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Nat Protoc. 2018 Nov;13(11):2501-2534. doi: 10.1038/s41596-018-0045-2.

Barcoded solid-phase RNA capture for Spatial Transcriptomics profiling in mammalian tissue sections.

Author information

1
Science for Life Laboratory, Department of Gene Technology, KTH Royal Institute of Technology, Stockholm, Sweden.
2
Hubrecht Institute-KNAW (Royal Netherlands Academy of Arts and Sciences) and University Medical Center Utrecht, Cancer Genomics Netherlands, Utrecht, The Netherlands.
3
Science for Life Laboratory, Department of Gene Technology, KTH Royal Institute of Technology, Stockholm, Sweden. patrik.stahl@scilifelab.se.
4
Broad Institute of Harvard and Massachusetts Institute of Technology, Cambridge, MA, USA.
5
Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden.
6
Science for Life Laboratory, Department of Gene Technology, KTH Royal Institute of Technology, Stockholm, Sweden. joakim.lundeberg@scilifelab.se.

Abstract

Spatial resolution of gene expression enables gene expression events to be pinpointed to a specific location in biological tissue. Spatially resolved gene expression in tissue sections is traditionally analyzed using immunohistochemistry (IHC) or in situ hybridization (ISH). These technologies are invaluable tools for pathologists and molecular biologists; however, their throughput is limited to the analysis of only a few genes at a time. Recent advances in RNA sequencing (RNA-seq) have made it possible to obtain unbiased high-throughput gene expression data in bulk. Spatial Transcriptomics combines the benefits of traditional spatially resolved technologies with the massive throughput of RNA-seq. Here, we present a protocol describing how to apply the Spatial Transcriptomics technology to mammalian tissue. This protocol combines histological staining and spatially resolved RNA-seq data from intact tissue sections. Once suitable tissue-specific conditions have been established, library construction and sequencing can be completed in ~5-6 d. Data processing takes a few hours, with the exact timing dependent on the sequencing depth. Our method requires no special instruments and can be performed in any laboratory with access to a cryostat, microscope and next-generation sequencing.

PMID:
30353172
DOI:
10.1038/s41596-018-0045-2
[Indexed for MEDLINE]

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