Optimized assay for transposase-accessible chromatin by sequencing (ATAC-seq) library preparation from adult Drosophila melanogaster neurons
Year: 2022
Type: article
Abstract: Assay for transposase-accessible chromatin by sequencing (ATAC-seq) is rapidly becoming the assay of choice to investigate chromatin-mediated gene regulation, largely because of low input requirements, a fast workflow, and the ability to interrogate the entire genome in an untargeted manner. Many studies using ATAC-seq use mammalian or human-derived tissues, and established protocols work well in these systems. However, ATAC-seq is not yet widely used in Drosophila. Vinegar flies present several advantages over mammalian systems that make them an excellent model for ATAC-seq studies, including abundant genetic tools that allow straightforward targeting, transgene expression, and genetic manipulation that are not available in mammalian models. Because current ATAC-seq protocols are not optimized to use flies, we developed an optimized workflow that accounts for several complicating factors present in Drosophila. We examined parameters affecting nuclei isolation, including input size, freezing time, washing, and possible confounds from retinal pigments. Then, we optimized the enzymatic steps of library construction to account for the smaller Drosophila genome size. Finally, we used our optimized protocol to generate ATAC-seq libraries that meet ENCODE quality metrics. Our optimized protocol enables extensive ATAC-seq experiments in Drosophila, thereby leveraging the advantages of this powerful model system to understand chromatin-mediated gene regulation.
Source: Scientific Reports
Institutions: University of Utah, VA Salt Lake City Healthcare System
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $2,190
Awards: Chromatin remodeling in GABA neurons contributes to alcohol use disorder, Mechanisms of alcohol-induced plasticitey mediated by Arf6, G3646000930, ATAC-ing dopaminergic cell identity with single-cell resolution, G6423704564, Regulation of WNK signaling by potassium and Mo25: structure, function and physiology, Control of Alcohol Responses by Actin-Regulating Genes, Cancer Center Support Grant
OpenAlex ID: W4223487635