Questions tagged [read-mapping]
When dealing with associating sequencing reads to a matching position in a set of reference sequences (typically a genome).
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Meaning of BWA-MEM MAPQ scores
Does anyone know what the MAPQ values produced by BWA-MEM mean?
I'm looking for something similar to what Keith Bradnam ...
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Why is bwa-mem the standard algorithm when using bwa?
The industry standard for aligning short reads seems to be bwa-mem. However, in my tests I have seen that using bwa backtrack (bwa-aln + bwa-sampe + bwa-samse) performs better. It is slightly slower, ...
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How does the BWA-MEM algorithm assign its mapping qualities?
Is there any resource (paper, blogpost, Github gist, etc.) describing the BWA-MEM algorithm for assigning mapping qualities? I vaguely remember that I have somewhere seen a formula for SE reads, which ...
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Library for computing BWT-based alignments
I am writing a software tool to which I would like to add the ability to compute alignments using the efficient Burrows-Wheeler Transform (BWT) approach made popular by tools such as BWA and Bowtie. ...
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Difference between BWA-backtrack and BWA-MEM
Many of my colleagues recommend I use BWA-MEM instead of regular old BWA. The problem is I don't understand why and reading the BWA man page doesn't seem to help the matter.
What is the difference ...
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Visualisation of long read RNA-Seq splicing
I have a dataset of Oxford Nanopore cDNA reads. Many of my reads are full-length or close to full-length transcripts, and I and am interested in examining alternative splicing. For this, I would like ...
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How do I generate a variant list (i.e. VCF file) using Illumina reads from a human genome?
This is a problem I have to solve frequently, and I'd be interested in knowing what other methods people use to solve the same problem.
About twice a year, I get asked to determine variants from ...
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What is mate rescue in bwa mem?
BWA mem has the -S and -P tags for skip mate rescue and skip pairing; mate rescue performed unless -S also in use.
What do ...
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What are "split reads" and "intron clusters?"
I'm working through the example data set for LeafCutter and the documentation mentions "split reads":
This will cluster together the introns fond [sic] in the junc files listed
in test_juncfiles....
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Highly heterozygous reads mapping
I have short (67bp) Hi-C reads from a highly heterozygous organism (~15% between-haplotype SNP divergence) and I have both reference haplotypes.
I wanted to try and compare different haplotyping ...
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Definition of "seed" in sequence alignment
I would like to know what is meant by "seed" for various sequence aligners. How is it important?
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What is the correct way to map Hi-C data with bwa mem?
Library Prep
I have a Hi-C library prepped using an enzyme that cuts at GATC, so it leaves GATCGATC as the junction sequences. ...
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How to trim adapter sequences from GSE65360 in order to map the reads?
I am trying to map single-cell chromatin accessibility data from doi:10.1038/nature14590, obtained using scATAC-seq, to the reference genome. Example paired-end reads are here.
The reads contain ...
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How to interpret contig-alignment.psa produced by velvet
I'm using velvet to align given reads of RNA to given CDSs (i.e. coding areas and genes) of an organism, so I can generate gene-expression profiles. But after using ...
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Why total RNA-seq usually yields low mapping rate?
Maybe this is a silly question, but I'm really wondering why we usually get low mapping rates if we map total RNA-seq, but not poly(A)-enriched (in particular, for human, mouse, and zebrafish datasets)...
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Filter Trinity transcriptome based on RNASeq reads
I have recently generated a genome-guided transcriptome with Trinity, and would like to apply an additional filter to exclude transcripts that don't have good support from the RNASeq reads. This is ...
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What is the difference between SAM mapping quality and Blast E-value?
Blast reports E-values, but short-read mappers report mapping qualities. Are they the same thing? Can they be converted to each other? If not, why blast doesn't report mapping quality while short-read ...
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How to estimate whether a long-read is meaningful sequence?
The setup
Imagine that I work on an organism without a reference genome, and that the closest reference genome I can get is quite diverged. E.g. ~10% diverged in terms of SNVs when measured with ...
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What is local realignment and what is the problem it solves?
I am trying to understand local realignment but I could not get a clear idea of what is the problem solved by it.
For example, reading Homer and Nelson (2010):
Because alignment algorithms map ...
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RapMap: reference transcriptome for simulated reads
I am very new to bioinformatics and trying to repeat the benchmark in the RapMap paper with an experimental tool working in a similar but different fashion.
In the paper (taken from their github) ...
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Control width of samtools tview "snapshot" when redirecting
I have a large number of loci I would like to examine manually with samtools tview. Rather than typing or copy-n-pasting dozens of coordinates, I was hoping to ...
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Why is SMALT better for microbial genomics than other mappers?
SMALT seems to be one of the most used read mappers for bacterial data, see, e.g., this query. I do not say that it is not a great mapper, but I cannot easily see what are its main strengths compared ...
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Displaying soft-clipped nucleotides in samtools tview
There are nicer genomics visualization tools available, but the samtools tview command is almost always my go-to for a quick first look at read alignments. I just ...
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what percentage of the human genome is MAPQ=0?
When doing Illumina 2x150bp sequencing of genomic DNA, and after aligning the reads to GRCh38, what percentage of the non-N fraction of the human genome is MAPQ=0? This is, what part corresponds to ...
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mapping nucleotide sequences
I am trying to do mapping with multiple sequences against a reference genome. I tried the following code: The outputfile is .fastq.sam. I need only .sam. How do I get it?
...
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Kallisto error: index input file could not be opened!
I am utilizing Kallisto in Anaconda/miniconda for RNA sequencing. I have successfully made ...
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Hisat2 : which option should mention for strand specific library read
I was trying hisat2 I get confused by the strand options...
The question has been already asked on github here but didn't get any satisfactory answer.
I'm surprise how much this strand information ...
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Problem: "Pair-end" reads scRNA seq data (Drop-seq)
In case of Drop-seq, we have paired end data.
Read 1: Cell code + UMI (unique molecule identifier)
Read 2: The transcript information
But I have a problem/doubt with the sample I am working on.
...
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Reference genome for allele specific expression
We are trying to sort out a pipeline for doing allele specific expression. Our plan is to call SNPs from RNA-seq data and combine with known SNP annotations. A well known problem in ASE is reference ...
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Can I run STAR without an annotation file?
I wish to use Rascaf to scaffold a fragmented draft genome.
For this, I need to provide a BAM file of aligned RNA-seq reads and the draft genome.
So, I indexed the draft genome with STAR like this:
<...
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Best way to detect long insertions in bisulfite sequencing data?
I am interested in identifying indels in whole genome bisulfite sequencing data (76bp paired end). Currently, I do this by setting the -rfg and ...
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Problems to Extract uniquely mapping reads from BWA MEM alignment
I did a mapping of genomic paired-end reads to a reference assembly using bwa mem. I need to extract the reads that mapped only once to my reference.
For that, I have tried to follow the method ...
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How to count the number of mapped read in 100-bp window from a BAM/SAM file
Although I know how to get total number of mapped read using samtools flagstat (samtools flagstat file_sorted.bam) but I want to count total number of mapped read ...
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Which reference to use for read mapping for popular model organisms
What is the "best" assembly for the popular model organisms:
human (GRCh37 and GRCh38 are obvious, I'd pick whatever bwakit uses)
mouse (GRCm37/GRCm38, OK)
but what about non-human/mouse ones?
...
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How to check what software has been used to generate bam file?
I have several bam files from unknown origin. I'd like to know what software was used to obtain those bam files, e.g. bwa-mem. Is it possible to somehow check it?
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Contamination on genome assembly
I had a question for the community.
I have a genome of a new species that has been sequenced via 150pb Illumina paired-end.
To verify the quality of the assembly I used the ...
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How to map reads shorter than 32bp with minimap2?
I mapped fasta sequences to a reference. The fasta sequences range in length from 17bp up to several hundred bp. I used minimap2 with the following command: ...
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Counts obtained by featureCounts seem much less than observed coverage
I have surprisingly low counts when running featureCounts on some (single-end) RNA-seq data mapped on C. elegans genome using ...
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Strange per sequence GC content results
I would be grateful if someone could take a quick look at these FASTQC results. This is rna-seq paired-end data. From the FASTQC manual, an unusual distribution seems to be suggestive of contamination ...
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Reference genome (bwa -mem)
I want to do mapping with bwa mem. I downloaded a reference genome from ncbi database which is (Danaus plexippus). I got one ncbi_database.zip file which contain 2 ...
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How do I generate a variant list using Illumina reads from a Salmonella genome?
I am planning on performing phylogenetic analysis of Salmonella specimens using WGS data from PulseNet and GenomeTrakr, for the purpose of public health surveillance & to provide context for ...
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RNASeq read coverage in protein space?
I used samtools depth to find the per base-pair read coverage over a number of isoform contigs from my Trinity assembly.
I have also conducted a multiple sequence ...
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Patient-sample mapping in GSE72056 dataset
I want to use the single-cell data from the following expression profiling which concerns the RNA-seq of metastatic melanoma:
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE72056
The data was ...
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Is there a read mapper that allows input in SAM format instead of FASTA/FASTQ?
When we get raw sequenced reads in FASTQ format, we usually need to do some pre-processing (QC, demultiplexing, etc.) prior to sequence alignment. The only way to register the results of this ...
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Error with BWA Mem input having multiple fastq files using cat and process substitution
Running BWA Mem on a number of paired end fastq files using process substitution on the inputs results in this error:
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Understanding PAF format
Does anyone have any experience with PAF files (minimap2 output)? I don’t fully understand what my output means.
I’ve looked at the minimap2 explanation (https://github.com/lh3/miniasm/blob/master/PAF....
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What do the symbols mean in minimap2's gap cost equation?
Heng Li's paper on minimap2 is understandable to me right up until the moment that it gives this definition of gap cost:
2.2.1 Alignment with 2-piece affine gap cost
Minimap2 performs DP-based ...
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bedtools: get name column of alignment file A
I'm using the coverage function of bedtools to check whether a certain set of regions (file B) has any overlap with known ...
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What tool I can use to map short-reads sequences to reference genome and get specific mapped size
I have about 300 90-bp sequences which I would like to map to a reference genome to make it longer to a 300bp sequence, wherein the 90bp is at the middle.
Anyone knows what bioinformatics tool I can ...
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Oxford Nanopore mapping Quality and sequencing error
I'm analyzing some Oxford Nanopore sequencing reads, with several articles stating the sequencing error is between 5% and 15%. Since its 1D and not 1D^2 sequencing i would assume the sequencing error ...
