Tag: flagstat

Randomized phase II study of preoperative afatinib in untreated head and neck cancers: predictive and pharmacodynamic biomarkers of activity

Study objectives and endpoints The main objective consisted in identifying predictive biomarkers of efficacy by exploring correlation between baseline potential biomarkers and radiological and metabolic responses to afatinib. Secondary objectives were to identify potential pharmacodynamic biomarkers, to evaluate the efficacy and safety of afatinib and to assess the metabolic and…

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Extracting chimeric reads from mapping

Hello, I am struggling to processing and analyse bam files (from bwa alignment), to extracting the chimeric read alignment. I am aligning human cell line RNA-seq data (paired end) to virus, aimed to find the viral integration sites in the genome. For that, after reading a bit here from following…

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Longitudinal detection of circulating tumor DNA

Analysis of Roche KAPA Target Enrichment kit experimental data obtained on an Illumina sequencing system is most frequently performed using a variety of publicly available, open-source analysis tools. The typical variant calling analysis workflow consists of sequencing read quality assessment, read filtering, mapping against the reference genome, duplicate removal, coverage…

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Filtering for primary and secondary reads using sam flags (0 properly paired reads in alignment step)

Hey everybody, I have just performed the alignment of paired-end reads to a reference using bwa mem with the -M flag, ran samtools markdup and flagstat. Flagstat produced the following output: 4985084 + 0 in total (QC-passed reads + QC-failed reads) 1806492 + 0 primary 3178592 + 0 secondary 0…

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mRNA vaccine quality analysis using RNA sequencing

Design and synthesis of reference plasmid A reference construct was first designed, with the intention of optimising the production of RNA therapeutics for pre-clinical research. The coding sequence of eGFP30 was selected as a reporter in the coding region, as its protein product can be assayed simply through Flow cytometry…

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Confused about % of mapped and unmapped reads output from STAR aligner

I am quite new to STAR aligner, and have some confusion in the numbers of unmapped/mapped reads output from STAR: I would like to know whether the STAR output BAM file if I do not use the argument (–outSAMunmapped within) is already filtered for unmapped, or duplicate reads or not…

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Blocky p300 ChIP-seq reads

I recently performed a p300 ChIP-seq on two different genotypes. I aligned my reads using STAR aligner, and made UCSC .bedGraph files that I loaded on IGV viewer. I noticed that the reads for one of my genotypes – genotypeX (red – combined replicates; purple and pink – individual replicates)…

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Bioinformatics Workflow

Bioinformatics Workflow 1 Hi everyone I am doing bioinformatics and computational biology Thesis project on genetic Characterisation of Plant Clones to a reference genome. The aims are to Read map all clones to the reference genome, Identify accumulated somatic mutations over time, compare all the 20 clones with each other…

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Exome sequencing identifies breast cancer susceptibility genes and defines the contribution of coding variants to breast cancer risk

UKB The UKB is a population-based prospective cohort study of more than 500,000 subjects. More detailed information on the UKB is given elsewhere34,35. The study received ethics approval from the North West Multi-center Research Ethics Committee. All participants signed written informed consent before participating. WES data for 450,000 subjects were…

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Nuclear genetic control of mtDNA copy number and heteroplasmy in humans

Overview of mtSwirl Here we develop mtSwirl, a scalable pipeline for mtCN and variant calling which makes calls relative to an internally generated per-sample consensus sequence before mapping all calls back to GRCh38. In addition to GRCh38 reference files and WGS data, the mtSwirl pipeline takes as input nuclear genome…

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What are bitsets in Samtools flagstat output?

What are bitsets in Samtools flagstat output? 0 Hello, I am am trying to use Samtools flagstat to analyze my BAM file after aligning nanopore dRNAseq reads to a reference transcriptome using minimap2. The output file indicates I have the following flagstats below (excluded zero values from output). 729779 +…

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Why extracting reads from BAM to fastq produces small amount of reads?

Hi, I got a peculiar issue with one of my datasets – when I try to extract reads from a BAM file (which has 20 mil reads) to a fastq – I get only ~400k reads. Here is what I do: I first sort the bam by name: samtools sort…

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Substantial reduction in read mapping following VG surject

Substantial reduction in read mapping following VG surject 0 I did a small comparison of total mapping of a small short-read dataset to: a graph constructed using cactus-minigraph composed of eight whole genome alignments using VG giraffe the same cactus-minigraph graph, but indexed using VG and mapped to with VG…

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Calculating Average Coverage or Read Depth for a Sequence (WES)

Calculating Average Coverage or Read Depth for a Sequence (WES) 0 Hi, I might as well ask this community as well just to be certain Background: I mapped my reads to reference genome, then used SAM tool flagstat and SAM tool stats. Felt like using both but at the same…

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group samples by population + boxplot ?

multiqc: group samples by population + boxplot ? 1 Hi All, is there a simple way to tell multiqc to group data by population ? for example, say I’ve got the output of samtools flagstat for 50 cases and 50 controls, how can I get a boxplot that would display…

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BAMboozle

BAMboozle 1 Hi, I am running BAMboozle to anonymize variant sequences using the GRCh37 human reference genome on my bam files. My bam files originally are 2-3 GB but when I get the output bam file from BAMboozle it is 500-600 Kb. Does BAMboozle decrease the size of the bam…

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High number of duplicates and low percentage properly paired

High number of duplicates and low percentage properly paired 0 I have some paired end sequencing data that I have trimmed using cutadapt. It was sequenced on an illumina novaseq 6000 and is low coverage RADseq data (2-3x). My cutadapt script used forward and reverse adapters from illumina : cutadapt…

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Comparing Alignment Files (CRAM)

Comparing Alignment Files (CRAM) 0 Hello all, Just checked different forums and generally, I see that it would be useful to use samtools or picard-tools for comparing alignment files. Here I want to compare the aligned output files using two different alignment algorithms. In this case, I had some general…

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10x Cellranger SN RNA-seq BAM file output looks off

I’m working with SN RNA-seq data that I processed using Cellranger v. 7.1.0 (which uses STAR for alignment with default settings). Using the samtools flagstat command to look at a BAM file output: 284890368 + 0 in total (QC-passed reads + QC-failed reads) 0 + 0 secondary 0 + 0…

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Nextflow rnaseq finishing early

Nextflow rnaseq finishing early 0 Hi I’m running the RNA-seq pipeline from nextflow and I have been running it without problems until this dataset it just stops prematurely saying it has finished when it doesn’t even aligns the reads with salmon. Any ideas what may be going on? I have…

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Wildcard error in Snakemake – clarification on inputs

Error: Building DAG of jobs… WildcardErrorin line 502 of /path/to/pipeline/workflow/Snakefile.py: Wildcards in input files cannot be determined from output files: ‘anc_r’ Code: import os import json from datetime import datetime from glob import iglob # ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Define Constants ~~~~~~~~~~~~~~~~~~~~~~~~~~~~ # # discover input files using path from run config SAMPLES…

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reads aligned concordantly exactly 1 time

Good evening, I’d like to compare the alignment quality of hisat2, bowtie2 and bwa for my files. The first 2 packages output the percentage of reads aligned concordantly exactly 1 time, bwa does not, because does not output alignment summary. The samtools flagstat report is not enough, because it outputs…

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Can’t convert paired end BAM to bed using bedtools

Can’t convert paired end BAM to bed using bedtools 0 Hello, I got .bam files from my pipeline and i merged them with samtools merge ALL.bam *.bam and I got this % of mapping 3825773 + 0 in total (QC-passed reads + QC-failed reads) 3825773 + 0 primary 0 +…

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TEMPERATURES Page Grand Canyon 45 Window Rock Flagstat! 46 Winsiow 57 Payson Show Low 54 FOX10

memepedia log in Animals & Nature Anime & Manga Art & Creative Cars Celebrities Gaming Girls Internet Memes Movies Other Politics Science & Tech Sports TV shows log in add meme new featured top memes memes catalog Animals & Nature Anime & Manga Art & Creative Cars Celebrities Gaming Girls…

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Samtools Convert Sam To Bam With Code Examples

Samtools Convert Sam To Bam With Code Examples In this session, we’ll try our hand at solving the Samtools Convert Sam To Bam puzzle by using the computer language. The code that follows serves to illustrate this point. # Basic syntax: samtools view -S -b sam_file.sam > bam_file.bam # Where:…

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Differing number of reads for read 1 and 2 in fastq’s from a subset bam file

I’m working with paired-end wgs data I downloaded from TCGA. I’m trying to extract the reads that align to a specific region, extract only those reads to two fastq files, one for each pair. Unfortunately, I am getting a different number of reads in both fastq files because some of…

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different result using minimap2 and pbmm2

Hi all! I am analysing CSS Pacbio data and each sample came from different run, in particular I have three files for each sample. I tested both pbmm2 and minimap2 to align my long reads, after getting the consensus sequences. This is the command I used to run mnimap2: minimap2…

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pjotrp/sambamba – sambamba – Genenetwork

10 years ago ​ 10 years ago ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ 10 years ago ​ ​ ​ ​ ​ ​ 10 years ago 10 years ago 10 years ago ​ ​ 10 years ago ​ 10 years…

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I wonder if someone please explain what secondary, supplementary, duplicates and paired in sequencing mean in samtools flagstat

I wonder if someone please explain what secondary, supplementary, duplicates and paired in sequencing mean in samtools flagstat 0 ”194492 + 0 in total (QC-passed reads + QC-failed reads) 80 + 0 secondary 0 + 0 supplementary 0 + 0 duplicates 193804 + 0 mapped (99.65% : N/A) 194412 +…

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Samtools flagstat confusing result of a merged bam file

Hi, I am a bioinformatics student and I am struggling with an issue, I had paired-end fastq files for one sample with some low-quality bases at the end and adapter contamination, so I went and I trimmed my reads with trimmomatic, it gave me 4 files that I used for…

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Overestimation of number of reads from nanopore data (flagstat)

Same issue as mentioned on the minimap2 tool: github.com/lh3/minimap2/issues/236#issue-361097444 For example nanopore reads aligned to the host transcriptome the flagstat output is: 5953480 + 0 in total (QC-passed reads + QC-failed reads) 2961480 + 0 secondary 22696 + 0 supplementary 0 + 0 duplicates 4195469 + 0 mapped (70.47% :…

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Samtools flagstat

Samtools flagstat 1 I aligned my ONT sequencing run with minimap2, subsequently I filtered the file using samtools view -b -F 256 aln_transcriptome_sorted_6.bam -o filtered_aln_transcriptome_6.bam to end up with primary alignments only. When I run samtools flagstat on the filtered file I get the following output: 3502608 + 0 in…

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Alignment report

Alignment report 0 Hi Guys, I did alignment of R1 and R2 fastq files with reference genome using bwa mem and got bam file. Now, I want to check whether the alignment is done correctly and alignment percentage,coverage etc. I run following command: bwa mem hg19.fasta R1.fastq R2.fastq | samtools…

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Attempting to generate a bam.bai file but the output is not readable

Attempting to generate a bam.bai file but the output is not readable 1 Hi, I am new a exome sequencing, and have tried to follow tutorials on the subject. I am stuck at the samtools index stage because the output files are in a non-human readable format and I believe…

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Problem with using flagstat after bowtie2 alignment

I’m running bowtie2 to align multiple samples to one reference genome, and then run samtools flagstats to output the results. All but two samples have aligned and I’ve managed to run flagstat on them. For those two samples, when I run flagstat, I first get: [W::bam_hdr_read] EOF marker is absent….

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Doubt samtools flagstat

Doubt samtools flagstat 0 I’d like to see the percentage of how many sequences align with my decrementing sequence and I’ve come to this sample table. But wanted to know what use? The mapped (80.94% : N/A)or properly paired (0.06% : N/A) percentage? 1036193 + 0 in total (QC-passed reads…

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MAPQ (Mapping quality) of 0 for most reads from BWA-MEM2 (with no secondary alignment or other apparent reason)

Hello, I got a very weird output from BWA-mem2 – most of the reads have mapping quality of 0, even though there is no secondary alignment or anything else suspicious. I got sequencing data that was aligned with Novoalign to hg18, the data was bam files. I needed to realign…

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very low coverage when mappin genomic DNA

very low coverage when mappin genomic DNA 0 Hi all, I’m having terrible problems to map/align single end RNA files from human genome (GRCh.38). It is genomic DNA but was prepared by using a RNA library kit to preserve strand specificity. I’ve first tried STAR and Kallisto and the coverage…

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