Minimap2 sam header. [main_samview] fail to read the header from "-".

Minimap2 sam header Well htslib has a lot of functionality not needed for minimap2. bam and. gz | samtools sort -o out. File example. fa sample. Where precision matters, you can use LAST, bearing in mind that it calculates local alignments, and can report multiple hits per query/reference pair. I'm trying to convert a SAM file into a BAM file. I want to generate a VCF file with as many sample as the multi-fasta sequence used in alignment. create_sam_cmd = BwaMemCommandline(reference=reference_genome, read_file1=in_file) Until now, I cannot figure out to add the RG to the bam file as the RG header is absent in the files. I am using Minimap2 for aligning it to the human genome. I only see this when aligning to a reference that is effectively a database of fasta files concatenated together. bam [sam_header_read2] 25 sequences loaded. Can the minimap2 aligned use the fasta header to generate @rg within the SAM file to preserve the downstream pipeline? 2. fa > align. bam [W::sam_hdr_create] Duplicated sequence "ptg000001c_rc_rotated" in file "-" [E::sam_hrecs_update_hashes] Duplicate entry "ptg000001c_rc_rotated" in sam header This suppresses all SQ lines for mappings against references with more than one sequence, right? No. Following the docs, I have tried --sam-hit-only but get: [ERROR] unknown option in "--sam-hit-only" Is it still possible to run minimap2 with this funct I didn't think the size of the header would be the issue, because I have another sam file with a bigger header (4971104417 bytes) where the sort function did not give any errors. Simply treat it as an ordinary gzip stream, skip/read the header as is and start parsing the records. VN: Format version: SO: Sorting order of alignments: GO: Grouping of alignments @PG A SAM file contains a "header" and a series of "records". sam sed -i '1d' align. 11. sam samtools view -bST hg18. sam trancriptome. sam file (see below), it gives an error: I saw a bunch of similar issues so here I try to summarize my understanding of minimap2's terms and control flags. The gtf2bed conversion seemed to function perfectly fine so I don't think that's the issue. Let’s step through some of the header compo‐ nents in more detail: @SQ header entries store information about the reference sequences (e. Dear @NJeanray. Care should be taken to I'd like to use minimap2 to generate SAM output that only includes the mapped reads. I concatenated reads fasta files and assembled the output with hifiasm. Introduction. I performed the alignment with minimap2 to get the SAM file and now I'm using this command to convert to BAM: samtools view -bs sample. fai sample. sam to align sample_name. More posts you may like Related Bioinformatics Computer Why would your SAM file not have a header? Did you not use-a output in the SAM format when you did the alignments? If you did not, then your alignment output is in PAF format (which is the default). README: general documentation; Manpage: explanation of command-line options; Peer-reviewed paper: algorithms and evaluations (please cite if you use minimap2); Preprint: similar to the paper but free of charge; GitHub Issues page: report bugs, the software dependencies will be automatically deployed into an isolated environment before execution. but this feels like quite an unstable solution relying on an undocumented argument and a non-standard FASTA/Q read-header formatting. Minimap2 is not only a command line tool, but also a programming library. fa data. How do I fix my headers so my command generates a bam file samtools view -bS joelle. Hi, I ran wtdbg2 in the following way and ran into the below 2 Errors: conda activate wtdbg2 # assemble long reads wtdbg2 -x $2 -g $1 -i $4 -t 16 -fo dbg # derive consensus wtpoa-cns -t 16 -i dbg. fq > aln. This suppresses all SQ lines if the index is too large to be hold in RAM (e. hifiasm. bam - Then it shows that: [E::sam_hrecs_update_hashes] Duplicate entry ":516332_sim4" in sam header samtools view: failed to add PG line to the header what should i do in this situation? I have this problem, too. I have mapped the reference chromosome assembly on other chromosome assemblies using minimap2 to generate sam file. raw. fq > aln2self. pafr processes data stored in the Pairwise mApping Format (PAF), as produced by minimap2 and other whole-genome aligners. nucest. pl this step is all piped: minimap2 -t 24 -2 -I 1000G -K 1000G -x map-pb Barley_Morex_V2_pseudomolecules. Sign in Product I added a very simple read group. I Instant search for meanings of SAM (Sequence Alignment/Map) format headers (faster than searching PDF) Toggle navigation SAM Format . Navigation Menu Toggle navigation. md at master · lh3/miniasm Hi Ben, Thanks a lot for your reply and valuable help! Yes, I am running that with the sample name ERR3357550. /contigs. fastq reference. bam # 自动建索引，并输出转换结果 Therefore when you remove some read groups from the header, samtools gets confused. samtools; transcriptome; trinity; Share. gz And yea I have an . nt). More posts you may like Related Bioinformatics (2) sam文件不包含header或者header不包含@SQ ，即sam中不包含了reference的信息，此时需要提供生成sam文件时使用的reference文件。 ${samtools} faidx ref. So I was wondering that is it possible to generate transcript_alignments. fasta > output. As a result, I decided to change my command line to try and maximise the alignment : minimap2 -cx map-ont --secondary=no --sam-hit-only -A 3 -B 2 -t 3 ref. At least for now, I am only specifying this for minimap2 output, which I believe only has the following Minimap2 automatically tests the file type. 2,498 11 11 silver badges 29 29 bronze badges. 1 Align all reads to a reference. Ram RS. I'm going to leave the issue open, because SNAP should both be able to take input as to what should go in the read group as well as copying the information when the reads come from a SAM file rather than FASTQ. All reactions where a line starting with R gives regions covered by one query contig, and a V-line encodes a variant in the following format: chr, start, end, query depth, mapping quality, REF allele, ALT allele, query name, query start, end and the query orientation. fa fastq > file. asked Aug 10, 2022 at 20:46. . fa hifi. sam > 111. Just take all lines that contain your chromosome name (header+reads) along with the @HD and @PG lines in the header, then convert to sam: BAM files have a mandatory table of reference names and lengths, plus optionally an embedded SAM header which may repeat that information as @SQ lines (and can supplement it with extra things like MD5 checksums). Eventually I got an answer from the developer of minimap2 saying that I did it correctly and the output in the bam file is in the format is should be. lay. and still have the same problem. U I am working on Oxford nanopore reads(1D). Although another possibility would be to make your own header using samtools dict and then no header in SAM file Maxine ▴ 50 I use the minimap2 to do alignment for my pacbio long-reads DNA data against reference genome. bam . bam - I think this issue might come up a lot now because with ONT methylation, you must run minimap2 -y to pass on the full fastq header with methylation tags (MM and ML from memory). Create alignment stats # SAM headers INTERSPERSED in the output SAM file, making it unparseable. So, using -a do increase runtime for it. sam > aln. The usual approach I use for LAST mapping is as follows: These ERRORs are all problems that we must address before using this BAM file as input for further analysis. I checked the line of 13394306, nothing special there. h gives Saved searches Use saved searches to filter your results more quickly Allow the header from in. g. STR is the prefix name of the temporary files which the intermediate values are written. ADD REPLY • link 4. In this step each read is aligned against the reference, and its best aligning position found. Here is the command I use below: minimap2 -t 30 -2 -I 100g -ax map-pb genome. Minimap2 generates a multi-part index when the total length of The solution took me a while but is very simple: if you check the help message of minimap2, you will see that the reference should be provided first. That position, along with a metric of the quality of the single alignment is reported in a SAM format file. The sort_extra allows for extra arguments for samtools/picard Contribute to jguhlin/minimap2-rs development by creating an account on GitHub. I'm not sure of the line's exact position in the SAM file, but I printed out the lines prior and after the offending line and they are intact with a nucleotide string followed by what I understand to be Illumina information (perhaps pertaining to quality of the nucleotide call?) Okay . [main_samview] fail to read the header from "-". I will appreciate your suggestions. The optix gene codes for a transcription factor that plays a key role in the class SAM2PAF (ConvBase): """Convert :term:`SAM` file to :term:`PAF` file The :term:`SAM` and :term:`PAF` formats are described in the :ref:`formats` section. 2. IBSC_v2. Is this a missing header issue or a RA Hello, When I try to run fly, the sam header that is generated from minimap appears to lack sequence names. sam file contains the most common header records types you’ll encounter in SAM/BAM files. Each line must contain the reference name in the first column and the length of the reference in the second column, with one line for each distinct reference. samminimap2 -a -x map-pb test. ***> wrote: I also meet this problem -- samtools view: failed to add PG line to the header. To get the agat_config. View On GitHub; Getting help. js call" ignores Library. The sort_extra allows for extra arguments for samtools/picard $ minimap2 -t 4 -ax sr -p0 -N 6 -k 10 transciptome. samtools faidx reference. fasta > minimap. It could be resolved (by counting the number of tab-delimited fields in a line for example) - on the other hand SAM specification does not allow read headers to be started from '@'. 1. "For a multi-p -t FILE A tab-delimited FILE. To get bam from minimap2 use the following command: minimap2 -ax splice:hq genome. gz input. fa transcripts. sam Minimap2 works perfectly fine and end [INFO] minimap2 -t 4 --secondary=no -ax map-pb all_potential_contigs. minimap2 -x map-ont -a ref. otherwise it takes the orignal agat_config. A part of the sam file is as follow: Yeah we are, unfortunately, aware of that absolutely horrifying bug in minimap2. samtools view -bT ref. fna. I tried. Hi! I am also facing a similar problem, could you please suggest some fix? Thank you. This would be a nice workaround and you will be able to control the parameters. reference sequence. sam This command generated Sam files. Thanks Reading in data with read_paf. Similar problem, but different cause (and far more duplicated headers) When passing multiple FASTQ files on the command line, a new SAM header is inserted in the output stream once (AFAICT) for each input FASTA/FASTQ. my sam file doesn't have header. For other BAM files this step is skipped. sam has to be omitted. fa seq_C11737035 --split-prefix string > long_headers. And use samtools read the sam file with I am already putting the reference first and reads second: minimap2 --secondary=no --sam-hit-only -t 48 -a -x map-hifi assembly. NB @lh3 example above using grep to filter out SAM headers until later works a treat incase you dont have the memory to do the above (or can't be bothered). In that case it does mess up the header section of the resulting sorted bam file and gives a warning: [W::sam_hdr_sanitise] Missing trailing newline on SAM header. You signed in with another tab or window. MM_F_RMQ: 2147483648: Use RMQ for read mapping quality estimation. Both target and query are in fasta format. Examples includes bwa mem, minimap2, and bowtie2 (unless in --end-to-end mode). toplevel. MM_F_QSTRAND: 4294967296: Consider query strand in mapping. Output only alignments in SAM (no headers). sam [M::mm_idx_gen::27. 0 in centos Hi , I got an error when I trasnfered sam to bam file after minimap2 for my nanorpore data. It would be best to realign with minimap2 and choose SAM output. I used the following command: minimap2 -ax map-ont -t 20 Hordeum_vulgare. gz >111. This can be caused by: adapter sequences (aren't in the reference) poor quality bases (mismatches only make the alignment score worse) header_index: a dict they associate an index to a header name; reads_index: associate a read name and its length to an index; matches: basic. gtf -o countFile *. Notes. index input. If BAM files are used as input (with --bam), only reads in files without a reference in the SAM header are aligned. with samtools view -b -T ref. g mikolmogorov/Flye#48). Getting started. sam -L Both the GTF file I am using, and the reference genome that the alignments were generated off of, are from UCSC table browser, HG38, knownGene, GENCODE v36. Thank you for your help! I initially thought the header wouldn’t be an issue since previous files ran perfectly without any problems. However, this appears not to be the This option makes minimap2 SAM. I saw that in file process_minimap2_alignments. I made an index file for the human genome to be used as a reference. It provides C APIs to build/load index and to align sequences against the index. py but it fails. The sample information is given only in the bam/sam header and not in each read. PAF against jPAF. That's because If I'm not mistaken StainedGlass breaks the input fasta into lots of smaller chunks that it maps back to. Unfortunately, I am busy with another project. fa in. paf Set index parameters for minimap2 using builder pattern Creates the index as well with the given number of threads (set at struct creation). If present, the header must be prior to the alignments. This option has no effect if seq is set. The sequence name will be set to N/A. 4. Li: You can count the number of gaps from CIGAR. I initially assumed this meant N from the CIGAR, however, when I took NM-N I obtained an array with negative values, so clearly I had been mistaken. I have used tried these commands: minimap2 -a -x map to map ONT reads to reference but have recently encountered the issue of bam not being binary as well as missing headers. my minimap2 code is : minimap2 -ax sr . 3. Nanopore: paf paf. preset: minimap2 preset. You could simply realign with this parameter and directly pipe into samtools to get a sorted BAM file. I will look into it . Summary: Not a bug report, not a feature request, not a documentation requst. sam I had to use the -I8g to get any output with sam header. The input and output files are specified accordingly, with the output being directed to SAM or PAF files. Although head works to take a quick peek at the top of a SAM file, keep the following points in mind: head won’t always provide the entire header. For example, MISSING_READ_GROUP errors This leads to SAM parsing issues (e. fastq > data. Based on samtools/samtools#1613 I'm assuming that that produces too many headers to fit into the bam file properly. bam [I] samtools mpileup /tmp/panphlan_3_46v4dq. Faroll Faroll. You signed out in another tab or window. You can set it to #!/usr/bin/python3: import argparse: from collections import defaultdict""" This utility converts a SAM file to a nucmer delta file. Minimap2 is a powerful tool that has revolutionized the way we align sequences in bioinformatics. bam samtools view -H sample. I did : minimap2 -cx map-p Download Citation | New strategies to improve minimap2 alignment accuracy | We present several recent improvements to minimap2, a versatile pairwise aligner for nucleotide sequences. h gives more detailed API documentation. The extra param allows for additional arguments for minimap2. I have been trying to use it but I don't seem to get past mapping. seq: a single sequence to index. sam In the manual is used to add @rg format -R STR | SAM read group line in a format like @rg\tID:foo\tSM:bar []. sam_parse1] no SQ lines present in the header #944. Minimap2 aims to keep APIs in this header stable. what should i do in this situation? Thank you very much. Header file minimap. Newer tools recognizes this tag and reconstruct. fasta query_sequence. 3331. Not sure if it will do what you are looking for: Building a bam file and vcf file from the alignment SAM file generated by minimap2 produces 1 sample i. If I use minimap2/2. I have used I have attempted outputting an intermediate SAM, but the issue persists. fastq. Minimap2 is a fast sequence mapping and alignment program that can find overlaps between long noisy reads, or map long reads or their assemblies to a reference genome optionally with In this case, minimap2 is unable to output a proper header. /111. I use the command minimap2 -t 30 -2 -I 5g -ax map-pb genome. sam" samtools sort: failed to read header from "20201032. [W::sam_hdr_create] Duplicated sequence 'CZBZ01000008. 22 it goes incredibly slowly for a 24Gb fastq that should be mapped relatively faster, and haven't manage to I am using the version 1. gz -fo dbg. So I apologize for the "free-format" report. However, I realised that at times (often) I had multiple primary alignments (see attachment). fa sample_name. gz | samtools view -S -bT reference. /4PF-0-1 I use the minimap2 to do alignment for my pacbio long-reads DNA data against reference genome. However, the length of the target is retrieved from the @SQ line that must be present. bam. fasta - > nanopore_reads_minimap2_mapped. fa # 建索引 ${samtools} view -bS -t ref. The input bam is a Promethion run mini bams merged via samtools merge into a single bam (140GB). xz Perhaps it isn't writing a sam file but something biostud1819 • I'm running minimap2 the command is: minimap2 -t 40 -ax splice -k14 -u f --secondary=no input. the software dependencies will be automatically deployed into an isolated environment before execution. sam One hinderance to this output is that (I believe) SAM format cannot be specified as the input to minimap2. Copy link Member. fa gbk. bam @SQ SN:utg000001l LN:5073047 @SQ SN:utg000002l LN:24568116 @SQ SN:utg000003l LN:22903238 @SQ SN No, it's not the last line in output. from minimap2. bam I am getting To cut a long story short, it looks like in some circumstances the SAM header gets spread out in chunks throughout the SAM file - almost like there are multiple SAM files concatenated together. Value to be consistent with the header RG-LB tag if @RG is present. francois Currently paftools. I used the bam file in GATK and there was no problem. I used the following command: minimap2 -t 20 -ayYL --MD --eqx -x asm20 ref. long_ref_headers seq_C11737035 long_headers Ask away! I'm using this workflow on our HPC via Singularity. fastq > aligned_reads. [sam_read1] missing header? Abort! So I wanted to know how did you do it . These are the commands I used: minimap2 -t 40 -ax map-ont reference. Improve this question. sam See here for more detailed information on the Minimap2 command. bz2 paf. So is there a way to turn off the SAM aux format parsing, or do I need to write alternative processes in my nextflow alignment pipeline to align either standard fastqs Why would your SAM file not have a header? Did you not use-a output in the SAM format when you did the alignments? If you did not, then your alignment output is in PAF format (which is the default). coords and . but when I am converting sam to bam samtools view -bS 111. 6 years ago. I mapped my read using the -c / -a option for obtaining the same alignment, but with a different format for the output. This document assumes you’ve already ran a sample an ONT Nanopore device and have generated fast5 files recording the squiggles of that run. Samtools is just complaining about a missing input file: [main_samview] fail to read the header from "-". In addition to some detailed information added in the header and footer, the following situation will happen-Some information is missing best wishes! The text was updated successfully, Well, sometimes sam headers do have spaces because the mapping tool put them there from the corresponding fasta header. I would like to skip the SAM file generation for performance and resource considerations, and pipe the minimap2 result to samtools sort in order to directly generate a BAM file. The main reason for this is each mapped BAM read doesn't store the reference name, but the index of the reference. samtools sort: failed to read header from "20201032. compatible with older tools. sam that it matches to only either one of them. lh3 commented on May 26, 2024 3 . In this case, minimap2 doesn't output any SAM header. Posted on May 15, 2018 by Thomas Cokelaer. fa align. Yes this is helping the problem is that the genome is too big for a normal GMAP build and run. Most ERRORs can typically be fixed using Picard tools to either correct the formatting or fill in missing information, although sometimes you may want to simply filter out malformed reads using Samtools. I have already tried mapping the reads with minimap2 a I was expecting that, when running minimap2 against the same input files, the alignments output into paf (ie without the -a option) would be the same as those output into sam (ie using the -a option). sam designates the minimap2 output with then SEQ, QUAL fields replaced by a '*'. To make sure the header is correctly written, I validated the file again. How does minima minimap2 -cx map-ont ref. The problem is that bwa mem is failing to produce a SAM file because the reference directory specified using params. bam both errors are: fail to read the header from sample. I've just updated the README to mention the bug in minimap2. The logic in Daijin unfortunately does not support providing custom command line arguments to However, implementing this feature would bring a heavy htslib dependency, which is larger and more difficult to compile than minimap2. sam > sample. (SQ lines) in the SAM header. no header in SAM file Maxine ▴ 50 I use the minimap2 to do alignment for my pacbio long-reads DNA data against reference genome. When using minimap2 to map sequencing reads onto a reference, you can use this kind of command (be careful, this is wrong as you will see later): minimap2 -a-x map-pb test. bam) as a post step. sample_name. gz -I8g > output. I have . This seems to be causing errors downstream: root@flye:/tank# Hi, When I use the command line: minimap2 -t 10 -ax splice --split-prefix -uf --secondary=no sequences. SAM files must contain an NM tag, which is default in minimap2 alignments. Now minimap2 1 The SAM Format Specification SAM stands for Sequence Alignment/Map format. Then, I try to run your workflow on m Hi, First of all: great tool to visualize the rearrangements! It works all perfectly fine as long as I use Mummer for the alignment, and the . sam >head. The same gencode hg38 annotation file is used for the minimap2 alignment and is the indexed miso file as well, so you wouldn't necessarily expect a header mismatch. In addition, a report and two charts are generated with complementary information. I end up with several question Skip to content. gz paf. View the Project on GitHub . yaml shipped with AGAT. It is a TAB-delimited text format consisting of a header section, which is optional, and an alignment section. Follow edited Aug 10, 2022 at 22:04. The version in This is just an introduction to the basics of the SAM format’s header section; see the SAM format specification for more detail. It won’t work with binary BAM files. fa reads. Methods used in this tutorial include: minimap2 - to create alignments of a long-read sequencing dataset, samtools - to inspect and filter SAM and BAM files, and I have aligned the Nanopore reads to reference genome command: minimap2 -ax map-ont -t 8 /ref . , the chromosomes if you’ve aligned to a reference genome). klein ▴ 30 4. sam The header of my sam: @sq SN:Chr1 LN:43270923 @sq SN:Chr2 LN:35937250 @sq SN:Chr3 LN:36413819 @sq SN:Chr4 LN: Hi, I have mapped my data to a large genome. See here for more info): On Tue, 16 Jan 2024 at 07:17, Enosh ***@***. In this tutorial we will align a piece of chromosome of two Heliconius butterfly species that includes the optix gene into a pan genome alignment. I am aware that full SAM format support may be out of scope, but hoping we might open some discussion Hi, I had a problem when repeating alignment in sam output. fasta Hi, thank you for the tool. Skip to content. fa > CI5791alnMorex. The tutorial presents an example using two haplotypes of the optix locus in Heli Eventually I got an answer from the developer of minimap2 saying that I did it correctly and the output in the bam file is in the format is should be. And is it possible to run funannotate predict using Previously generated misc files? (To skip the time-consuming steps such as parsing soft missing SAM header with minimap2 and samtools Posted on May 15, 2018 by Thomas Cokelaer When using minimap2 to map sequencing reads onto a reference, you can use this kind of command (be careful, this is wrong as you will see later): minimap2 -a -x map-pb test. the real CIGAR in memory. I have the sam file created by bwa. MM_F_NO_INV: Hi, I am mapping pacbio reads using minimap2-2. e. The sam file was created with minimap2 version 2. delta files. Hello Im using next command minimap2 -ax -R map-pb -I8g -d 5Gb_genomefile. sam designates the minimap2 output in sam format, short. The file size becomes large because in SAM/BAM each line contains query sequence, but agat_convert_minimap2_bam2gff. TP Mapping quality, flags, headers and the number of secondary mappings are the quantities that are recovered. sam how can I create bam file Hi, I followwd the instruction minimap2 -ax map-ont ref. I used the ValidateSamFile tool from Picard to get the different RG tags in the alignment section. To use bam with this script you will need samtools in your path. It turns out that your scripts outputs a sam file without sam header. fa ont-reads. Both the GTF file I am using, ERROR: Unable to find chromosome 'hg38_knownGene_ENST00000407684. paf. Only the @pg line is present in the file minimap. However, I’m currently using output from Prodigal, which generates headers minimap2 does not do basepair alignment without the -a option (as PAF does not output that). It is not that difficult to parse a BAM stream. fa When using minimap2 to map sequencing reads onto a reference, you can use this kind of command (be careful, this is wrong as you will see later): minimap2 -a -x map-pb test. fasta . Most short read aligners perform local alignment of reads to the reference genome. Hi, I'm using minimap2 for WGS with the GRCh38 reference. CMD is passed to the system's command shell. fastq Map query sequences to targets and output SAM file. sam" You may have been intending to pipe the output to samtools Hello, I have been struggling with running samtools because the program can not read the header of my sam file so i get the following error: samtools sort: failed to read header from "20201032 Skip to content. rule dict: input: contigs = os. Rust bindings to minimap2 library. Newer tools recognizes this tag and reconstruct the real CIGAR in memory. TP. Hi @wwood,. sam -o head. -I 64g), if you have access to a machine with lots of RAM, or alternatively to use another aligner. flt. So formally, minimap2 is producing an incorrect SAM, given those reads as input. bam > A versatile pairwise aligner for genomic and spliced nucleotide sequences - Issues · lh3/minimap2. sam -L. minimap2 -p 28 -ax map which was useful to create a SAM header from a chrom. This means the ends of the read may not be part of the best alignment. fa head. bam [main_samview] fail to open file for reading. PAF is a plain text tabular format where each row represents an Ultrafast de novo assembly for long noisy reads (though having no consensus step) - miniasm/PAF. Using a recent samtools, you can however coordinate sort the SAM and write a sorted BAM using: samtools sort -o "${baseName}. gff3 using the bam file I generated, and pass the gff file to funannotate predict to skip the minimap2 alignment step?. sam sample. Description: The header of the SAM file (lines starting with @) are dropped. fq > out. Parameters: targetfile (str) – Alignment targets (reference), typically FASTA (can be gzipped). fa | samtools sort -O BAM -o output. This is sent to the file Minimap_erato_melp. The tutorial is intended as a gentle introduction to Sequence Alignment/Map (SAM) formatted files and their binary equivalents BAM. # To work around this mind-boggling bug, we remove all header lines from # minimap2's SAM output by grepping, then re-add the header created in this # rule. CMD must take the original header through stdin in SAM format and output the modified header to stdout. However, when I try to use a . Because minimap2 is a minimiser-based mapper, it's less useful for determining mapping accuracy to single-base precision. You should convert your bam to sam first, update the header, then convert back to sam. fa. js sam2paf cannot handle spaces in fasta headers. HiFiMapped. Add MD Tag to BAM: Header lines start with ‘@’, while alignment lines do not. sam It seems to aligned all the reads just no header All reactions I had a small number of RG tags in each BAM file. An improvement possible to MarkDuplicates is to validate headerlines before doing the hard work of actually markin Input reads are aligned against the combined reference with Minimap2. fasta CI5791_rawreads. path. So the top command should when I used minimap2 to map Illunima paired reads to assemebled contigs, the header of the sam file seems to be wrong. 62] collected minimizers [M::mm_idx_gen::3 Hello,everyone: I have used minimap2 to map pb ccs reads to a genome assembly by hifiasm. 3. fa aln. The sort param allows to enable sorting (if output not PAF), and can be either ‘none’, ‘queryname’ or ‘coordinate’. 0 years ago Not a duplicate of issue #15. I NUM. Not a direct answer, but if you have access to the files used to generate the PAF, you can use them and generate sam files with minimap2. In each of these commands, -ax specifies the preset for different types of reads, and -x specifies the preset for the overlap layout. The text was updated successfully, but these errors were encountered: All reactions. Generally, you should only look at variants where column 5 is one. Dorsal (top) and ventral (bottom) sides of Heliconius melpomene rosina (left) and Heliconius erato demophoon (right). Each alignment line has 11 mandatory Minimap2 is not only a command line tool, but also a programming library. fa N006_merge. sam" srun: error: node2-069: tasks 0-3: Exited with exit code 1 [E::hts_open_format] Failed to open file "20201032_sorted The celegans. 10 and gcc 5. gz") Hello, This the first time I am using Cupcake to handle IsoSeq data. fixed. longer500. bam to be processed by external CMD and read back the result. fa Asecodes_parviclava. I may not be able to implement this feature soon. Then I manually wrote the header files and added the new header to the existing bam file using the samtools reheader tool. mmi index file I've built using minimal2 and input One thing to try is piping it into less to see if the header is missing A versatile pairwise aligner for genomic and spliced nucleotide sequences. Closed theo-allnutt-bioinformatics opened this Hi Dr. After getting the SAM file, I usually count using HTSeq, but there are some format issues when using the minimap2 SAM file. So I was using samtools to convert sam to bam but I am getting same answer as you have got [samopen] no @SQ lines in the header. collect the sequence names and the lengths of all dumps to generate a better SAM header (SQ lines) Minimap2 is not only a command line tool, but also a programming library. Guide through code for a Minimmap2 genome alignment and a seq-seq-pan pan-genome alignment with visualizations in R. Unless a file or directory is specified using the input path qualifier, Nextflow will not know to stage the Samtools sort will work on a Sam, no need to convert to . p_ctg. And use samtools read the sam file with Samtools sort will work on a Sam, no need to convert to . fa # polish consen The best (and likely fastest) solution would be to use the minimap2-I option and give minimap lots of memory, as suggested in the FAQ. /. 1 years ago by GenoMax 148k 5. fastq > Hi, I have seen the other closed issues in this topic, but the solution does not work. There Introduction to SAM and BAM files. Entering edit mode. which finished correctly. I want to know all possible transcripts The procedure will be definitely cleaner and perhaps even simpler than manipulating SAM/PAF from the current multi-part minimap2 output. fasta | samtools view -@ 4 -b -F4 -F 0x800 -q 0 -o HiFi-vs-potential_contigs. When used in this manner, the external header file in. and I use bowtie to index my reference index file. 14-r883: minimap2 -ax map-ont long_ref_headers. By default, when calling variants, "paftools. Reply reply More replies. Hi, I’m trying to better understand how minimap2 assigns the primary and secondary alignments and how these relate to the primary and secondary mapping. I added an example reference, reads and resulting sam file below. ADD REPLY • link 24 months ago by susan. sam. The workaround is to either increase the memory consumption of the minimap2 index (e. I used the lattes version and the version 0. yaml locally type: "agat config --expose". Header lines start with ‘@’, while alignment lines do not. bam Reply reply guepier • and -h in conjunction with -b makes no sense, since -b always writes a header anyway. I minimap2 option1 option2 genome. sam > out. fa looks like this, it has more than 500 headers, I see in my . A versatile pairwise aligner for genomic and spliced nucleotide sequences - Issues · lh3/minimap2. There is also this evaluation method inside minimap2. SAMv1 All the SAM records in a chimeric alignment have the same QNAME and the same values for 0x40 and head -13394305 xxx. bam" "mapped_${baseName}. You have to manually add it (e. I'm trying with window=64 right now but I'm wondering if there's a way to split those alignment steps per chromosome so each individual [E::sam_hdr_create] Invalid header line: must start with @HD/@SQ/@RG/@PG/@co However, I have checked many times about the header of sam files, which are fit to the format. head -13394306 xxx. 1' [E::sam_hrecs_update_hashes] Duplicate entry "CYYQ01000001. dna. SAM Flag; SAM Flag (single) Base Quality; Header; Alignment Tags; Ambiguity Codes The header line The first line if present. Sign in Product 2. sam files aligned with minimap2, and am running the following command to try to get a count table: featureCounts -a knownGene_v36. Methods used in this tutorial include: minimap2 - to create Given a reference longer than 4Gb, minimap2 is unable to see all the sequences and thus can't produce a correct SAM header. The sequence file can be optionally gzip'd. You switched accounts on another tab or window. bam # 输出转换结果 # 或者 ${samtools} view -bS -T ref. samtools ref. The | cut -f 1-12 will only take the first twelve columns of the Minimap2 output. [rehab@iria N006-minimap]$ ~/minimap2/minimap2 -a human-refseq. We incorporated multi-index merging into the Minimap2 aligner and demonstrate that long read alignment to the human genome can be performed on a system with 2 GB RAM with negligible impact on accuracy. The output should look like this (without the column titles as shown here. You always get a uni-part index if -I is larger than the total lengths of all sequences. sam ## to add headers from the reference and being able to produce a bam file. fasta nanopore_reads_raw. ctg. Top 2% Rank by size . 18: H0: i: Number of perfect hits: 19: H1: i: Number of 1-difference hits (see also NM) 20: H2: i: Number of 2-difference hits: 21: HI: i: Query hit index, indicating the alignment record is the i-th one stored in SAM: 22: IH: i: Number of stored alignments in SAM that My header seems OK (see below), but I am new at this. fa to reference genome. Consider this SAM file with two alignements only. align_ref does not exist inside the container. fai aln. 1" in sam header samtools view: failed to add PG line to the header samtools sort: failed to read header from "-" [I] samtools index /tmp/panphlan_3_46v4dq. when I started the alignment I got following warning. sam #to see if there was anything wrong with the SAM headers This option makes minimap2 SAM compatible with older tools. 1' in the Hello, I have had success using your workflow for several jobs but now I try to run it on a concatenated version of my samples. SYNOPSIS# 1. sam > joelle. This is the main step, and with minimap2 it can be accomplished with a single command-line. pl# DESCRIPTION# The script converts output from minimap2 (bam or sam) into GFF file. I am trying to collapse HQ isoforms from IsoSeq reads (FASTA file) with collapse_isoforms_by_sam. samtools view: failed to add PG line to the header". I am now assuming that you Minimap2 is not only a command line tool, but also a programming library. daviesrob commented May 25, 2022. sizes file. join(OUTDIR,"contigs. fasta in sam header samtools view: failed to add PG line to the header. sam ## to remove the headers samtools view -b -T genome. c demonstrates typical uses of C APIs. Reload to refresh your session. So I think it is worth pointing out. Users must manually add SQ lines to the header for compatibility with downstream analysis tools. fa > N006_sam_alignment. I created a conda environment with your yml file that is inside MitoHiFi/environment after the MitoFinder was installed and made accessible via the PATH environment variable, so the version of samtools is 1. Converting SAM to BAM with samtools view [W::sam_hdr_create] Duplicated sequence The above will now avoid the 'concatenated' SAM file effect. queryfile (str) – Queries to align, The tutorial is intended as a gentle introduction to Sequence Alignment/Map (SAM) formatted files and their binary equivalents BAM. For example: missing SAM header with minimap2 and samtools. If a sequence file is provided, minimap2 builds an index. #What is a SAM "record"? A record is a single line in a SAM file, and it generally corresponds to a single read, minimap2 -a reference_genom. sam . You must set the number of threads before calling this function. header. fq. Questions You can have one primary alignment in the SAM file format. sam # for Oxford Nanopore reads for my nanopore data and produced the sam file. mlwsq hnxgvv gap wqhmhrk vfjt ebmrz omiez fblyo feqp rpvkjt