Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox

ABSTRACT: Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox

PROVIDER: PRJNA1011332 | ENA |

REPOSITORIES: ENA

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	SRR25841400.fastq.gz	Fastqsanger.gz
	SRR25841401.fastq.gz	Fastqsanger.gz
	SRR25848706.fastq.gz	Fastqsanger.gz
	SRR25848707.fastq.gz	Fastqsanger.gz
	SRR25848708.fastq.gz	Fastqsanger.gz

Items per page:

1 - 5 of 36

Similar Datasets

Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox [Screen]

Project description:Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox [Screen]

| PRJNA1011336 | ENA

Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox [RNA-seq]

Project description:Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox [RNA-seq]

| PRJNA1011334 | ENA

Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox [LR-RNA-Seq]

Project description:Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox [LR-RNA-Seq]

| PRJNA1011333 | ENA

Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox [Screen]

Project description:Alternative splicing is an essential mechanism for diversifying proteins, with different mature RNA isoforms producing different proteins with potentially distinct functions. Two major challenges in characterizing the cellular function of isoforms are the lack of both experimental methods to specifically and efficiently modulate isoform expression and computational tools for complex experimental design. To address these gaps, we developed and methodically tested a strategy which pairs the RNA-targeting CRISPR/Cas13d system with guide RNAs (gRNAs) that overlap exon-exon junctions in the mature RNA. We perform a high-throughput essentiality screen, numerous qPCR assays, and PacBio long read sequencing to affirm that our strategy works to specifically target the unique junctions of isoforms to robustly knockdown their RNA. In parallel, we provide computational tools for experimental design and screen analysis. Considering all possible splice junctions in current GENCODE annotations and our predictions of gRNA efficacy, we estimate that this strategy can target 94% of all human isoforms, including 29,238 protein-coding and 9,638 lncRNA isoforms that are specifically targetable with a gRNA spanning a unique splice junction.

2023-09-13 | GSE242106 | GEO

Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox [RNA-seq]

2023-09-13 | GSE242105 | GEO

Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox [LR-RNA-Seq]

2023-09-13 | GSE242104 | GEO

Cas13d-mediated isoform-specific RNA knockdown with a unified computational and experimental toolbox

Project description:This SuperSeries is composed of the SubSeries listed below.

2023-09-13 | GSE242107 | GEO

Cas13d experimental viral genome-wide screen identifies regions of RNA vulnerability in SARScov2

Project description:We performed an experimental Cas13d-SARScov2 genome-wide screen to identify gRNAs that would allow Cas13d to degrade the viral RNA. We built mCherry reporter plasmids that express mCherry with a 3kb 3'UTR deriving from the SARScov2 genome. In total we designed 11 reporters covering the entire plus strand of the viral genome and 11 other reporters covering the entire minus strand. Each of the 22 mCherry reporter plasmids carries a U6 expression cassette containing a Cas13d gRNA that targets the 3'UTR of the mCherry reporter. Each reporter is represented by a pool of reporters each containing a different gRNA that targets mCherry 3'UTR for a total average of ~300 gRNA per 3'UTR. The entire pool of 22 reporters, each with a pool of ~300 different gRNAs constitutes a comprehensive set ~6,500 reporters (~ 6,500 different gRNAs) that allowed us to interrogate the entire SARScov2 plus and minus strand viral RNA for regions of vulnerability and targetability. In order to specifically interrogate Cas13d activity an remove the biases that would be introduced in the reporter expression by the presence of a large 3kb 3'UTR we used a case (presence of Cas13d) control (absence of Cas13d) design. Briefly, the ~6,500 reporters were lentiviral transduced in RKO cells, the cells were split in 2 populations, 1 population was transduced with Cas13d and the other serving as control did not. The population expressing Cas13d was FACS sorted in low mCherry (efficient gRNAs) and high mCherry (un-efficient gRNAs) in 2 biological replicates and the genomic DNA of these populations was extracted, gRNAs were PCR amplified and sequenced. For the population that did not express Cas13d, a low mCherry, one high mCherry and unsorted population were sequenced as control libraries.

2023-10-29 | E-MTAB-11111 | biostudies-arrayexpress

Optimized RNA-targeting CRISPR/Cas13d technology outperforms shRNA in identifying functional circRNAs

Project description:Circular RNAs (circRNAs) are widely expressed, but their functions remain largely unknown. To study circRNAs in a high-throughput manner, short hairpin RNA (shRNA) screens1 have recently been used to deplete circRNAs by targeting their unique back-splicing junction (BSJ) sites. Here, we report frequent discrepancies between shRNA-mediated circRNA knockdown efficiency and the corresponding biological effect, raising pressing concerns about the robustness of shRNA screening for circRNA functional characterization. To address this issue, we leveraged the CRISPR/Cas13d system2 for functional study of circRNAs by optimizing the strategy for designing single guide RNAs to deplete circRNAs. We then performed shRNA and CRISPR/Cas13d parallel screenings and demonstrated that shRNA-mediated circRNA screening yielded a high rate of false positive phenotypes. Furthermore, using a CRISPR/Cas13d screening library targeting over 2,500 human hepatocellular carcinomas (HCC) related circRNAs, we identified a group of circRNAs, whose inhibition increased the therapeutic efficacy of sorafenib, an approved HCC drug. Collectively, these data demonstrate that CRISPR/Cas13d system is an effective approach to study the function of circRNAs in a high-throughput manner.

2020-12-06 | GSE162720 | GEO

Gene expression profiling by CRISPR/Cas13d-mediated CTSL knockdown in 293FT cells

Project description:To examine the potential off-target effects by CRISPR/Cas13d-mediated CTSL knockdown in 293FT cells

2022-06-09 | GSE203052 | GEO