Project description:Many new alternative splice forms have been detected at the transcript level using next generation sequencing (NGS) methods, especially RNA-Seq, but it is not known how many of these transcripts are being translated. Leveraging the unprecedented capabilities of NGS, we collected RNA-Seq and proteomics data from the same cell population (Jurkat cells) and created a bioinformatics pipeline that builds customized databases for the discovery of novel splice-junction peptides. Results: Eighty million paired-end Illumina reads and ~500,000 tandem mass spectra were used to identify 12,873 transcripts (19,320 including isoforms) and 6,810 proteins. We developed a bioinformatics workflow to retrieve high-confidence, novel splice junction sequences from the RNA data, translate these sequences into the analogous polypeptide sequence, and create a customized splice junction database for MS searching. Jurkat T-cell mRNA was analyzed on an Illumina HiSeq2000. ~80 million paired end reads (2x200bp, ~350bp lengths) were collected.
Project description:Follicular lymphoma (FL) is one of the most common types of indolent B-cell lymphoma in Western countries. FL commonly transforms to more aggressive diffuse large B-cell lymphoma (DLBCL) at reported frequencies between 15 - 60%. We have used microarray comparative genomic hybridisation (aCGH) at 1 Mb resolution to study copy number changes in paired tumor samples (primary FL and a subsequent tDLBCL) as well as de novo DLBCL cases to outline genetic mechanisms of transformation from follicular lymphoma to diffuse large B-cell lymphoma.
Project description:We report the gene expression profile of 8 metastatic castration resisistant prostate cancer samples analyzed by paired-end RNA-seq. We found evidence of extensive abnormal splicing as well as several novel fusion genes. Finally, we also observed several recurrent high-confidence somatic mutations. Paired-end RNA-seq by rRNA depletion
Project description:<p>We used massively parallel, paired-end sequencing of expressed transcripts (RNA-seq) to detect novel gene fusions in short-term cultures of glioma stem-like cells freshly isolated from nine patients carrying primary glioblastoma multiforme (GBM). The culture of primary GBM tumors under serum-free conditions selects cells that retain phenotypes and genotypes closely mirroring primary tumor profiles as compared to serum-cultured glioma cell lines that have largely lost their developmental identities.</p>
Project description:Total RNA extracted from prostate cancer LNCaP cells transfected with siRNA against CTCF(siCTCF), or negative control siRNA (si-)were processed, and sequenced by two different companies using Illumina Hi-seq 2000 platform to generate RNA sequencing with two output sequences: paired-end 50bp and 101bp in read length. Nearly 100 million and 50 million raw reads were yielded from each sample respectively. We used FastQC to confirm the quality of raw fastq sequencing data, and SOAPfuse software to detect fusion transcripts. Discovering fusion genes from siCTCF and si- in LNCaP cells.
Project description:We report the design and implementation of a "breakpoint analysis" pipeline to discover novel gene fusions by tell-tale transcript level or genomic DNA copy number transitions occurring within genes. We use this method to prioritize candidate rearrangements from high density array CGH datasets as well as exon-resolution expression microarrays. We mine both publicly available data as well as datasets generated in our laboratory. Several gene fusion candidates were chosen for further characterization, and corresponding samples were profiled using paired end RNA sequencing to discover the identity of the gene fusion. Using this approach, we report the discovery and characterization of novel gene fusions spanning multiple cancer subtypes including angiosarcoma, pancreatic cancer, anaplastic astrocytoma, melanoma, breast cancer, and T-cell acute lymphoblastic leukemia. Taken together, this study provides a robust approach for gene fusion discovery, and our results highlight a more widespread role of fusion genes in cancer pathogenesis. Breakpoint analysis for the discovery of novel gene fusions across human cancers