Tag: HTSeq-count

smallRNA profiling using HTSeq error

smallRNA profiling using HTSeq error 1 Hello, I want to create a “count” file using HTseq. I have both BAM file and gtf file: htseq-count -f bam -s no -i AK1a_clean_Aligned.sortedByCoord.out.bam gencode.v42.chr_patch_hapl_scaff.annotation.gtf >> AK1a_counts.txt It still gives an error: htseq-count: error: the following arguments are required: featuresfilename Can someone please…

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HISAT2 and HTSEQ command

HISAT2 and HTSEQ command 0 @4fedfa78 Last seen 10 hours ago Japan Hello, For analysis of the data by rna-sequencing I selected HISAT2 and HTSeq-count for mapping and counting the genes levels, the libraryLayout is paired, I am using the below command for both but the results are not exact…

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Bash script to automate htseq-count

Hi everyone- I am trying to write a script to automate htseq-count on a large number of samples. The script runs but then throws the following error: “Please provide 2 arguments”. Does anyone see something obvious I am missing: #!/bin/bash for samples in *.sam do gtf = “Galaxy135-\[Escherichia_coli_str_k_12_substr_mg1655.GCA_000005845.2.29.gtf\].gtf” echo $sample …

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can gff2 reference used in htseq-count?

Dear all We are recently working with E.coli plasmid and tried to summarize the gene counts from our RNA-Seq samples. The short reads were mapped to E.coli plasmid using tophat which generated bam files accordingly. However, we were unable to obtain a gff3 version of our target plasmid genome, the…

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HTseq-Count: Long processing time

HTseq-Count: Long processing time 1 Hi everyone, I’m processing BAM files using htseq-count and it takes very long time to produce the counts for each file. It is about pair-end reads (around 50 million sequence each). It takes 75 minutes to count this pair; is that normal? Thanks. htseq-count –max-reads-in-buffer=24000000000…

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htseq-count error

htseq-count error 1 Hi, htseq-count -f bam -s yes ~/htseq-trial/SRR13826419_Aligned.sortedByName.out.bam ~refgen/gencode.v39.primary_assembly.annotation.gtf > counts.txt I am trying to run htseq-count with command above but in the err file [E::idx_find_and_load] Could not retrieve index file for ‘~/htseq-trial/SRR13826419_Aligned.sortedByName.out.bam’ 100000 GFF lines processed. 200000 GFF lines processed. 300000 GFF lines processed. 400000 GFF lines…

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Feature count is very low using htseq-count

Feature count is very low using htseq-count 0 Hello all, I performed bbmap on my RNA-seq paired sequence data using following cmd bbmap.sh in1=J2_R1.fastq in2=J2_R2.fastq out=output_J2.sam ref=im4.fasta nodisk The header of generated sam file is @HD VN:1.4 SO:unsorted @SQ SN:k141_1006 LN:2503 @SQ SN:k141_5512 LN:5393 @SQ SN:k141_4772 LN:4387 @SQ SN:k141_3267 LN:4531…

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Htseq is giving me 0 counts using the GFF3 of miRBase

Hello! I am trying to annotate a miRNA-seq so that it gives me mature miRNAs where I already have 5p and 3p. For this, I have used the index mm10.fa and the miRBase mmu.gff3. I have aligned with HISAT2 and am trying to count with HTSeq, however I get 0…

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The role of ATXR6 expression in modulating genome stability and transposable element repression in Arabidopsis

Significance The plant-specific H3K27me1 methyltransferases ATXR5 and ATXR6 play integral roles connecting epigenetic silencing with genomic stability. However, how H3K27me1 relates to these processes is poorly understood. In this study, we performed a comprehensive transcriptome analysis of tissue- and ploidy-specific expression in a hypomorphic atxr5/6 mutant and revealed that the…

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How to label columns in HTSeq output

How to label columns in HTSeq output 0 I’ve been working to process RNAseq data and I’ve used hisat2 to align my reads to the reference genome. When I take those output files and put them into HTSeq-count using the below code, I get a count matrix but the columns…

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htseq-count -t gene not working

I found a little problem. When I set the “-t gene”, the reads is mark “__no_feature”. But when I set the “-t exon”, the reads is mark “ENSG00000276104”. The gene “ENSG00000276104” is a single exon gene. I don’t know why this happens. reads: “TGTCTGTGGCGGTGGGATCCCGCGGCCGTGTTTTCCTGGTGGCCCGGCCGTGCCTGAGGTTTCTCCCCGAGCCGCCGCCTCTGCGGGCTCCCGGGTGCCCTTGCCCTCGCGGTCCCCGGCCCTCGCCCGTCTGTGCCCTCTTCCCCGCCCGCCGATCCTCTTCTTCCCCCCGAGCGGCTCACCGGCTTCACGTCCGTTGGTGGCCCCGCCTGGGAC”. I had aligned to hg38 by…

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htseq-count python tutorial attribute counts error

Hello, I’m following the htseq-count tutorial for RNA-seq (counting the overlapping genes and exons) here htseq.readthedocs.io/en/master/tour.html. However, when I get to the point where I need to find the overlaps in the .sam file and .gtf file, I get an error. This is the code I ran originally that gave…

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htseq-count Error ‘_StepVector_Iterator_obj’ object has no attribute ‘next’

htseq-count Error ‘_StepVector_Iterator_obj’ object has no attribute ‘next’ 0 I am trying to run htseq-count (v. 0.13.5) on a sorted and indexed bam file. The command I entered looks like this: htseq-count -f bam -r pos -s yes -t CDS -i gene_id -m union filename_sorted.bam filename.gtf I get the following…

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HTSeq is a Python library to facilitate processing and analysis of data from high-throughput sequencing (HTS) experiments.

Hello, I’m following the htseq-count tutorial for RNA-seq (counting the overlapping genes and exons) here htseq.readthedocs.io/en/master/tour.html. However, when I get to the point where I need to find the overlaps in the .sam file and .gtf file, I get an error. This is the code I ran originally that gave…

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Running htseq-count to “grab” long non coding gene_id names

Running htseq-count to “grab” long non coding gene_id names 0 hi all, new to bioinformatics. so bare with me.. I am trying find long non coding RNA from RNA-seq data. As i checked the human gtf file there are 2 different types of long non coding RNA, “lnc_RNA” and “lncRNA”,…

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gffread error

hello I am currently trying to do RNA-seq using public data in brassica juncea. To use htseq-count for making count table, I have to convert gff file which downloaded in brassica database to gtf file. So I used gffread for converting gff file with below command gffread Bju.genome.gff -T -o…

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HTseq doesn’t support Multi-Threading ?

HTseq doesn’t support Multi-Threading ? 1 Hello, everyone ! I’m looking for a way to use HTseq with multi-thread. I couldn’t find any options about multi-thread. Anybody knows how to, please ? (I know there are tools support multi-thread like STAR, HISAT2. but just wonder whether HTseq doesn’t support it.)…

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Fastqc user manual – vodosp.ru

FASTQ format – Wikipedia 06 September 2021 – by TC Collin · 2020 · Cited by 3 — Be accompanied by a step-by-step user-friendly manual, If the user performs FastQC prior to the removal of adapters (step 3), the length Both programs can be used on Linux/MacOS X machines and quite…

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does not contain a ‘gene’ attribute

htseq-count returns : does not contain a ‘gene’ attribute 1 Dear BIOSTAR community, I’m trying to make count matrix with htseq-count, htseq-count -s yes -t gene -i gene 01.sorted.sam annotation_cattle.gff > 01.txt even with –idattr=gene , it returns error: Error processing GFF file (line 1864255 of file annotation_cattle.gff): Feature gene-D1Y31_gp1…

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Mapping reads and quantifying genes

Mapping reads and quantifying genes – Metagenomic workshop 0 Hello, I am using the following metagenomic workshop tutorial to analyse my own metagenomic data. metagenomics-workshop.readthedocs.io/en/latest/annotation/quantification.html I performed the following steps: mapped reads with bowtie2 and generated .bam file with samtools sort. Removed duplicates with picard Extracted gene information from prokka…

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Error creating DESeq2 Data Set from HTSeq-Count

I am trying to run DESeq2 using gene counts generated by HTSeq-Count. I combine files for different conditions: directory <- “~/GeneCountFiles/” WT_Files <- c( “P0CTRS3.aligned.sam.genecount”, “P0CTRS4.aligned.sam.genecount”, “P0CTRS5.aligned.sam.genecount” ) KO_Files <- c( “P0CTRS1.aligned.sam.genecount”, “P0CTRS2.aligned.sam.genecount”, “P0CTRS6.aligned.sam.genecount” ) I then create the sample table: sampleTable <- data.frame( sampleName=c(WT_Files, KO_Files), fileName=c(WT_Files, KO_Files), genotype=c(rep(“WT”, length(WT_Files)),…

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how htseq-count counts unstranded RNA-seq data

how htseq-count counts unstranded RNA-seq data 1 preliminary Say I have some unstranded RNA-seq data and im mapping to the reference human genome using htseq-count (–stranded=no) My understanding (biologically) was that for a given protein_coding gene, reading DNA in the sense strand gives the protein_coding transcript, reading the gene in…

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HTSeq-count TruSeq RNA Exome Lib Prep

HTSeq-count TruSeq RNA Exome Lib Prep 0 Hello, I observed a high percentage of “no features” while running HTseq w/ the –stranded yes option enabled (>80%). The library prep kit I am using is Illumina TruSeq RNA Exome which generates stranded data. If I run HTseq-count w/ strand == “no”…

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hisat2 compatibility for long read

hisat2 compatibility for long read 0 Hi, I am trying to align PacBio transcriptome reads against the genome to count the gene number. For pair end read i used the following workflow: # convert gff to gtf /home/software/cufflinks-2.2.1/gffread xxx.gff -T -o xxx.gtf # build index /home/software/hisat2-2.2.1/hisat2_extract_exons.py xxx.gtf > xxx.exon /home/software/hisat2-2.2.1/hisat2_extract_splice_sites.py…

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