STAR Protocols (Dec 2022)
Direct isolation of single cells from living brains of Drosophila melanogaster without dissociation for transcriptome analysis
Abstract
Summary: Here, we describe a protocol to remove single identified cells directly from Drosophila living brains and analyze their transcriptome. We detail the steps to harvest fluorescent cells using a capillary under epifluorescence and transmitted light to avoid contamination. We then outline the procedure to obtain the transcriptome by reverse transcription and amplification. The process from cell harvesting to the initiation of reverse transcription only takes 2 min, thus avoiding transcriptional activation of cell damage response or cell death genes.For complete details on the use and execution of this protocol, please refer to Barros and Bossing (2021), Bossing et al. (2012), Gil-Ranedo et al. (2019), and Liu and Bossing (2016). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
