in particular in the brain. Furthermore, by applying single-nuclei sequencing, all information about the spatial location of the cells in the body and of about possible subcellular localization of the mRNAs within these cells is lost. Spatial transcriptomics promises to tackle these issues. In a proof-of-concept study, we have here applied spatial transcriptomics using a selected gene panel to pinpoint the locations of 150 mRNA species in the adult fly. This enabled us to map unknown clusters identified in the Fly Cell Atlas to their spatial locations in the fly brain. Additionally, spatial transcriptomics discovered interesting principles of mRNA localization and transcriptional diversity within the large and crowded muscle cells that may spark future mechanistic investigations. Furthermore, we present a set of computational tools that will allow for easier integration of spatial transcriptomics and single-cell datasets.

developmental biology↗

bioRxiv2021

HyDrop: droplet-based scATAC-seq and scRNA-seq using dissolvable hydrogel beads

Single-cell RNA-seq and single-cell ATAC-seq technologies are being used extensively to create cell type atlases for a wide range of organisms, tissues, and disease processes. To increase the scale of these atlases, lower the cost, and allow for more specialized multi-ome assays, custom droplet microfluidics may provide complementary solutions to commercial setups. We developed HyDrop, a flexible and generic droplet microfluidic platform encompassing three protocols. The first protocol involves creating dissolvable hydrogel beads with custom oligos that can be released in the droplets. In the second protocol, we demonstrate the use of these beads for HyDrop-ATAC, a low-cost non-commercial scATAC-seq protocol in droplets. After validating HyDrop-ATAC, we applied it to flash-frozen mouse cortex and generated 8,502 high-quality single-cell chromatin accessibility profiles in a single run. In the third protocol, we adapt both the reaction chemistry and the capture sequence of the barcoded hydrogel bead to capture mRNA, and demonstrate a significant improvement in throughput and sensitivity compared to previous open-source droplet-based scRNA-seq assays (Drop-seq and inDrop). Similarly, we applied HyDrop-RNA to flash-frozen mouse cortex and generated 9,508 single-cell transcriptomes closely matching reference single-cell gene expression data. Finally, we leveraged HyDrop-RNAs high capture rate to analyse a small population of FAC-sorted neurons from the Drosophila brain, confirming the protocols applicability to low-input samples and small cells. HyDrop is currently capable of generating single-cell data in high throughput and at a reduced cost compared to commercial methods, and we envision that HyDrop can be further developed to be compatible with novel (multi-) omics protocols.

genomics↗