Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Highly multiplexed mRNA quantitation with CRISPR-Cas13

ABSTRACT: RNA quantitation tools are often either high-throughput or cost-effective, but rarely are they both. Existing methods can profile the transcriptome at great expense or are limited to quantifying a handful of genes by labor constraints. A technique that permits more throughput at a reduced cost could enable multi-gene kinetic studies, gene regulatory network analysis, and combinatorial genetic screens. Here, we introduce quantitative Combinatorial Arrayed Reactions for Multiplexed Evaluation of Nucleic acids (qCARMEN): an RNA quantitation technique which leverages the programmable RNA-targeting capabilities of CRISPR-Cas13 to address this challenge by quantifying over 4,500 gene-sample pairs in a single experiment. Using qCARMEN, we studied the response profiles of interferon-stimulated genes (ISGs) during interferon (IFN) stimulation and flavivirus infection. Additionally, we observed isoform switching kinetics during epithelial-mesenchymal transition. qCARMEN is a simple and inexpensive technique that greatly enhances the scalability of RNA quantitation for novel applications with performance similar to gold-standard methods.

ORGANISM(S): Homo sapiens

PROVIDER: GSE289345 | GEO | 2025/10/31

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Highly multiplexed mRNA quantitation with CRISPR-Cas13

Project description:Highly multiplexed mRNA quantitation with CRISPR-Cas13

| PRJNA1222449 | ENA

Analysis of RNA quantitation in formaldehyde-fixed cells

Project description:Fluorescence-activated cell sorting (FACS) is a sensitive and valuable technique to characterize cellular subpopulations and great advances have been made using this approach. Cells are often fixed with formaldehyde prior to the sorting process to preserve cell morphology and maintain the expression of surface molecules, as well as to ensure safety in the sorting of infected cells. It is widely recognized that formaldehyde fixation alters RNA and DNA structure and integrity, thus analyzing gene expression in these cells has been difficult. We therefore examined the effects of formaldehyde fixation on the stability and quantitation of nucleic acids in cell lines, primary leukocytes and also cells isolated from SIV-infected pigtailed macaques. We developed a method to extract RNA from fixed cells that yielded the same amount of RNA as our common method of RNA isolation from fresh cells. Quantitation of RNA by RT-qPCR in fixed cells was not always comparable with that in unfixed cells. In comparison, when RNA was measured by the probe-based NanoString system, there was no significant difference in RNA quantitation. In addition, we demonstrated that quantitation of proviral DNA in fixed cells by qPCR is comparable to that in unfixed cells when normalized by a single-copy cellular gene. These results provide a systematic procedure to quantitate gene expression in cells that have been fixed with formaldehyde and sorted by FACS.

2013-09-09 | GSE50690 | GEO

An integrated 5'- and 3'-end RNA sequencing platform for high-throughput transcriptome quantitation and annotation

Project description:RNA-Seq provides an effective way to annotate and measure the transcriptome, but the combination of cost, analysis and end-mapping challenges has limited its adoption for high-throughput quantitation and annotation. Here, we present an integrated experimental and computational suite for transcriptome annotation and quantitation based on the sequencing of mRNA ends. It consists of: a novel, simple, one-step, strategy 5' RNA-Seq; an optimized library construction method for strand-specific 3' RNA-seq that reduces costs and time; and a complete computational analysis toolkit. We demonstrate the power and versatility of our approach to study the transcriptome of LPS stimulation in mouse dendritic cells, promoter structure of the TCRb locus in mouse T cells, and gene expression in circadian cycles in Drosophila. Our platform provides a comprehensive solution for high-throughput, cost-effective transcriptome quantification and end annotation.

2013-06-01 | GSE47207 | GEO

Intrinsic RNA targeting constrains the utility of CRISPR-Cas13 systems [Screen]

Project description:CRISPR-Cas13 systems have been adapted as versatile toolkits for RNA-related applications. Here we systematically evaluate the performance of several prominent Cas13 family effectors (Cas13a, Cas13b and Cas13d) under lentiviral vectors and reveal surprisingly differential defects and characteristics of these systems. Using RNA immunoprecipitation sequencing, transcriptome profiling, biochemistry analysis and high-throughput CRISPR-Cas13 screening approaches, we determine that each Cas13 system has its intrinsic RNA targets in mammalian cells. Viral process-related host genes can be targeted by Cas13 and affect the production of fertile lentiviral particles, thereby restricting the utility of lentiviral Cas13 systems. Multiple RNase activities of Cas13 are involved in endogenous RNA targeting. Unlike target-induced collateral effect, intrinsic RNA targeting can be specific, target-independent and dynamically tuned by varied states of Cas13 nucleases. Our work not only provides guidance to appropriately utilize lentiviral Cas13 systems, but also raises cautions about intrinsic RNA targeting during Cas13-based basic and therapeutic applications.

2023-10-09 | GSE202841 | GEO

Intrinsic RNA targeting constrains the utility of CRISPR-Cas13 systems [RNA-seq]

2023-10-09 | GSE202838 | GEO

Intrinsic RNA targeting constrains the utility of CRISPR-Cas13 systems [RIP-seq]

2023-10-09 | GSE202898 | GEO

An integrated 5'- and 3'-end RNA sequencing platform for high-throughput transcriptome quantitation and annotation

2013-06-01 | E-GEOD-47207 | biostudies-arrayexpress

Transcriptome changes upon Cas13-based knockdown

Project description:We used RNA-seq to detect genome-wide gene expression change when a reporter RNA was targeted by Cas13

2023-02-24 | GSE155134 | GEO

Analysis of RNA quantitation in formaldehyde-fixed cells

2013-09-09 | E-GEOD-50690 | biostudies-arrayexpress

Efficient combinatorial targeting of RNA transcripts in single cells with Cas13 RNA Perturb-seq

Project description:Pooled CRISPR screens coupled with single-cell RNA-sequencing have enabled systematic interrogation of gene function and regulatory networks. Here, we introduce Cas13 RNA Perturb-seq (CaRPool-seq) which leverages the RNA-targeting CRISPR/Cas13d system and enables efficient combinatorial perturbations alongside multimodal single-cell profiling. CaRPool-seq encodes multiple perturbations on a cleavable array which is associated with a detectable barcode sequence, allowing for the simultaneous targeting of multiple genes. We compared CaRPool-seq to existing Cas9-based methods, highlighting its unique strength to efficiently profile combinatorially perturbed cells. Finally, we apply CaRPool-seq to perform multiplexed combinatorial perturbations of myeloid differentiation regulators in an acute myeloid leukemia (AML) model system and identify extensive interactions between different chromatin regulators that can enhance or suppress AML differentiation phenotypes.

2022-09-29 | GSE213957 | GEO

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data