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| Status |
Public on Jan 16, 2022 |
| Title |
Single-nuclei isoform RNA sequencing reveals combination patterns of transcript elements across human brain cell types |
| Organisms |
Homo sapiens; Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Single-nuclei RNA-Seq is widely employed to investigate cell types, especially of human brain and frozen samples. In contrast to single-cell approaches, many single-nuclei reads are purely intronic. Here, using microfluidics, PCR-based artifact removal, target enrichment, and long-read sequencing, we developed single-nuclei isoform RNA-sequencing (‘SnISOr-Seq’), and applied it to human adult frontal cortex. SnISOr-Seq dramatically increased the fraction of informative reads. We found that exons associated with autism exhibit coordinated and highly cell-type specific inclusion. We discovered two distinct combination patterns: first, those distinguishing neural cell types, enriched in TSS-exon, exon-polyA-site, and non-adjacent exon pairs. Second, those with multiple configurations within one cell type, enriched in adjacent exon pairs. Furthermore, adjacent exons are predominantly mutually-associated, while distant exons are frequently mutually-exclusive. Finally, we observed that human-specific exons are almost as tightly coordinated as conserved exons. SnISOr-Seq enables single-nuclei long-read isoform analysis in human brain, and in any frozen or hard-to-dissociate sample.
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| Overall design |
10X single-nuclear RNA sequencing done on two replicates of human Frontal Cortex samples. PacBio done on the same cDNA, after linear PCR. 10X single-cell RNA sequencing followed by Oxford Nanopore long read sequencing as a control sample. FACS sorted nuclei obtained from Human prefrontal cortex samples for bulk RNA sequencing on Illumina, also used as control.
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| Contributor(s) |
Hardwick S, Hu W, Joglekar A, Tilgner H |
| Citation(s) |
35256815, 35301264, 36593406 |
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| Submission date |
Jun 14, 2021 |
| Last update date |
Sep 12, 2023 |
| Contact name |
Anoushka Joglekar |
| E-mail(s) |
anj2026@med.cornell.edu
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| Organization name |
Weill Cornell Medicine
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| Department |
Dept. of Neurogenetics
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| Lab |
Tilgner Lab
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| Street address |
413 E 69th St
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| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10021 |
| Country |
USA |
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| Platforms (5)
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| GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
| GPL26167 |
PromethION (Homo sapiens) |
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| Samples (12)
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| Relations |
| BioProject |
PRJNA737508 |
| SRA |
SRP324021 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE178175_RAW.tar |
312.0 Mb |
(http)(custom) |
TAR (of GFF, MTX, TAB, TSV) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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