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

Autor:	Gulko A; Division of Endocrinology, Diabetes, and Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA; Boston Nutrition and Obesity Research Center/Boston Area Diabetes and Endocrinology Research Center Functional Genomics and Bioinformatics Core, Boston, MA, USA., Essene A; Division of Endocrinology, Diabetes, and Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA; Boston Nutrition and Obesity Research Center/Boston Area Diabetes and Endocrinology Research Center Functional Genomics and Bioinformatics Core, Boston, MA, USA., Belmont-Rausch DM; Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, 2200 Copenhagen, Denmark., Veregge M; Division of Endocrinology, Diabetes, and Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA; Boston Nutrition and Obesity Research Center/Boston Area Diabetes and Endocrinology Research Center Functional Genomics and Bioinformatics Core, Boston, MA, USA., Pant D; Division of Endocrinology, Diabetes, and Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA; Boston Nutrition and Obesity Research Center/Boston Area Diabetes and Endocrinology Research Center Functional Genomics and Bioinformatics Core, Boston, MA, USA., Tenen D; Division of Endocrinology, Diabetes, and Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA; Boston Nutrition and Obesity Research Center/Boston Area Diabetes and Endocrinology Research Center Functional Genomics and Bioinformatics Core, Boston, MA, USA., Kapel BS; Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, 2200 Copenhagen, Denmark., Emont MP; Division of Endocrinology, Diabetes, and Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA., Pers TH; Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, 2200 Copenhagen, Denmark., Rosen ED; Division of Endocrinology, Diabetes, and Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA; Boston Nutrition and Obesity Research Center/Boston Area Diabetes and Endocrinology Research Center Functional Genomics and Bioinformatics Core, Boston, MA, USA., Tsai LT; Division of Endocrinology, Diabetes, and Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA; Boston Nutrition and Obesity Research Center/Boston Area Diabetes and Endocrinology Research Center Functional Genomics and Bioinformatics Core, Boston, MA, USA. Electronic address: ltsai@bidmc.harvard.edu.
Jazyk:	angličtina
Zdroj:	STAR protocols [STAR Protoc] 2024 Mar 15; Vol. 5 (1), pp. 102893. Date of Electronic Publication: 2024 Feb 27.
DOI:	10.1016/j.xpro.2024.102893
Abstrakt:	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. Competing Interests: Declaration of interests The authors declare no competing interests. (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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