Back to Search
Start Over
HyPR-seq: Single-cell quantification of chosen RNAs via hybridization and sequencing of DNA probes.
- Source :
-
Proceedings of the National Academy of Sciences of the United States of America [Proc Natl Acad Sci U S A] 2020 Dec 29; Vol. 117 (52), pp. 33404-33413. Date of Electronic Publication: 2020 Dec 21. - Publication Year :
- 2020
-
Abstract
- Single-cell quantification of RNAs is important for understanding cellular heterogeneity and gene regulation, yet current approaches suffer from low sensitivity for individual transcripts, limiting their utility for many applications. Here we present Hybridization of Probes to RNA for sequencing (HyPR-seq), a method to sensitively quantify the expression of hundreds of chosen genes in single cells. HyPR-seq involves hybridizing DNA probes to RNA, distributing cells into nanoliter droplets, amplifying the probes with PCR, and sequencing the amplicons to quantify the expression of chosen genes. HyPR-seq achieves high sensitivity for individual transcripts, detects nonpolyadenylated and low-abundance transcripts, and can profile more than 100,000 single cells. We demonstrate how HyPR-seq can profile the effects of CRISPR perturbations in pooled screens, detect time-resolved changes in gene expression via measurements of gene introns, and detect rare transcripts and quantify cell-type frequencies in tissue using low-abundance marker genes. By directing sequencing power to genes of interest and sensitively quantifying individual transcripts, HyPR-seq reduces costs by up to 100-fold compared to whole-transcriptome single-cell RNA-sequencing, making HyPR-seq a powerful method for targeted RNA profiling in single cells.<br />Competing Interests: Competing interest statement: J.L.M., V.S., S.G.R., F.C., and J.M.E. are inventors on patent applications filed by the Broad Institute related to this work (62/676,069 and 62/780,889). E.S.L. serves on the Board of Directors for Codiak BioSciences and Neon Therapeutics, and serves on the Scientific Advisory Board of F-Prime Capital Partners and Third Rock Ventures; he is also affiliated with several nonprofit organizations, including serving on the Board of Directors of the Innocence Project, Count Me In, and Biden Cancer Initiative, and the Board of Trustees for the Parker Institute for Cancer Immunotherapy. E.S.L. has served and continues to serve on various federal advisory committees.
- Subjects :
- Animals
CRISPR-Cas Systems genetics
Gene Expression
Humans
Introns genetics
K562 Cells
Kidney cytology
Mice
Polyadenylation
RNA, Messenger genetics
RNA, Messenger metabolism
THP-1 Cells
Time Factors
DNA Probes genetics
High-Throughput Nucleotide Sequencing methods
Nucleic Acid Hybridization
RNA metabolism
Single-Cell Analysis
Subjects
Details
- Language :
- English
- ISSN :
- 1091-6490
- Volume :
- 117
- Issue :
- 52
- Database :
- MEDLINE
- Journal :
- Proceedings of the National Academy of Sciences of the United States of America
- Publication Type :
- Academic Journal
- Accession number :
- 33376219
- Full Text :
- https://doi.org/10.1073/pnas.2010738117