Dataset Information

RAMPAGE: promoter activity profiling by paired-end sequencing of 5'-complete cDNAs.

ABSTRACT: RNA annotation and mapping of promoters for analysis of gene expression (RAMPAGE) is a method that harnesses highly specific sequencing of 5'-complete complementary DNAs to identify transcription start sites (TSSs) genome-wide. Although TSS mapping has historically relied on detection of 5'-complete cDNAs, current genome-wide approaches typically have limited specificity and provide only scarce information regarding transcript structure. RAMPAGE allows for highly stringent selection of 5'-complete molecules, thus allowing base-resolution TSS identification with a high signal-to-noise ratio. Paired-end sequencing of medium-length cDNAs yields transcript structure information that is essential to interpreting the relationship of TSSs to annotated genes and transcripts. As opposed to standard RNA-seq, RAMPAGE explicitly yields accurate and highly reproducible expression level estimates for individual promoters. Moreover, this approach offers a streamlined 2- to 3-day protocol that is optimized for extensive sample multiplexing, and is therefore adapted for large-scale projects. This method has been applied successfully to human and Drosophila samples, and in principle should be applicable to any eukaryotic system.

SUBMITTER: Batut P

PROVIDER: S-EPMC4372803 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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Publications

RAMPAGE: promoter activity profiling by paired-end sequencing of 5'-complete cDNAs.

Batut Philippe P Gingeras Thomas R TR

Current protocols in molecular biology 20131111

RNA annotation and mapping of promoters for analysis of gene expression (RAMPAGE) is a method that harnesses highly specific sequencing of 5'-complete complementary DNAs to identify transcription start sites (TSSs) genome-wide. Although TSS mapping has historically relied on detection of 5'-complete cDNAs, current genome-wide approaches typically have limited specificity and provide only scarce information regarding transcript structure. RAMPAGE allows for highly stringent selection of 5'-comple ...[more]

PMID: 24510412

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Project description:BackgroundA cancer genome is derived from the germline genome through a series of somatic mutations. Somatic structural variants - including duplications, deletions, inversions, translocations, and other rearrangements - result in a cancer genome that is a scrambling of intervals, or "blocks" of the germline genome sequence. We present an efficient algorithm for reconstructing the block organization of a cancer genome from paired-end DNA sequencing data.ResultsBy aligning paired reads from a cancer genome - and a matched germline genome, if available - to the human reference genome, we derive: (i) a partition of the reference genome into intervals; (ii) adjacencies between these intervals in the cancer genome; (iii) an estimated copy number for each interval. We formulate the Copy Number and Adjacency Genome Reconstruction Problem of determining the cancer genome as a sequence of the derived intervals that is consistent with the measured adjacencies and copy numbers. We design an efficient algorithm, called Paired-end Reconstruction of Genome Organization (PREGO), to solve this problem by reducing it to an optimization problem on an interval-adjacency graph constructed from the data. The solution to the optimization problem results in an Eulerian graph, containing an alternating Eulerian tour that corresponds to a cancer genome that is consistent with the sequencing data. We apply our algorithm to five ovarian cancer genomes that were sequenced as part of The Cancer Genome Atlas. We identify numerous rearrangements, or structural variants, in these genomes, analyze reciprocal vs. non-reciprocal rearrangements, and identify rearrangements consistent with known mechanisms of duplication such as tandem duplications and breakage/fusion/bridge (B/F/B) cycles.ConclusionsWe demonstrate that PREGO efficiently identifies complex and biologically relevant rearrangements in cancer genome sequencing data. An implementation of the PREGO algorithm is available at http://compbio.cs.brown.edu/software/.

Dataset Information

RAMPAGE: promoter activity profiling by paired-end sequencing of 5'-complete cDNAs.

Publications

RAMPAGE: promoter activity profiling by paired-end sequencing of 5'-complete cDNAs.

Similar Datasets

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