Homo sapiens
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA147303 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:We have developed two methods for efficiently consructing RNA-seq libraries using transposition. Each method constructs high quality RNA-seq libraries when compared to standard approaches. One of the methods (Directional Tn-RNA-seq) maintains strand-of-origin information and exhibits strand specificity comparable to current approaches. RNA-seq libraries were constructed from ECC-1, a human cell line, and Universal Human Reference RNA using transposon-based and standard RNA-seq library construciton methods.
2011-10-10 | E-GEOD-32307 | biostudies-arrayexpress
Project description:Tn5 transposon tagments double-strand DNA and RNA/DNA hybrid to generate nucleic acids ready to be amplified for high throughput sequencing. The nucleic acid substrates of Tn5 transposon need to be explored to increase the applications of Tn5. Here, we find that Tn5 transposon is able to transport oligos into the 5’ end of single-strand DNA, which are more than 140 base pairs of nucleic acids. Based on this property of Tn5, we develop a tagmentation-based and ligation-enabled single-strand DNA sequencing method, TABLE-seq. Through a series of reaction temperature, time, and enzyme concentration tests, we managed to apply this TABLE-seq to strand-specific RNA sequencing, starting with as low as 30 picograms of total RNA. Moreover, compared with traditional dUTP based strand-specific RNA sequencing, this method detects more genes, has higher strand-specificity, and shows more evenly distributed reads across genes. Together, our results provide insights into the properties of Tn5 transposons and expand the applications of Tn5 in cutting-edge sequencing techniques.
2023-01-30 | GSE208316 | GEO
Project description:Strand-specific massively-parallel cDNA sequencing (RNA-Seq) is a powerful tool for novel transcript discovery, genome annotation, and expression profiling. Despite multiple published methods for strand-specific RNA-Seq, no consensus exists as to how to choose between them. Here, we developed a comprehensive computational pipeline for the comparison of library quality metrics from any RNA-Seq method. Using the well-annotated Saccharomyces cerevisiae transcriptome as a benchmark, we compared seven library construction protocols, including both published and our own novel methods. We found marked differences in complexity, strand-specificity, evenness and continuity of coverage, agreement with known annotations, and accuracy for expression profiling. Weighing each method’s performance and ease, we identify the dUTP second strand marking and the Illumina RNA ligation methods as the leading protocols, with the former benefitting from the availability of paired-end sequencing. Our analysis provides a comprehensive benchmark, and our computational pipeline is applicable for assessment of future protocols in any organism. Examination of 11 different strand-specific RNA-Seq libraries from 7 distinct methods; also 2 control non-strand-specific RNA-Seq libraries. To assess the performance of each strand-specific library in digital expression profiling, we compared them to reference expression measurements estimated from expression profiles using competitive hybridization of a mid-log RNA sample vs. genomic DNA using Agilent arrays.
2010-08-15 | E-GEOD-21739 | biostudies-arrayexpress
Project description:We have examined the nuclear (nuc) and cytoplasmic (cyt) polyA+ transcriptomes of undifferentiated mouse embryonic stem cells (un) and cells differentiated to neural precursors (d5) using strand-specific RNA-Seq. The 46C mouse embryonic stem cell line was used for this study. Two cell types were examined: undifferentiated mouse embryonic stem cells (un) and cells differentiated to neural precursors (d5). For each cell type, cells were fractionated to nuclear and cytoplasmic components. RNAs were extracted from each component and were fragmented enzymatically for library construction. For each cell type and component, strand-specific RNA-Seq libraries were generated using at least two different fragmentation protocols.
2012-08-01 | E-GEOD-38990 | biostudies-arrayexpress
Project description:Intervention type:DRUG. Intervention1:Huaier, Dose form:GRANULES, Route of administration:ORAL, intended dose regimen:20 to 60/day by either bulk or split for 3 months to extended term if necessary. Control intervention1:None.
Primary outcome(s): For mRNA libraries, focus on mRNA studies. Data analysis includes sequencing data processing and basic sequencing data quality control, prediction of new transcripts, differential expression analysis of genes. Gene Ontology (GO) and the KEGG pathway database are used for annotation and enrichment analysis of up-regulated genes and down-regulated genes.
For small RNA libraries, data analysis includes sequencing data process and sequencing data process QC, small RNA distribution across the genome, rRNA, tRNA, alignment with snRNA and snoRNA, construction of known miRNA expression pattern, prediction New miRNA and Study of their secondary structure Based on the expression pattern of miRNA, we perform not only GO / KEGG annotation and enrichment, but also different expression analysis.. Timepoint:RNA sequencing of 240 blood samples of 80 cases and its analysis, scheduled from June 30, 2022..
| 2612481 | ecrin-mdr-crc
Project description:One method of directional cloning of fragmented mRNA is based on single strand RNA ligation, for which ligation bias occurrs if the adapters have single sequences. The sequencing bias affects the mRNA quantification and subsequently the differential expression analysis. High definition (HD) adapters [Sorefan et al 2012, Xu et al 2015] can be used to diminish the cloning bias during library construction. HD adapters and standard illumina adapters were used to construct mRNA-seq libraries for a side by side comparison.
2019-10-01 | GSE102510 | GEO
Project description:Primary human hepatocytes were treated with 200µM DHA/EPA/OA or vehicle(ethanol) for 16 hours before cells were harvested for RNA extraction. Total RNA was isolated from primary human hepatocytes using KingFisher PURE RNA tissue Kit. The eluted RNA was subjected for strand specific sequencing libraries construction with illumina TruSeq RNA sample Prep Kit and subsequently sequenced by NHLBI DNA sequencing and Genomic Core using Illumina HiSeq 3000 paired-end sequencing platform.
2021-03-10 | GSE165354 | GEO
Project description:Strand-specific RNA-seq libraries were constructed for two samples, including (I) wild-type strain NBRC0988 grown in YEP medium containing 2% w/v glucose;(II) wild-type strain NBRC0988 grown in YEP medium containing 2% w/v xylose. For preparation of RNA samples, NBRC0988 cells grown overnight were inoculated into 100 ml liquid Yeast Extract Peptone Dextrose (YEPD) medium with the initial inoculation amount of OD600= 0.1, and cultured for 15 hours at 30℃ and 250 rpm. The cells were collected by centrifugation at 6,000g for 5 minutes. After washing twice with phosphate buffer saline (PBS), they were inoculated into new 100 mL YEP medium containing 2% w/v glucose or xylose.After flask culturing at 30°C and 250 rpm for an additional 5 hours, the yeast cells were collected by centrifugation for total RNA isolation and Illumina RNA-seq library construction. Total RNA for samples were isolated using TRIzol reagent (Invitrogen, Grand Island, USA), then used for high-throughput RNA sequencing. The 150-nt paired-end strand-specific RNA-seq libraries (SS_lib_type RF) were generated commercially at Novogene Biotechnology Co. Ltd (Tianjin, China) by using Illumina’s novaseq 6000 platform (Illumina, San Diego, USA).
2022-12-11 | GSE220542 | GEO
Project description:Next-generation sequencing has been widely used for the genome-wide profiling of histone modifications, transcription factor binding and gene expression through chromatin immunoprecipitated DNA sequencing (ChIP-seq) and cDNA sequencing (RNA-seq). Here, we describe a versatile library construction method that can be applied to both ChIP-seq and RNA-seq on the widely used Illumina platforms. Standard methods for ChIP-seq library construction require nanograms of starting DNA, substantially limiting its application to rare cell types or limited clinical samples. By minimizing the DNA purification steps that cause major sample loss, our method achieved a high sensitivity in ChIP-seq library preparation. Using this method, we achieved the following: (1) generated high-quality epigenomic and transcription factor-binding maps using ChIP-seq for murine adipocytes; (2) successfully prepared a ChIP-seq library from as little as 25 pg of starting DNA; (3) achieved paired-end sequencing of the ChIP-seq libraries; (4) systematically profiled gene expression dynamics during murine adipogenesis using RNA-seq; and (5) preserved the strand specificity of the transcripts in RNA-seq. Given its sensitivity and versatility in both double-stranded and single-stranded DNA library construction, this method has wide applications in genomic, epigenomic, transcriptomic and interactomic studies. Pre-adipocytes and mature adipocytes were collected. Their chromatin and RNA were subjected to ChIP and mRNA extraction. Sequencing libraries from ChIP DNA or mRNA were generated following either standard protocols or TELP method. The quality and features of TELP libraries were proved and demonstrated in comparison with standard libraries or other published data.
2015-12-15 | E-GEOD-75966 | biostudies-arrayexpress