Collection Citation: Heidi Monroe, Nayra Cardenes, Oliver Eickelberg, Melanie Königshoff, Melanie Königshoff, Robert Lafyatis 2024. Single nuclei RNA sequencing (snRNA-seq) of frozen human lung tissue and hPCLS. protocols.io https://dx.doi.org/10.17504/protocols.io.36wgqndb5gk5/v1
License: This is an open access collection distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
TriState SenNET (Lung and Heart) Tissue Map and Atlas consortium - NIA
Grant ID: U54AG075931
Abstract
Single-cell RNA sequencing (scRNA-seq) has become an essential tool for delineating cellular diversity in normal tissues and alterations in disease states. This technique requires the dissociation of tissue specimens into cell suspensions. However, the isolation of intact cells can be challenging due to factors such as fragility, large size and tight interconnections. Additionally, single-nuclei isolation can be performed on frozen tissue, enabling the analysis of biobanked samples in a single batch. This protocol for single-nuclei RNA sequencing (snRNA-seq) provides an alternative approach to scRNA-seq, overcoming these limitations to generate high-quality transcriptomic data.
The analysis of gene expression at the cellular level has proven to be a powerful tool for understanding various aspects of lung biology and disease, particularly the process of lung aging. Aging affects different cell types within the human lung heterogeneously, leading to a range of associated changes in their function. Examining the patterns of gene expression in the numerous lung cell types provides insights into the aging and disease processes that contribute to cellular dysfunction. By analyzing these changes at the single-cell level, we can delineate the complex cellular diversity in the human lung and track alterations in molecular pathways involved in the dynamic process of lung aging. Understanding these gene expression patterns will offer opportunities for timely interventions and the identification of biomarker for early prognosis and personalized treatment therapies.
This protocol collection describes the process of single nuclei RNA sequencing from nuclei isolation from frozen tissue (whole, or from PCLS), to barcoding, library construction and sequencing.
The protocol content here is for informational purposes only and does not constitute legal, medical, clinical, or safety advice, or otherwise; content added to protocols.io is not peer reviewed and may not have undergone a formal approval of any kind. Information presented in this protocol should not substitute for independent professional judgment, advice, diagnosis, or treatment. Any action you take or refrain from taking using or relying upon the information presented here is strictly at your own risk. You agree that neither the Company nor any of the authors, contributors, administrators, or anyone else associated with protocols.io, can be held responsible for your use of the information contained in or linked to this protocol or any of our Sites/Apps and Services.
