High-purity circular RNA isolation method (RPAD) reveals vast collection of intronic circRNAs (IcircRNAs)
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ABSTRACT:
OTHER RELATED OMICS DATASETS IN: GSE92632
PROVIDER: PRJNA358203 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR5122011_1.fastq.gz | Fastqsanger.gz | |||
| SRR5122011_2.fastq.gz | Fastqsanger.gz | |||
| SRR5122012_1.fastq.gz | Fastqsanger.gz | |||
| SRR5122012_2.fastq.gz | Fastqsanger.gz | |||
| SRR5122013_1.fastq.gz | Fastqsanger.gz |
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Similar Datasets
Project description:High-throughput RNA sequencing methods coupled with specialized bioinformatic analyses have recently uncovered tens of thousands of unique circular (circ)RNAs, but their complete sequences, genes of origin, and functions are largely unknown. Given that circRNAs lack free ends and are thus relatively stable, their association with microRNAs (miRNAs) and RNA-binding proteins (RBPs) can influence gene expression programs. While exoribonuclease treatment is widely used to degrade linear RNAs and enrich circRNAs in RNA samples, it does not efficiently eliminate linear RNAs. Here, we describe a novel method for the isolation of highly pure circRNA populations involving RNase R treatment followed by Polyadenylation and poly(A)+ RNA Depletion (RPAD), which removes linear RNA to near completion. High-throughput sequencing of RNA prepared using RPAD from human cervical carcinoma HeLa cells and mouse C2C12 myoblasts led to two surprising discoveries: (1) many circRNA isoforms share an identical backsplice sequence but have different body sizes and sequences, and (2) thousands of novel intronic circular RNAs (IcircRNAs) are expressed in cells, including many IcircRNAs that do not follow gt/ag splicing patterns. In sum, isolating high-purity circRNAs using the RPAD method can enable quantitative and qualitative analyses of circRNA types and sequence composition, paving the way for the elucidation of circRNA functions.
2017-05-03 | GSE92632 | GEO
Project description:High-throughput RNA sequencing methods coupled with specialized bioinformatic analyses have recently uncovered tens of thousands of unique circular (circ)RNAs, but their complete sequences, genes of origin and functions are largely unknown. Given that circRNAs lack free ends and are thus relatively stable, their association with microRNAs (miRNAs) and RNA-binding proteins (RBPs) can influence gene expression programs. While exoribonuclease treatment is widely used to degrade linear RNAs and enrich circRNAs in RNA samples, it does not efficiently eliminate all linear RNAs. Here, we describe a novel method for the isolation of highly pure circRNA populations involving RNase R treatment followed by Polyadenylation and poly(A)+ RNA Depletion (RPAD), which removes linear RNA to near completion. High-throughput sequencing of RNA prepared using RPAD from human cervical carcinoma HeLa cells and mouse C2C12 myoblasts led to two surprising discoveries: (i) many exonic circRNA (EcircRNA) isoforms share an identical backsplice sequence but have different body sizes and sequences, and (ii) thousands of novel intronic circular RNAs (IcircRNAs) are expressed in cells. In sum, isolating high-purity circRNAs using the RPAD method can enable quantitative and qualitative analyses of circRNA types and sequence composition, paving the way for the elucidation of circRNA functions.
| S-EPMC5499592 | biostudies-literature
Project description:The study aimed to identify circular RNAs (circRNAs) commonly back-spliced to intronic region of different sets of endothelial cells (human cardiac microvascular endothelial cells (HCMEC), human aortic endothelial cells (HAoEC), human umbilical vein endothelial cells (HUVECs)) and to evaluate their overall expression and their expression compared to their respective host gene. Identified circRNAs were quality controlled by their detection in an additional exonuclease RNase R treated RNA-Seq dataset performed with RNA of HUVECs. CircRNAs were compared for overlapping detection between datasets and filtered by annotation for circRNAs back-spliced to intronic regions. Common endothelial intronic circRNAs candidates were compared to respective murine circRNAs stored in the circATLAS database. The prime candidate cZNF292 was functionally characterized in vivo and in vitro.
2023-11-30 | E-MTAB-11171 | biostudies-arrayexpress
Project description:To explore the overall circRNAs involved in growth and development of Arabidopsis thaliana across the lifespan, we deeply sequenced samples of whole plants from different developmental stages (cotyledons emergence, rosette leavesï¹¥1 mm, rosette growth complete, first flower open, flourishing florescence, first silique shattered, senescence). The total RNA was purified by rRNA-depletion and linear RNA removal with RNAseR, and sequenced by the Illumina HiSeq2500 platform. We obtained 31 Gb raw data and identified 1217 circRNAs with expression quantity. We annotated these circRNAs and predicted their targeted microRNA. The circRNAs involved in growth and development of Arabidopsis thaliana across lifespan were identified and analyzed using the Illumina HiSeq2500 platform.
2016-09-01 | E-GEOD-77672 | biostudies-arrayexpress
Project description:Mouse Human Purity mixing studies
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Project description:We used human + mouse cell mixtures to demonstrate the purity of PIP-seq single cell RNA-sequencing
2023-01-12 | GSE202911 | GEO
Project description:We propose a statistical algorithm MethylPurify that uses regions with bisulfite reads showing discordant methylation levels to infer tumor purity from tumor samples alone. With purity estimate, MethylPurify can identify differentially methylated regions (DMRs) from individual tumor samples without genomic variation information or prior knowledge from other datasets. In simulations with mixed bisulfite reads from cancer and normal cell lines, MethylPurify correctly inferred tumor purity and identified over 96% of the DMRs. On real patient data where tumor to normal comparison were used as golden standard, MethylPurify called DMR from tumor samples alone at over 57% sensitivity and 91% specificity.
2014-07-08 | GSE56712 | GEO
Project description:BACKGROUND: Transcription of large numbers of non-coding RNAs originating from intronic regions of human genes has been recently reported, but mechanisms governing their biosynthesis and biological functions are largely unknown. In this work, we evaluated the existence of a common mechanism of transcription regulation shared by protein-coding mRNAs and intronic RNAs by measuring the effect of androgen on the transcriptional profile of a prostate cancer cell line. RESULTS: Using a custom-built cDNA microarray enriched in intronic transcribed sequences, we found 39 intronic non-coding RNAs for which levels were significantly regulated by androgen exposure. Orientation-specific reverse transcription-PCR indicated that 10 of the 13 were transcribed in the antisense direction. These transcripts are long (0.5-5 kb), unspliced and apparently do not code for proteins. Interestingly, we found that the relative levels of androgen-regulated intronic transcripts could be correlated with the levels of the corresponding protein-coding gene (asGAS6 and asDNAJC3) or with the alternative usage of exons (asKDELR2 and asITGA6) in the corresponding protein-coding transcripts. Binding of the androgen receptor to a putative regulatory region upstream from asMYO5A, an androgen-regulated antisense intronic transcript, was confirmed by chromatin immunoprecipitation. CONCLUSIONS: Altogether, these results indicate that at least a fraction of naturally transcribed intronic non-coding RNAs may be regulated by common physiological signals such as hormones, and further corroborate the notion that the intronic complement of the transcriptome play functional roles in the human gene-expression program. Keywords: Time course study – effect of androgen on gene expression
2007-01-01 | GSE5345 | GEO
Project description:The purity of tissue samples can affect the accuracy and utility of DNA methylation array analyses. This is particularly important for the placenta which is hypomethylated. Placental villous tissue from early pregnancy terminations can be difficult to separate from the non-villous tissue, resulting in potentially inaccurate results. We used several methods to identify mixed placental samples using DNA methylation array datasets from our laboratory and those contained in the NCBI GEO database, highlighting the importance of determining sample purity during quality control processes.
2020-06-09 | GSE131945 | GEO
Project description:In the process of clone-based genome sequencing, initial assemblies frequently contain cloning gaps that can be resolved using cloning-independent methods, but the reason for their occurrence is largely unknown. By analyzing 9,328,693 sequencing clones from 393 microbial genomes, we systematically mapped more than 15,000 genes residing in cloning gaps and experimentally showed that their expression products are toxic to the Escherichia coli host. A subset of these toxic sequences was further evaluated through a series of functional assays exploring the mechanisms of their toxicity. Among these genes, our assays revealed novel toxins and restriction enzymes, and new classes of small, non-coding toxic RNAs that reproducibly inhibit E. coli growth. Further analyses also revealed abundant, short, toxic DNA fragments that were predicted to suppress E. coli growth by interacting with the replication initiator DnaA. Our results show that cloning gaps, once considered the result of technical problems, actually serve as a rich source for the discovery of biotechnologically valuable functions, and suggest new modes of antimicrobial interventions.
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