Questions tagged [sam]
Use this tag for questions specifically related to the text SAM format. For general questions about the SAM/BAM formats use the tag BAM.
226
questions
42
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What is the difference between FASTA, FASTQ, and SAM file formats?
I'd like to learn the differences between 3 common formats such as FASTA, FASTQ and SAM. How they are different? Are there any benefits of using one over another?
Based on Wikipedia pages, I can't ...
- 1,305
20
votes
12
answers
2k
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Random access on a FASTQ file
I would like to select a random record from a large set of n unaligned sequencing reads in log(n) time complexity (big O ...
- 2,166
19
votes
2
answers
12k
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Obtaining uniquely mapped reads from BWA mem alignment
This is based on a question from betsy.s.collins on BioStars. The original post can be found here.
Does anyone have any suggestions for other tags or filtering steps on BWA-generated BAM files that ...
- 13k
18
votes
3
answers
4k
views
Convert a BAM file from one reference to another?
I have a set of BAM files that are aligned using the NCBI GRCh37 human genome reference (with the chromosome names as NC_000001.10) but I want to analyze it using a BED file that has the UCSC hg19 ...
- 540
18
votes
3
answers
13k
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How can I downsample a BAM file while keeping both reads in pairs?
I know how to downsample a BAM file to lower coverage. I know I can randomly select lines in SAM, but this procedure can't guarantee two reads in a pair are always sampled the same time. Is there a ...
- 777
17
votes
3
answers
5k
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BAM to BigWig without intermediary BedGraph
I have a pipeline for generating a BigWig file from a BAM file:
BAM -> BedGraph -> BigWig
Which uses bedtools genomecov ...
16
votes
2
answers
9k
views
Merge hundreds of small BAM files into a single BAM file
I am working with over a million (long) reads, and aligning them to a large genome. I am considering running my alignment jobs in parallel, distributing horizontally across hundreds of nodes rather ...
- 2,113
15
votes
1
answer
4k
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What is the difference between samtools, bamtools, picard, sambamba and biobambam?
After some google searches, I found multiple tools with overlapping functionality for viewing, merging, pileuping, etc. I have not got time to try these tools, so will just see if anyone already know ...
- 777
14
votes
1
answer
870
views
Why would someone use a CRAM instead of a BAM?
I had this question from a graduate student yesterday, and I was stuck.
What should I say? Why use a CRAM instead of a BAM?
When is it a good idea to use a CRAM instead of a BAM?
When is it a bad ...
- 1,373
14
votes
1
answer
2k
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Are soft-clipped bases used for variant calling in samtools + bcftools?
If there are soft clipped base pairs specified in the CIGAR string for a read in a SAM/BAM file, will these be used for variant calling in a samtools + bcftools workflow?
The GATK HaplotypeCaller, ...
- 744
14
votes
1
answer
232
views
Compare alignment quality of multiple sequencing runs aligned against the same reference genome
I have run Oxford Nanopore Technologies' MinION sequencing on the same DNA sample using three flowcells, each aligned against the same reference genome (E.coli K12 MG1655) using both BWA MEM and ...
- 2,113
13
votes
5
answers
1k
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Good / recommended way to archive fastq and bam files?
We have a lot of Illumina sequenced exome data. Currently we are using spring for its great lossless compression, but we are looking if there is anything better (and most preferably opensource) which ...
- 151
11
votes
2
answers
378
views
Do variant calls change when you call from CRAM?
We're considering switching our storage format from BAM to CRAM. We work with human cancer samples, which may have very low prevalence variants (i.e. not diploid frequency).
If we use lossy CRAM to ...
- 540
11
votes
1
answer
967
views
Quantifying reads mapping to multiple loci
I have been using STAR for our RNA-Seq samples. The final.out log file reports percentage of uniquely mapped reads along with percentage of reads that map to ...
- 971
11
votes
1
answer
275
views
Can I create a CRAM file with a relative reference path?
I’m trying to create a CRAM file that stores its path to the FASTA reference as a relative path, rather than an absolute path, so that I can move the files around. Unfortunately I can’t get this to ...
- 4,815
10
votes
2
answers
283
views
What are the de facto required fields in a SAM/BAM read group?
The SAM specification indicates that each read group must have a unique ID field, but does not mark any other field as required.
I have also discovered that htsjdk throws exceptions if the sample (...
- 744
9
votes
2
answers
11k
views
Converting a BAM file into VCF
I have NGS illumina RNA-seq reads from M. musculus (mm10). I am trying to find variants along the strand portion of the reads in the refseq (mm10).
I mapped a pair of sequence files and generated a ...
- 361
9
votes
1
answer
681
views
Why does this human bam file only have one copy of each chromosome?
As we know that in human DNA sequence, one copy of chromosome comes from mother's DNA and another copy comes from father's DNA so as to form two copies of each chromosome in human DNA. So, if we ...
- 550
9
votes
2
answers
1k
views
Why do BAM files created by different tools have different file sizes?
I have a BAM created by Picard. I want to filter alignments by flags with samtools view. However, I noticed that even if I apply no filters, the output BAM is ...
- 777
9
votes
1
answer
5k
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How to count reads in bam per bed interval with bedtools
I recently installed Ubuntu 16.04 (because I was still using 12.04). But it seems my bedtools scripts don't work properly anymore. I can't figure out how to use the new bedtools for my old ways. What ...
- 3,561
9
votes
2
answers
5k
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samtools depth print out all positions
I am trying to use samtools depth (v1.4) with the -a option and a bed file listing the human chromosomes chr1-chr22, chrX, chrY, and chrM to print out the coverage ...
- 2,274
8
votes
7
answers
9k
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How to subset a BAM by a list of QNAMEs?
I have a text file 'qnames.txt' with QNAMEs in the following format:
EXAMPLE:QNAME1
EXAMPLE:QNAME2
EXAMPLE:QNAME3
EXAMPLE:QNAME4
EXAMPLE:QNAME5
I would like to ...
- 1,373
8
votes
4
answers
1k
views
How to manipulate a reference FASTA or bam to include variants from a VCF?
I have some software which takes fastas as the input. I need to include SNVs and InDels from a VCF into the reference hg38 and then use this.
The problem is, I don't know of an algorithmically sound ...
- 1,651
8
votes
2
answers
443
views
Convert bam file to highly compressible bam
I have a large collection of bam files and I want to post-process each of them into another bam where I can make queries about:
the reads position and pair-endness,
insert sizes,
MAPQ and other ...
- 2,274
8
votes
3
answers
3k
views
How can I count the number of reads that support a variant in a bam file?
I am calling variants from a human sample using bwa mem to align the reads and gatk to call the variants. I'm trying to understand why a specific variant was not called in my sample. I have checked ...
- 9,352
8
votes
3
answers
3k
views
How to merge sam files together with adding read groups
I have three sequencing libraries of single individual mapped to a reference using bwa-mem. I would like to merge the three unsorted ...
- 5,467
8
votes
2
answers
545
views
How to import a large amount of .bed, .gff, .vcf, .paf, .sam files into an SQL database?
Are there best practices to load different bioinformatics file formats such as VCF, BED, GFF, and SAM to SQL databases? I am wondering how people out there do that efficiently.
All of these three ...
- 1,417
8
votes
1
answer
261
views
Convert local alignments to spliced alignments in SAM file
I mapped RNA reads to reference genome, using LAST in split mode, and converted the MAF alignment to SAM with maf-convert.
My problem is that the transcripts are not reported in a spliced manner, ...
- 2,656
8
votes
0
answers
436
views
How can I read multiple different regions from a BAM file in R?
I’m trying to process BAM records (of a coordinate-sorted, indexed BAM file) successively in fixed-size, non-overlapping, sliding genome coordinate windows. Unfortunately I am unable to read records ...
- 4,815
7
votes
3
answers
671
views
Is there a way to retrieve several SAM fields faster than `samtools view | cut -f`?
I am constructing a bit of software which pipes the outputs of the bam file via samtools view into a script for parsing. My goal is to (somehow) make this process ...
- 1,651
7
votes
3
answers
3k
views
How to check whether all BAM read contain defined read groups?
I'm trying to investigate whether there are errors within my BAM. After looking at the BAM header to see whether the read groups exists (using samtools, i.e.
...
- 1,651
7
votes
1
answer
749
views
ethnicity check either from bam or vcf files
What tool could I use to check the ethnicity of a human bam or vcf file? I would like to use the results as a QC check to know whether a given sample or set of samples match the ethnicity information ...
- 2,274
7
votes
2
answers
589
views
How does htslib/samtools access optional BAM fields?
I am confused regarding how various software libraries deal with optional fields in a BAM:
Based upon the BAM specification, there are 11 mandatory fields to a BAM:
...
- 1,373
7
votes
1
answer
2k
views
How to remove all BAM read groups from all reads (not just the header)?
I have problem with one my BAMs---it appears to have invalid read groups.
Normally when I have such a problem, I remove all the read groups from the BAM header as follows:
...
- 1,373
7
votes
1
answer
491
views
Split FASTQ and matching BAM into matching chunks
I am running a slow downstream analysis on a large set of nanopore reads (approx 3 million), and would like to split them into smaller chunks, run the analysis in massively parallel, and then ...
- 2,113
7
votes
2
answers
3k
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Extracting the CIGAR string from a BAM via Python?
Is there a standard method in Python to extract a CIGAR string from the BAM?
There are great libraries which parse the CIGAR, e.g. https://pypi.python.org/pypi/cigar/0.1
...
- 1,651
7
votes
1
answer
774
views
Filtering bases based on phred qualities with pysam
Is there a way to filter bases in BAM files based on phred quallities through python's pysam ?
I have a code here that
Takes the nucleobases per position from a BAM file using pysam's pileup ...
7
votes
1
answer
304
views
scanBam from Rsamtools is not importing one of my reads into R
I have this read in my BAM file. It maps on chromosome 1.
I open this BAM file in IGV, and I can see the alignment on chromosome 1.
But when I open this file in R with Rsamtools:
...
- 2,449
7
votes
1
answer
175
views
How can I speed up INDEL calling/correction on BAM files?
The samtools mpileup command has quite a neat feature that it is able to correct mapping errors associated with misalignment of INDELs. By default, the ...
- 13k
7
votes
1
answer
702
views
low-memory high-speed replacement for Picard MarkDuplicates
I am running Picard MarkDuplicates with the following parameters below. On the file described, it takes about 41.6Gb of RAM memory and about 20-25 minutes to compute (only uses 1 core AFAICS).
...
- 2,274
7
votes
1
answer
429
views
samtools mpileup empty when filtering out flags
I produced a bam file by aligning reads to a small set of synthetic sequences using bwa-mem.
I am heavily filtering reads that are not paired and of a certain orientation.
Applying the filtering, I ...
- 2,274
7
votes
1
answer
570
views
Reject reads with low quality bases from a Bam file through pysam
I have a code below:
...
6
votes
1
answer
471
views
Why do SAM and BAM use different coordinate systems?
BAM files are, at least as far as I know, simply binary compressed versions of SAM files. They have the exact same information and are used in the same way. Why then does the SAM format use a 1-based ...
- 9,352
6
votes
3
answers
1k
views
Total reads aligning to each reference within a bam file
I have two PCR amplicons that have been multiplexed and sequenced using the nanopore minion.
I have aligned the fastq reads using minimap2 with a reference file containing both amplicon sequences and ...
- 63
6
votes
5
answers
3k
views
Subset smaller BAM to contain several thousand rows from multiple chromosomes
There are many cases whereby I would like to subset a BAM to create a small file in order to work with (e.g. algorithmic testing, debugging, etc.)
Normally I do the following, which will subset the ...
- 1,373
6
votes
1
answer
298
views
How to validate that BAMs have been downloaded correctly?
I currently have several hundred BAM files which were downloaded by someone else. These have remained untouched---before working with them, I would like to double-check that these BAMs have been fully ...
- 1,373
6
votes
3
answers
751
views
Generating the reconstructed alignment from BAM
I have a (small) BAM file with CIGAR and MD fields.
Question 1: What tools exists in Python and/or R to reconstruct the alignment between the reference and the read in a BAM? Given that this is a ...
- 1,651
6
votes
2
answers
4k
views
Get the mapping statistics of a single read from a BAM file
I have a BAM file, and I have a read ID. What is the simplest way to get mapping statistics of that read in human-readable format?
E.g. I might want: % identity of aligned bases; number of insertions ...
- 952
6
votes
1
answer
577
views
Why most aligners do not output the "X" CIGAR operation?
As I read the SAM spec, the "X" CIGAR operator represents a mismatch. This seems useful as we can know where are the mismatches without looking at the reference genome. However, many popular aligners ...
- 777
6
votes
1
answer
2k
views
Read counts from BAM file
I have few BAM files which are generated from Ion Torrent Server (ampliseq) aligned to hg19 genome. I want to extract read counts from the bam files and I know that "featureCounts" can be used for ...
- 1,252