Ion Proton Sequencing
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ABSTRACT:
PROVIDER: PRJEB27429 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:Evaluation of AmpliSeq and SureSelect whole exome sequencing methods on Ion Proton
| PRJNA481671 | ENA
Project description:Five libraries from 100 HEK293 cells each were prepared using a Smartseq based custom library preparation approach with unique molecular identifiers. One batch of 2 replicates (A) and one batch of 3 replicates (B) were prepared from different cell cultures. Libraries were sequenced on an Ion Proton
2016-12-19 | GSE79123 | GEO
Project description:Evaluation of Whole-Genome Sequencing of Four Chinese Crested Dogs for Variant Detection Using the Ion Proton System
| PRJEB10523 | ENA
Project description:Background: Whole transcriptome sequencing (RNA-seq) represents a powerful approach for whole transcriptome gene expression analysis. However, RNA-seq carries a few limitations, e.g., the requirement of a significant amount of input RNA and complications led by non-specific mapping of short reads. The Ion AmpliSeqTM Transcriptome Human Gene Expression Kit (AmpliSeq) was recently introduced by Life Technologies as a whole-transcriptome, targeted gene quantification kit to overcome these limitations of RNA-seq.To assess the performance of this new methodology, we performed a comprehensive comparison of AmpliSeq with RNA-seq using two well-established next-generation sequencing platforms (Illumina HiSeq and Ion Torrent Proton). We analyzed standard reference RNA samples and RNA samples obtained from human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs). Results: Using published data from two standard RNA reference samples, we observed a strong concordance of log2 fold change for all genes when comparing AmpliSeq to Illumina HiSeq (Pearson’s r=0.92) and Ion Torrent Proton (Pearson’s r=0.92). We used ROC, Matthew’s correlation coefficient and RMSD to determine the overall performance characteristics. All three statistical methods demonstrate AmpliSeq as a highly accurate method for differential gene expression analysis. Additionally, for genes with high abundance, AmpliSeq outperforms the two RNA-seq methods. When analyzing four closely related hiPSC-CM lines, we show that both AmpliSeq and RNA-seq capture similar global gene expression patterns consistent with known sources of variations. Conclusions: Our study indicates that AmpliSeq excels in the limiting areas of RNA-seq for gene expression quantification analysis. Thus, AmpliSeq stands as a very sensitive and cost-effective approach for very large scale gene expression analysis and mRNA marker screening with high accuracy.
2016-02-11 | GSE74760 | GEO
Project description:CD3+ T cells were enriched using the EasySep human T cell isolation kit (Stem cell technology). T cells from normal controls or patients with CARD9 mutations were co-cultured with monocytes of one normal control in the presence of heat killed candida. After 3 days, T cell were enriched again to remove the monocytes and total RNA was extracted from the T cells for the RNA-sequencing (RNASeq) evaluation. Targeted RNA sequencing library preparation was carried out using the Ion AmpliSeq Transcriptome Human Gene Expression Kit (Life Technologies), which profiles more than 20,000 human genes; each amplicon (~150 bp) represents a unique targeted gene (one transcript per gene). For library preparation, each sample was run in duplicate, and a cDNA library was generated from a minimum of 10 ng of total RNA. The cDNA was barcoded and amplified with Ion AmpliSeq technology, and the amplified cDNA Libraries were evaluated for quality and quantified with Agilent Bioanalyzer high-sensitivity chip. Libraries were then diluted to 100 pM and pooled equally, with 4 individual samples per pool. Pooled libraries were amplified and enriched with the Ion Chef System (Life Technologies). Templated libraries were then sequenced on an Ion Torrent Proton sequencing system (Life Technologies) with Ion PI HiQ kit and chip version 3. We performed gene-level differential expression analysis of targeted RNASeq data using R (v.3.5.3) and the Bioconductor packages DESeq2 (v.1.22.2).
2021-08-12 | GSE181718 | GEO
Project description:Whole exome sequencing data of 19 snap-frozen peritoneal mesothelioma (tumor) samples and 16 matched normal samples. Sequencing library was prepared using Ion AmpliSeq Exome RDY Library Preparation. Samples were sequenced on the Ion Proton System using the Ion PI Hi-Q Sequencing 200 Kit and Ion PI v3 chip.
| EGAD00001004503 | EGA
Project description:Ion AmpliSeq transcriptome analysis of atypical hepatocytes
| PRJNA643843 | ENA
Project description:Binge drinking contributes to a rising number of emergency room visits in the United States. Our previous work demonstrated that binge drinking impairs cardiac performance and exerts complex hemodynamic alterations through activation of the endocannabinoid–CB1 receptor (CB1R) signaling pathway. Anandamide, an endogenous CB1R agonist, is produced in response to stimuli such as endotoxin challenge and oxidative stress; however, the effect of binge drinking on myocardial anandamide production and subsequent CB1R-mediated cardiodepression remains unclear. Here, we studied the impact of endotoxins derived from intestinal Gram-negative bacteria on myocardial anandamide formation and CB1R-dependent cardiac dysfunction following acute alcohol intoxication. Using a mouse model of binge drinking (5 g/kg orally), we observed a reduced rate of mesenteric microcirculation concurrent with elevated circulating endotoxin levels. Selective intestinal decontamination significantly lowered circulating endotoxin levels, coinciding with reduced cardiac anandamide levels at 3 hours post-binge. These changes were paralleled with improved left ventricular performance, vascular tone, and reduced myocardial oxidative stress. Moreover, cardiac RNA sequencing revealed marked alterations in gene expression following alcohol gavage that were absent in mice with prior selective intestinal decontamination. These findings provide novel mechanistic insights that highlight the crucial role of intestinal Gram-negative bacteria in modulating cardiac anandamide production after binge drinking, thereby exacerbating cardiovascular function and blood redistribution. Cardiac tissues were homogenized in Trizol (Invitrogen, Carlsbad, CA, USA), and total RNA was isolated with a Direct-zol RNA Miniprep Kit (Zymo Research, Irvine, CA, USA) according to the manufacturer’s instructions. RNA concentrations were determined using a NanoDrop 2000 Spectrophotometer (Thermo Scientific). AmpliSeq libraries were prepared using the Ion AmpliSeq™ Library Kit Plus (Thermo Fisher Scientific, 4488990) per the manufacturer’s instructions. Library quality was determined using the Agilent High Sensitivity DNA Kit (Agilent Technologies, 5067-4626), and library concentrations were quantified using the Ion Library TaqMan™ Quantitation Kit (Life Technologies, 4468802), per manufacturer’s instructions. Barcoded AmpliSeq libraries were loaded on the sequencing chips using the Ion 540TM Chip Kit (ThermoFisher, A27766) and Ion Chef™ Instrument. Libraries were sequenced on the Ion Torrent S5 Sequencing System via the Ion Torrent 540-OT2 kit (ThermoFisher, A27753). 8 to 9 libraries were loaded per sequencing chip, and 2 sequencing ships were loaded and sequenced at a time. An average of 8.5 million counts per sample were sequenced. Alignment and gene expression count were performed using Ion Torrent AmpliSeq RNA Plugin v0.5.4.0 (Thermo Fisher) using mm10 genome.
2025-09-01 | GSE294937 | GEO
Project description:Next-generation sequencing (NGS) technology applications like RNA-sequencing (RNA-seq) have dramatically expanded the potential for novel genomics discoveries, but the proliferation of various platforms and protocols for RNA-seq has created a need for reference data sets to help gauge the performance characteristics of these disparate methods. Here we describe the results of the ABRF-NGS Study on RNA-seq, which leverages replicate experiments across multiple sites using two reference RNA standards tested with four protocols (polyA selected, ribo-depleted, size selected, and degraded RNA), and examined across five NGS platforms (Illumina’s HiSeqs, Life Technologies’ Personal Genome Machine and Proton, Roche 454 GS FLX, and Pacific Biosciences RS). These results show high (R2 >0.9) intra-platform consistency across test sites, high inter-platform concordance (R2 >0.8) for transcriptome profiling, and a large set of novel splice junctions observed across all platforms. Also, we observe that protocols using ribosomal RNA depletion can both salvage degraded RNA samples and also be readily compared to polyA-enriched fractions. These data provide a broad foundation for standardization, evaluation and improvement of RNA-seq methods. Two reference RNA standards tested with four protocols (polyA selected, ribo-depleted, size selected, and degraded RNA), and examined across five NGS platforms (Illumina’s HiSeqs, Life Technologies’ Personal Genome Machine and Proton, Roche 454 GS FLX, and Pacific Biosciences RS). Please note that the samples were named following the ABRF-Platform-Site-Sample-Replicate# format. For example, ABRF-454-CNL-A-1 means Sample A was run on 454 platform at Cornell and this is the first replicate, and ABRF-454-CNL-A-2 means the same exact sample was ran with same machine at same location and is 2nd replicate.
2014-05-14 | E-GEOD-48034 | biostudies-arrayexpress
Project description:Next-generation sequencing (NGS) technology applications like RNA-sequencing (RNA-seq) have dramatically expanded the potential for novel genomics discoveries, but the proliferation of various platforms and protocols for RNA-seq has created a need for reference data sets to help gauge the performance characteristics of these disparate methods. Here we describe the results of the ABRF-NGS Study on RNA-seq, which leverages replicate experiments across multiple sites using two reference RNA standards tested with four protocols (polyA selected, ribo-depleted, size selected, and degraded RNA), and examined across five NGS platforms (Illumina’s HiSeqs, Life Technologies’ Personal Genome Machine and Proton, Roche 454 GS FLX, and Pacific Biosciences RS). These results show high (R2 >0.9) intra-platform consistency across test sites, high inter-platform concordance (R2 >0.8) for transcriptome profiling, and a large set of novel splice junctions observed across all platforms. Also, we observe that protocols using ribosomal RNA depletion can both salvage degraded RNA samples and also be readily compared to polyA-enriched fractions. These data provide a broad foundation for standardization, evaluation and improvement of RNA-seq methods.
2014-05-14 | GSE48034 | GEO