Fluorescent in situ sequencing (FISSEQ) of RNA for gene expression profiling in intact cells and tissues.
IF: 17.021
Cited by: 268


RNA-sequencing (RNA-seq) measures the quantitative change in gene expression over the whole transcriptome, but it lacks spatial context. In contrast, in situ hybridization provides the location of gene expression, but only for a small number of genes. Here we detail a protocol for genome-wide profiling of gene expression in situ in fixed cells and tissues, in which RNA is converted into cross-linked cDNA amplicons and sequenced manually on a confocal microscope. Unlike traditional RNA-seq, our method enriches for context-specific transcripts over housekeeping and/or structural RNA, and it preserves the tissue architecture for RNA localization studies. Our protocol is written for researchers experienced in cell microscopy with minimal computing skills. Library construction and sequencing can be completed within 14 d, with image analysis requiring an additional 2 d.


Gene Expression

MeSH terms

DNA, Complementary
Gene Expression Profiling
Microscopy, Confocal
RNA, Messenger
Sequence Analysis, RNA


Lee, Je Hyuk
Daugharthy, Evan R
Scheiman, Jonathan
Kalhor, Reza
Ferrante, Thomas C
Terry, Richard
Turczyk, Brian M
Yang, Joyce L
Lee, Ho Suk
Aach, John
Zhang, Kun
Church, George M

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