TECHNOLOGICAL AND COMPUTATIONAL ADVANCEMENTS IN DROSOPHILA SINGLE-CELL OMICS

Regulatory programs encoded in the genome are required for the correct differentiation of cell types into their final fates. These programs are formed by cis-regulatory regions throughout the genome that are activated/repressed by the binding of transcription factors, causing expression of a specific repertoire of genes; it is this combination of genes being expressed that defines the cell type and function of a cell. The main aim of this thesis was to implement and develop advances in technologies and software for the analysis of both genome regulation and single cell omics. To achieve this goal, we implemented the novel technique, ATAC-seq in Drosophila, built and implemented systems for high throughput, droplet-based single cell RNA-seq to profile an entire tissue, and developed software solutions for the analysis and visualisation of the data generated by these techniques. At the time a just-published technique, ATAC-seq provided advantages over similar techniques such as FAIRE-seq and DNAse-seq, particularly its ability to work well on small amount of input material, but it had not yet been applied to Drosophila tissues. After optimising the ATAC-seq protocol to work on Drosophila tissues, comparing the three techniques we found that ATAC-seq was able to work on 10-20x less input material with a marked increase in signal-to-noise ratio. We next began working on implementing single cell technologies in Drosophila with the aim of increasing resolution and providing the first single cell atlas of the adult fly brain. We used the 10X genomics platform to profile >125,000 single cells from the adult Drosophila brain across a range of ages. Using this data we were able to annotate over 80 unique cell types across the entire brain as well as use cutting-edge bioinformatics techniques to identify regulatory networks across the entire dataset. To make this, and future single cell data readily available to a wide audience, we developed the webtool SCope for easy and fast visualisation of single cell datasets. Finally, building on our experiences from generating the fly brain atlas, we developed a Nextflow-powered pipeline for the analysis of single cell omics data. Alongside this, we further developed SCope to provide a collaborative annotation system that allows multiple users to work and annotate clusters/cell types in a single dataset based on controlled vocabularies. Combined, the work presented in this thesis has helped progress the field of drosophila omics, the adapted ATAC-seq protocol has been used by many other groups to answer their own research questions. Moreover, with the fly brain atlas as a catalyst, both SCope and vsn-pipelines have become integral parts in the Fly Cell Atlas, a multi-national project to profile all tissues in the adult fly, and we hope they find use in many other such large-scale projects in the future.

Jaar van publicatie:2021

Toegankelijkheid:Open