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• W2465461003 abstract "In recent years, the sequencing of RNA (RNA-seq) using next generation sequencing (NGS) technology has become a powerful tool for analyzing the transcriptomic state of a cell. Modern NGS platforms allow for performing RNA-seq experiments in a few days, resulting in millions of short sequencing reads. A crucial step in analyzing RNA-seq data generally is determining the transcriptomic origin of the sequencing reads (= read mapping). In principal, read mapping is a sequence alignment problem, in which the short sequencing reads (30 - 500 nucleotides) are aligned to much larger reference sequences such as the human genome (3 billion nucleotides).In this thesis, we present ContextMap, an RNA-seq mapping approach that evaluates the context of the sequencing reads for determining the most likely origin of every read. The context of a sequencing read is defined by all other reads aligned to the same genomic region. The ContextMap project started with a proof of concept study, in which we showed that our approach is able to improve already existing read mapping results provided by other mapping programs. Subsequently, we developed a standalone version of ContextMap. This implementation no longer relied on mapping results of other programs, but determined initial alignments itself using a modification of the Bowtie short read alignment program. However, the original ContextMap implementation had several drawbacks. In particular, it was not able to predict reads spanning over more than two exons and to detect insertions or deletions (indels). Furthermore, ContextMap depended on a modification of a specific Bowtie version. Thus, it could neither benefit of Bowtie updates nor of novel developments (e.g. improved running times) in the area of short read alignment software.For addressing these problems, we developed ContextMap 2, an extension of the original ContextMap algorithm. The key features of ContextMap 2 are the context-based resolution of ambiguous read alignments and the accurate detection of reads crossing an arbitrary number of exon-exon junctions or containing indels. Furthermore, a plug-in interface is provided that allows for the easy integration of alternative short read alignment programs (e.g. Bowtie 2 or BWA) into the mapping workflow. The performance of ContextMap 2 was evaluated on real-life as well as synthetic data and compared to other state-of-the-art mapping programs. We found that ContextMap 2 had very low rates of misplaced reads and incorrectly predicted junctions or indels. Additionally, recall values were as high as for the top competing methods. Moreover, the runtime of ContextMap 2 was at least two fold lower than for the best competitors.In addition to the mapping of sequencing reads to a single reference, the ContextMap approach allows the investigation of several potential read sources (e.g. the human host and infecting pathogens) in parallel. Thus, ContextMap can be applied to mine for infections or contaminations or to map data from meta-transcriptomic studies. Furthermore, we developed methods based on mapping-derived statistics that allow to assess confidence of mappings to identified species and to detect false positive hits. ContextMap was evaluated on three real-life data sets and results were compared to metagenomics tools. Here, we showed that ContextMap can successfully identify the species contained in a sample. Moreover, in contrast to most other metagenomics approaches, ContextMap also provides read mapping results to individual species. As a consequence, read mapping results determined by ContextMap can be used to study the gene expression of all species contained in a sample at the same time. Thus, ContextMap might be applied in clinical studies, in which the influence of infecting agents on host organisms is investigated.The methods presented in this thesis allow for an accurate and fast mapping of RNA-seq data. As the amount of available sequencing data increases constantly, these methods will likely become an important part of many RNA-seq data analyses and thus contribute valuably to research in the field of transcriptomics." @default.
• W2465461003 created "2016-07-22" @default.
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• W2465461003 date "2016-04-22" @default.
• W2465461003 modified "2023-09-24" @default.
• W2465461003 title "Context-based RNA-seq mapping" @default.
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