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Series GSE156988 Query DataSets for GSE156988
Status Public on Aug 28, 2020
Title FD-seq: Droplet-based RNA sequencing of fixed single cells
Organisms Homo sapiens; Mus musculus
Experiment type Expression profiling by high throughput sequencing
Summary Current high-throughput single-cell RNA sequencing (scRNA-seq) methods are incompatible with paraformaldehyde (PFA) fixation, a common cell/tissue preservation and viral inactivation technique, which has prevented transcriptomic analysis of rare cells that require protein staining and enrichment. Here we present FD-seq, a method for high-throughput RNA sequencing of PFA-fixed, stained and FACS-sorted single cells. We used FD-seq to address two important questions in virology. First, by analyzing a rare population of cells supporting herpesvirus reactivation, we identified TMEM119 to mediate reactivation of Kaposi’s sarcoma-associated herpesvirus (KSHV), a tumor virus. Second, we studied the innate immune activation in cells infected with the coronavirus OC43, which causes the common cold and also serves as a safer model pathogen for SARS-CoV-2 studies. We found that pro-inflammatory cytokine induction, which is a major contributor to the severe pathogenicity in SARS-CoV-2-infected individuals, is primarily driven by uninfected or lowly infected bystander cells that are exposed to the virus but fail to express high level of viral genes. FD-seq is a simple method that is suitable for characterizing the transcriptome of rare cell populations of interest, and for studying high-containment biological samples such as SARS-CoV-2-infected cells after PFA inactivation.
 
Overall design KSHV reactivation is chemically induced, and the mRNA from single reactivated and non-reactivated cells are sequenced. Bulk RNA-seq was performed to identify the mechanism of TMEM119-induced reactivation. OC43 infected and mock infected cells were sequenced to identify subpopulation of bystander cells. The performance of FD-seq was also compared to the standard Drop-seq method for methanol-fixed cells.
 
Contributor(s) Phan HV
Citation(s) 32995793, 34561439
Submission date Aug 27, 2020
Last update date Oct 27, 2021
Contact name Hoang Van Phan
E-mail(s) [email protected]
Organization name The University of Chicago
Department Priztker School of Molecular Engineering
Street address 5640 S Ellis Ave
City Chicago
State/province Illinois
ZIP/Postal code 60637
Country USA
 
Platforms (2)
GPL21697 NextSeq 550 (Homo sapiens)
GPL24625 NextSeq 550 (Homo sapiens; Mus musculus)
Samples (31)
GSM4749729 Live species mixing
GSM4749730 Fixed species mixing
GSM4749731 KSHV K8.1 positive
Relations
BioProject PRJNA659731
SRA SRP279053

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE156988_RAW.tar 27.4 Mb (http)(custom) TAR (of TXT)
GSE156988_counts_KSHV.txt.gz 3.4 Mb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file
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