Details

Autor(en) / Beteiligte

• Gupta, Ishaan
• Collier, Paul G
• Haase, Bettina
• Mahfouz, Ahmed
• Joglekar, Anoushka
• Floyd, Taylor
• Koopmans, Frank
• Barres, Ben
• Smit, August B
• Sloan, Steven A
• Luo, Wenjie
• Fedrigo, Olivier
• Ross, M Elizabeth
• Tilgner, Hagen U

Titel

Single-cell isoform RNA sequencing characterizes isoforms in thousands of cerebellar cells

Ist Teil von

• Nature biotechnology, 2018-12, Vol.36 (12), p.1197-1202

Ort / Verlag

New York: Nature Publishing Group US

Erscheinungsjahr

2018

Beschreibungen/Notizen

• Full-length spliced RNA isoforms are identified in thousands of single cerebellar cells. Full-length RNA sequencing (RNA-Seq) has been applied to bulk tissue, cell lines and sorted cells to characterize transcriptomes 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , but applying this technology to single cells has proven to be difficult, with less than ten single-cell transcriptomes having been analyzed thus far 12 , 13 . Although single splicing events have been described for ≤200 single cells with statistical confidence 14 , 15 , full-length mRNA analyses for hundreds of cells have not been reported. Single-cell short-read 3′ sequencing enables the identification of cellular subtypes 16 , 17 , 18 , 19 , 20 , 21 , but full-length mRNA isoforms for these cell types cannot be profiled. We developed a method that starts with bulk tissue and identifies single-cell types and their full-length RNA isoforms without fluorescence-activated cell sorting. Using single-cell isoform RNA-Seq (ScISOr-Seq), we identified RNA isoforms in neurons, astrocytes, microglia, and cell subtypes such as Purkinje and Granule cells, and cell-type-specific combination patterns of distant splice sites 6 , 7 , 8 , 9 , 22 , 23 . We used ScISOr-Seq to improve genome annotation in mouse Gencode version 10 by determining the cell-type-specific expression of 18,173 known and 16,872 novel isoforms.