Whole genome bisulfite sequencing on 10 multiple myeloma cases
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PROVIDER: EGAS00001004346 | EGA |
REPOSITORIES: EGA
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Project description:Whole genome bisulfite sequencing on 10 multiple myeloma cases. Data quality control and adaptor-trimmed were performed with the Trimomatic tool. Paired-reads were mapped to the hg19 human reference with methylCtools aligner.
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2020-04-23 | GSE130241 | GEO
Project description:Small RNAs (<100 bases in length) were purified from total RNA samples using the miRNeasy kit (Qiagen), and multiplexed miRNA libraries were prepared using a previously described protocol and sequenced on a HiSeq 2000 (Illumina). Image analysis, base calling, demultiplexing and conversion of BCL to FASTQ format were carried out using CASAVA 1.8.2 software (Illumina). Adaptor sequences were removed using mirExpress software. Fastq were 3p-adaptor trimmed. The sequences were then aligned on hg19 (GRCh37) human reference genome with STAR (v.2.5.2a), allowing no mismatch with reference. Reads were counted using HTSeq-count (HTSeq v 0.6.1p1 Python package) using “intersection-strict” mode, to count mature miRNA abundance according to miRbase v.20.
2020-08-12 | E-MTAB-6553 | biostudies-arrayexpress
Project description:For ATAC-seq data processing, we used the ENCODE ATAC-seq pipeline (https://www.encodeproject.org/atac-seq/). In detail, for each sample, ATAC-seq reads were first checked for adaptor contamination. Then, adaptor trimmed reads were aligned to hg19 using Bowtie2 under paired-end mode with parameter “-X2000”, which permits 2000 bp for the maximum allowable insert size between two paired ends of each read. Using the ENCODE ATAC-seq pipeline, duplicated reads were properly filtered out. ATAC-seq enriched peaks were called using MACS2 (Zhang et al., 2008) based on remaining unique reads with parameters “-f BAMPE -q 0.05 --nomodel”. In total, we collected 45,569 peaks for the ARID1AWT sample and 27,480 peaks for the ARID1AKO sample.
2020-06-08 | GSE106660 | GEO
Project description:Small RNA sequencing has been performed using eight commonly used breast cancer cell lines by Illumina as part of the Illumina iDEA Challange 2011. Total RNA of the cell lines was isolated and the sequencing library was prepared using Illuminas Small RNA v1.5 library prep protocol. This data was analyzed with Flicker, an add-on to the Illumina Genome Analyzer Pipeline software that allows for the preliminary analysis of small RNA runs on the GA. Flicker was used in this case for: * Trimming: Attempts to trim the adaptor sequence from small RNA reads Trimming is accomplished by aligning the adaptor with each read, both internally (the adaptor is fully contained within the read) and as a suffix (the adaptor overlaps the 3' end of the read). A similarity matrix is used to generate the matching score, taking into account both substitutions and indels. The alignment with the best matching score is returned, and that best alignment is trimmed from the read. A trimming threshold may be specified, and the adaptor is not trimmed unless the best score exceeds the threshold. * Aligning with Iterated ELAND: Attempts to aligns trimmed reads to targets (squashed genomes or databases such as miRBase) using the ELAND short read aligner. Each read is aligned by ELAND with specified alignment targets. ELAND was designed to align reads of a uniform length, and the trimmed reads from a small RNA run are several different lengths. We currently address this by running ELAND for reads of each length: reads are binned by length, and each bin of a given length is aligned against a specified target. Once all bins are aligned against all targets (in this case, gDNA and miRBase), the results are recombined into a single file with all the annotation information. ===== Key to Experiments HCT ID, Lane, Sample ID, Cell Line, ER +/- Status, Reads HCT20061, Lane 1, HS378, MCF7, +, 14,555,390 HCT20062, Lane 2, HS379, MDA-MB-231, -, 15,147,037 HCT20063, Lane 3, HS381, T47D, +, 14,701,254 HCT20064, Lane 4, HS377, BT-20, -, 16,529,823 HCT20065, Lane 5, HS375, BT-474, +, 16,443,361 HCT20066, Lane 6, HS380, MDA-MB-468, -, 16,746,981 HCT20067, Lane 7, HS382, ZR-75-1, +, 17,631,005 HCT20068, Lane 8, HS376, MCF10A, normal, 19,155,607
2016-03-14 | E-MTAB-4539 | biostudies-arrayexpress
Project description:We performed the RNA-seq in control samples and FXR1 knockdown samples, and compared the gene expression profiles to explore the effect of FXR1 knockdown on gene expression. The study was performed in H358 cells. Doxycycline inducible shRNA3 (sh3) was used to knockdown FXR1. Control shRNA (ctrl) samples were used to get rid of the effect of Doxycycline treatment. Both the Doxycycline treament for 3 days (D3) and 5 days (D5) samples were collected. Each sample has three repeats (rep 1, rep 2, and rep 3). The mRNA profiles were generated by deep sequencing using Illumina.Sequenced reads were trimmed for adaptor sequence, then mapped to hg19 whole genome using STAR v2.5.3 with parameters --bamRemoveDuplicatesType UniqueIdentical --outSAMmultNmax 1. Raw reads and Reads Per Kilobase per Megabase of library size (RPKM) were calculated using HOMER (PMID: 20513432). Differential gene expression was analyzed using R package DESeq2 using the raw reads.
2017-10-04 | GSE101754 | GEO
Project description:This experiment is to explore the mechanism of anti-inflammatory activity of celastrol derivatives on Rheumatoid arthritis fibroblast-like synoviocytes (RAFLS), compared with celastrol. Immortalized RAFLS cells were simulated with 1 ng/mL TNFα for 24h, and exposed to different treatments during the last 4 hours. Total RNA was extracted with Trizol. Transcriptomic profiling was performed with an in-house, proprietary method that captures the 3’ primed end of mRNA molecules for sequencing. The data was processed with a custom script. Briefly, sequencing reads were demultiplexed based on sample barcode with idemux, trimmed with cutadapt to remove polyA, adaptor and low-quality sequences, mapped to human GRCh38 reference assembly with STAR aligner. PCR duplicates were removed based on UMI with UMI-tools. Htseq-count was used to generate read counts. The count matrix was analyzed with DESeq2 package to determine differential genes. Enrichment and GSEA was performed with clusterprofiler.
2025-09-01 | GSE300831 | GEO
Project description:LSEC fom WT and Bmp9-KO 25-weeks female mice were isolated using collagenase and treated for bulk RNAsequencing. The objective was to determine the biological processes that are altered in Bmp9-KO compared to WT, as BMP9 is know to be a vascular quiescence factor produced by hepatic stellate cells. RNAseq reads were trimmed to remove possible adapter sequences and nucleotides with poor quality using Trimmomatic v.0.36 16. The trimmed reads were mapped to the Mus Musculus MM10 reference genome available on ENSEMBL using the STAR aligner v.2.5.2b 17. Unique gene hit counts were calculated by using ‘featureCounts’ from the Subread package v.1.5.2 WT samples are F126, F121, F120, F119, F135 KO samples are F131, F132, F124 and F133
2024-04-30 | E-MTAB-14012 | biostudies-arrayexpress
Project description:This data was generated by ENCODE. If you have questions about the data, contact the submitting laboratory directly (Jonathan Preall jpreall@cshl.edu (Generation 0 Data from Hannon Lab), Carrie Davis davisc@cshl.edu (experimental), Alex Dobin dobin@cshl.edu (computational), Wei Lin wlin@cshl.edu (computational), Tom Gingeras gingeras@cshl.edu (primary investigator)). If you have questions about the Genome Browser track associated with this data, contact ENCODE (mailto:genome@soe.ucsc.edu). hg18: This data was produced by Hannon lab part of Cold Spring Harbor as part of the ENCODE Project. The series depicts NextGen sequencing information for RNAs between the sizes of 20-200 nt isolated from RNA samples from tissues or sub cellular compartments of cell lines. hg19: This track depicts NextGen sequencing information for RNAs between the sizes of 20-200 nt isolated from RNA samples from tissues or sub cellular compartments from ENCODE cell lines. The overall goal of the ENCODE project is to identify and characterize all functional elements in the sequence of the human genome. hg19: This cloning protocol generates directional libraries that are read from the 5' ends of the inserts, which should largely correspond to the 5' ends of the mature RNAs. The libraries were sequenced on a Solexa platform for a total of 36, 50 or 76 cycles however the reads undergo post-processing resulting in trimming of their 3' ends. Consequently, the mapped read lengths are variable. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf hg18: Small RNAs between 20-200 nt were ribominus treated according to the manufacturer's protocol (Invitrogen) using custom LNA probes targeting ribosomal RNAs (some datasets are also depleted of U snRNAs and high abundant microRNAs). The RNA was treated with Tobacco Alkaline Pyrophosphatase to eliminate any 5' cap structure. Poly-A Polymerase was used to catalyze the addition of C's to the 3' end. The 5' ends were phosphorylated using T4 PNK and an RNA linker was ligated onto the 5' end. Reverse transcription was carried out using a poly-G oligo with a defined 5' extension. The inserts were then amplified using oligos targeting the 5' linker and poly-G extension and containing sequencing adapters. The library was sequenced on an Illumina GA machine for a total of 36, 50 or 76 cycles. Initially 1 lane is run. If an appreciable number of mappable reads are obtained, additional lanes are run. Sequence reads underwent quality filtration using Illumina standard pipeline (Gerlad). The read lengths may exceed the insert sizes and consequently introduce 3' adaptor sequence into the 3' end of the reads. The 3' sequencing adaptor was removed from the reads using a custom clipper program, which aligned the adaptor sequence to the short-reads, allowing up to 2 mismatches and no indels. Regions that aligned were "clipped" off from the read. The trimmed portions were collapsed into identical reads, their count noted and aligned to the human genome (NCBI build 36, hg18 unmasked) using Nexalign (Lassmann et al., not published). The alignment parameters are tuned to tolerate up to 2 mismatches with no indels and will allow for trimmed portions as small as 5 nucleotides to be mapped. We report reads that mapped 10 or fewer times. Data obtained from each lane is processed and mapped independently. The processed/mapped data from each lane is then complied as a single track without additional processing and submitted to UCSC. Consequently, identical reads within a lane were collapsed and their value is reported as the "transfrag" signal value. However, the redundancy between lanes has not been eliminated so the same transfrag may appear multiple times within a signal. hg19: Small RNAs between 20-200 nt were ribominus treated according to the manufacturer's protocol (Invitrogen) using custom LNA probes targeting ribosomal RNAs (some datasets are also depleted of U snRNAs and high abundant microRNAs). The RNA was treated with Tobacco Alkaline Pyrophosphatase to eliminate any 5' cap structures. Poly-A Polymerase was used to catalyze the addition of C's to the 3' end. The 5' ends were phosphorylated using T4 PNK and an RNA linker was ligated onto the 5' end. Reverse transcription was carried out using a poly-G oligo with a defined 5' extension. The inserts were then amplified using oligos targeting the 5' linker and poly-G extension and containing sequencing adapters. The library was sequenced on an Illumina GA machine for a total of 36, 50 or 76 cycles. Initially, one lane was run. If an appreciable number of mappable reads were obtained, additional lanes were run. Sequence reads underwent quality filtration using Illumina standard pipeline (GERALD). The Illumina reads were initially trimmed to discard any bases following a quality score less than or equal to 20 and converted into FASTA format, thereby discarding quality information for the rest of the pipeline. As a result, the sequence quality scores in the BAM output are all displayed as "40" to indicate no quality information. The read lengths may exceed the insert sizes and consequently introduce 3' adapter sequence into the 3' end of the reads. The 3' sequencing adapter was removed from the reads using a custom clipper program (available at http://hannonlab.cshl.edu/fastx_toolkit/), which aligned the adapter sequence to the short-reads using up to 2 mismatches and no indels. Regions that aligned were "clipped" off from the read. Terminal C nucleotides introduced at the 3' end of the RNA via the cloning procedure are also trimmed. The trimmed portions were collapsed into identical reads, their count noted and aligned to the human genome (version hg19, using the gender build appropriate to the sample in question - female/male) using Bowtie (Langmead B. et al). The alignment parameter allowed 0, 1, or 2 mismatches iteratively. We report reads that mapped 20 or fewer times. Discrepancies between hg18 and hg19 versions of CSHL small RNA data: The alignment pipeline for the CSHL small RNA data was updated upon the release of the human genome version hg19, resulting in a few noteworthy discrepancies with the hg18 dataset. First, mapping was conducted with the open-source Bowtie algorithm (http://bowtie-bio.sourceforge.net/index.shtml) rather than the custom NexAlign software. As each algorithm uses different strategies to perform alignments, the mapping results may vary even in genomic regions that do not differ between builds. The read processing pipeline also varies slightly, in that we no longer retain information regarding whether a read was 'clipped' off adapter sequence.
2010-10-08 | E-GEOD-24565 | biostudies-arrayexpress
Project description:Purpose: To asses the transcriptome of 4 strains of the Tasmanian Devil facial tumor and one healthy fibroblast cell line. Methods: DFTD cells were grown from primary cell cultures derived from fine needle aspirates that have been collected from the wild. Total RNA from devil facial tumor strains 1-4 and the fibroblast was isolated from approximately 1 x 10^6 cells using QIAzol lysis reagent according to the manufacturer’s instructions (Qiagen). RNA-seq libraries were prepared with TruSeq Stranded mRNA LT sample preparation kit (Illumina) using Sciclone and Zephyr liquid handling robotics (PerkinElmer). The 5 libraries were pooled, diluted and sequenced on Illumina HiSeq 3000/4000 using 75 bp paired-end chemistry. Base calls provided by the Illumina Realtime Analysis software were converted into BAM format using Illumina2bam and demultiplexed using BamIndexDecoder. Paired-end reads were trimmed for adaptor sequences and filtered with the trimmomatic tool. Trimmed reads were aligned on the version 7.0 of the Tasmanian Devil with the STAR aligner. Counting of reads on annotated transcripts (Sarcophilus_harrisii.DEVIL7.0.90.gtf from Ensembl) was performed with featurecounts on fragments. The DESeq2 Biconductor library has been used for counts normalization and differential analysis between the transcriptomes of the four DFTD and fibroblast cell lines
2019-01-14 | GSE108107 | GEO