Protocol for flow cytometry-assisted single-nucleus RNA sequencing of human and mouse adipose tissue with sample multiplexing

Autor: Anton Gulko, Adam Essene, Dylan Matthew Belmont-Rausch, Molly Veregge, Deepti Pant, Danielle Tenen, Benedicte Schultz Kapel, Margo P. Emont, Tune H. Pers, Evan D. Rosen, Linus T. Tsai
Jazyk: angličtina
Rok vydání: 2024
Předmět:
Zdroj: STAR Protocols, Vol 5, Iss 1, Pp 102893- (2024)
Druh dokumentu: article
ISSN: 2666-1667
DOI: 10.1016/j.xpro.2024.102893
Popis: Summary: Adipocyte size and fragility and commercial kit costs impose significant limitations on single-cell RNA sequencing of adipose tissue. Accordingly, we developed a workflow to isolate and sample-barcode nuclei from individual adipose tissue samples, integrating flow cytometry for quality control, counting, and precise nuclei pooling for direct loading onto the popular 10× Chromium controller. This approach can eliminate batch confounding, and significantly reduces poor-quality nuclei, ambient RNA contamination, and droplet loading-associated reagent waste, resulting in pronounced improvements in information content and cost efficiency. : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
Databáze: Directory of Open Access Journals