| 1. |
- Karami, Moein, et al.
(författare)
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Designing efficient randstrobes for sequence similarity analyses
- 2024
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Ingår i: Bioinformatics. - 1367-4803 .- 1367-4811. ; 40:4
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Tidskriftsartikel (refereegranskat)abstract
- Motivation: Substrings of length k, commonly referred to as k-mers, play a vital role in sequence analysis. However, k-mers are limited to exact matches between sequences leading to alternative constructs. We recently introduced a class of new constructs, strobemers, that can match across substitutions and smaller insertions and deletions. Randstrobes, the most sensitive strobemer proposed in Sahlin (Effective sequence similarity detection with strobemers. Genome Res 2021a;31:2080–94. https://doi.org/10.1101/gr.275648.121), has been used in several bioinformatics applications such as read classification, short-read mapping, and read overlap detection. Recently, we showed that the more pseudo-random the behavior of the construction (measured in entropy), the more efficient the seeds for sequence similarity analysis. The level of pseudo-randomness depends on the construction operators, but no study has investigated the efficacy.Results: In this study, we introduce novel construction methods, including a Binary Search Tree-based approach that improves time complexity over previous methods. To our knowledge, we are also the first to address biases in construction and design three metrics for measuring bias. Our evaluation shows that our methods have favorable speed and sampling uniformity compared to existing approaches. Lastly, guided by our results, we change the seed construction in strobealign, a short-read mapper, and find that the results change substantially. We suggest combining the two results to improve strobealign’s accuracy for the shortest reads in our evaluated datasets. Our evaluation highlights sampling biases that can occur and provides guidance on which operators to use when implementing randstrobes.Availability and implementation: All methods and evaluation benchmarks are available in a public Github repository at https://github.com/Moein-Karami/RandStrobes. The scripts for running the strobealign analysis are found at https://github.com/NBISweden/strobealign-evaluation.
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| 2. |
- Petri, Alexander J., et al.
(författare)
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isONform : reference-free transcriptome reconstruction from Oxford Nanopore data
- 2023
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Ingår i: Bioinformatics. - 1367-4803 .- 1367-4811. ; 39, s. i222-i231
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Tidskriftsartikel (refereegranskat)abstract
- Motivation With advances in long-read transcriptome sequencing, we can now fully sequence transcripts, which greatly improves our ability to study transcription processes. A popular long-read transcriptome sequencing technique is Oxford Nanopore Technologies (ONT), which through its cost-effective sequencing and high throughput, has the potential to characterize the transcriptome in a cell. However, due to transcript variability and sequencing errors, long cDNA reads need substantial bioinformatic processing to produce a set of isoform predictions from the reads. Several genome and annotation-based methods exist to produce transcript predictions. However, such methods require high-quality genomes and annotations and are limited by the accuracy of long-read splice aligners. In addition, gene families with high heterogeneity may not be well represented by a reference genome and would benefit from reference-free analysis. Reference-free methods to predict transcripts from ONT, such as RATTLE, exist, but their sensitivity is not comparable to reference-based approaches.Results We present isONform, a high-sensitivity algorithm to construct isoforms from ONT cDNA sequencing data. The algorithm is based on iterative bubble popping on gene graphs built from fuzzy seeds from the reads. Using simulated, synthetic, and biological ONT cDNA data, we show that isONform has substantially higher sensitivity than RATTLE albeit with some loss in precision. On biological data, we show that isONform's predictions have substantially higher consistency with the annotation-based method StringTie2 compared with RATTLE. We believe isONform can be used both for isoform construction for organisms without well-annotated genomes and as an orthogonal method to verify predictions of reference-based methods.Availability and implementation
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| 3. |
- Sahlin, Kristoffer, et al.
(författare)
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Accurate spliced alignment of long RNA sequencing reads
- 2021
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Ingår i: Bioinformatics. - : Oxford University Press (OUP). - 1367-4803 .- 1367-4811 .- 1460-2059. ; 37:24, s. 4643-4651
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Tidskriftsartikel (refereegranskat)abstract
- MOTIVATION: Long-read RNA sequencing technologies are establishing themselves as the primary techniques to detect novel isoforms, and many such analyses are dependent on read alignments. However, the error rate and sequencing length of the reads create new challenges for accurately aligning them, particularly around small exons.RESULTS: We present an alignment method uLTRA for long RNA sequencing reads based on a novel two-pass collinear chaining algorithm. We show that uLTRA produces higher accuracy over state-of-the-art aligners with substantially higher accuracy for small exons on simulated and synthetic data. On simulated data, uLTRA achieves an accuracy of about 60% for exons of length 10 nucleotides or smaller and close to 90% accuracy for exons of length between 11 to 20 nucleotides. On biological data where true read location is unknown, we show several examples where uLTRA aligns to known and novel isoforms containing small exons that are not detected with other aligners. While uLTRA obtains its accuracy using annotations, it can also be used as a wrapper around minimap2 to align reads outside annotated regions.
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| 4. |
- Sahlin, Kristoffer, et al.
(författare)
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Assembly scaffolding with PE-contaminated mate-pair libraries
- 2016
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Ingår i: Bioinformatics. - : Oxford University Press (OUP). - 1367-4803 .- 1367-4811. ; 32:13, s. 1925-1932
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Tidskriftsartikel (refereegranskat)abstract
- Motivation: Scaffolding is often an essential step in a genome assembly process, in which contigs are ordered and oriented using read pairs from a combination of paired-end libraries and longer-range mate-pair libraries. Although a simple idea, scaffolding is unfortunately hard to get right in practice. One source of problems is so-called PE-contamination in mate-pair libraries, in which a non-negligible fraction of the read pairs get the wrong orientation and a much smaller insert size than what is expected. This contamination has been discussed before, in relation to integrated scaffolders, but solutions rely on the orientation being observable, e.g. by finding the junction adapter sequence in the reads. This is not always possible, making orientation and insert size of a read pair stochastic. To our knowledge, there is neither previous work on modeling PE-contamination, nor a study on the effect PE-contamination has on scaffolding quality. Results: We have addressed PE-contamination in an update to our scaffolder BESST. We formulate the problem as an integer linear program which is solved using an efficient heuristic. The new method shows significant improvement over both integrated and stand-alone scaffolders in our experiments. The impact of modeling PE-contamination is quantified by comparing with the previous BESST model. We also show how other scaffolders are vulnerable to PE-contaminated libraries, resulting in an increased number of misassemblies, more conservative scaffolding and inflated assembly sizes.
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| 5. |
- Sahlin, Kristoffer, et al.
(författare)
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Improved gap size estimation for scaffolding algorithms
- 2012
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Ingår i: Bioinformatics. - Oxford : Oxford University Press (OUP). - 1367-4803 .- 1367-4811. ; 28:17, s. 2215-2222
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Tidskriftsartikel (refereegranskat)abstract
- Motivation: One of the important steps of genome assembly is scaffolding, in which contigs are linked using information from read-pairs. Scaffolding provides estimates about the order, relative orientation and distance between contigs. We have found that contig distance estimates are generally strongly biased and based on false assumptions. Since erroneous distance estimates can mislead in subsequent analysis, it is important to provide unbiased estimation of contig distance.Results: In this article, we show that state-of-the-art programs for scaffolding are using an incorrect model of gap size estimation. We discuss why current maximum likelihood estimators are biased and describe what different cases of bias we are facing. Furthermore, we provide a model for the distribution of reads that span a gap and derive the maximum likelihood equation for the gap length. We motivate why this estimate is sound and show empirically that it outperforms gap estimators in popular scaffolding programs. Our results have consequences both for scaffolding software, structural variation detection and for library insert-size estimation as is commonly performed by read aligners.
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