Category: hisat2

Weird WGCNA plot

Weird WGCNA plot 2 Dear Seniors and All members, I am wondering whether anyone has done weighted gene co-expression network analysis (WGCNA) from Bulk-RNAseq before. I followed the WGNCA tutorial using my data and the module detection outputs are here. However, once I visualized the modules in clustering, I got…

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Exploration of Prognostic Biomarkers of Muscle-Invasive Bladder Cancer (MIBC) by Bioinformatics

This article was originally published here Evol Bioinform Online. 2021 Oct 28;17:11769343211049270. doi: 10.1177/11769343211049270. eCollection 2021. ABSTRACT We aimed to discover prognostic factors of muscle-invasive bladder cancer (MIBC) and investigate their relationship with immune therapies. Online data of MIBC were obtained from The Cancer Genome Atlas (TCGA) and Gene Expression…

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RNA-seq normalization methods between samples

RNA-seq normalization methods between samples 0 I’m a beginner in RNA-seq. I’m trying to learn RNA-seq analysis with a practical and simple analysis with public RNA-seq data. I downloaded 9 RNA-seq files (same sample prepared on 9 different days) I’ve done mapping them with Hisat2. so far it seems pretty…

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Running Hisat2 but it seems not working at all

Running Hisat2 but it seems not working at all 0 I’m running Hisat2 with the code below: hisat2 -p 9 -f ../0.Reference/CHO-PICRH -1 ../2.Trimmomatic/ERR2593198_1_trimmed.fastq.gz -2 ../2.Trimmomatic/ERR2593198_2_trimmed.fastq.gz 2> ../3.HISAT/ERR25593198.log| samtools view -Sbo ../3.HISAT/ERR2593198.bam It’s been running over 24 hours but it doesn’t seem work well. Can you please check the code…

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Error in Galaxy using Deseq2 “Error in data.frame(…, check.names = FALSE) :”

Hello everyone How are things going? Currently, I am processing ARN-seq data using Galaxy. I would like to observe the differential expression between controls and cases. For that, I follow the following steps: I did a quality check of my library first (obviously) Second, I used HISAT2 for reference genome…

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hisat2 SyntaxError: invalid syntax

hisat2 SyntaxError: invalid syntax 0 Hi all, I am really struggling with hisat2 recently and getting confused. I’m trying to run hisat2: hisat2 -t -p 2 -x shiryn RNAseq data/reference/grch38_genome/grch38 -1 shiryn RNAseq data/trimmed data/ly1_paired_1.trim.fq.gz shiryn RNAseq data/trimmed data/ly1_paired_2.trim.fq.gz -S shiryn RNAseq data/alighed/ly1.sam This is my error: File “/home/yun/Downloads/hisat2-2.2.0/hisat2_read_statistics.py”, line…

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hisat SyntaxError: invalid syntax

hisat SyntaxError: invalid syntax 2 Hi all, I am really struggling with hisat2 recently and getting confused. I’m trying to run hisat2 python2 /home/yun/Downloads/hisat2-2.2.0/hisat2 -t -p 2 -x shiryn RNAseq data/reference/grch38_genome/grch38 -1 shiryn RNAseq data/trimmed data/ly1_paired_1.trim.fq.gz shiryn RNAseq data/trimmed data/ly1_paired_2.trim.fq.gz -S shiryn RNAseq data/alighed/ly1.sam Heie is my error: File “/home/yun/Downloads/hisat2-2.2.0/hisat2”,…

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RNA seq trimming

RNA seq trimming 1 I am writing to know if 2-4 percent n content near the both ends of a sequence (RNA seq, read length:150) would be a problem and need trimming? RNA-Seq • 1.4k views It depends on the aligner that you use. If you’re using something like hisat2…

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Paired-end reads somehow counted twice?

Paired-end reads somehow counted twice? 0 Hi. I’m new in Bioinformatics and try to extract read counts from fastq files. I compared my result with answer count matrix, and read counts are doubled. (Left one is from the answer read count matrix, and right one is my result.) I used…

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Identification of Hub mRNAs and lncRNAs in Atrial Fibrillation Using Weighted Co-expression Network Analysis With RNA-Seq Data

This article was originally published here Front Cell Dev Biol. 2021 Oct 4;9:722671. doi: 10.3389/fcell.2021.722671. eCollection 2021. ABSTRACT Atrial fibrillation (AF)/paroxysmal AF (PAF) is the main cause of cardiogenic embolism. In recent years, the progression from paroxysmal AF to persistent AF has attracted more and more attention. However, the molecular…

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Numerical result out of range

samtools: Numerical result out of range 1 I am working with a SAM file (created with hisat2) with header: @SQ SN:chr1A LN:594006513 @SQ SN:chr1B LN:693261537 … When doing sorting and indexing, I get this error with samtools: [E::hts_idx_check_range] Region 536907741..536907892 cannot be stored in a bai index. Try using a…

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bowtie2 and then HISAT2? What does it mean in Data processing details on a paper?

bowtie2 and then HISAT2? What does it mean in Data processing details on a paper? 1 IMHO that must be a mistake: the data seems to be RNA so I would’ve directly gone for TopHat2 (or HISAT2). (Unless they wanted to run Fastqc in BAM, not fastq mode – in…

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Severe trauma and burns accompanied by sepsis

Introduction Trauma accounts for approximately 10% of deaths and 16% of disabilities worldwide.1 Billions of dollars have been spent on research into new biological therapeutics for severe injuries, as well as post-traumatic sepsis and septic shock.2 Burn injuries cause unpredictable trauma and sepsis is a complication associated with high morbidity…

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Opposite Signed Genes in WGCNA Signed Consensus Analysis

Hello, I was wondering if anyone could tell me how it would be possible to have genes that have negative correlations (as determined by kME values) with a module in a signed WGCNA analysis. My understanding is that, by definition, all the genes in a module should have a positive…

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how to get ride of duplicated genes when we also have duplicated Ensemble ID in the expression profile?

how to get ride of duplicated genes when we also have duplicated Ensemble ID in the expression profile? 0 Hi all, I have a mouse expression profile that is annotated with gene symbols and many of them are duplicated. I usually use collapseRows function with maxMean method from WGCNA package…

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Study Modules significance (or Modules Preservation) in ONE network

Study Modules significance (or Modules Preservation) in ONE network 1 I am studying only one co-expression network utilizing WGCNA. I would like to validate that the modules identified by WGCNA are significantly above randomly generated modules. 1. What would be the best way to do it? 2. Is there a…

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Identification of Hub Genes Associated With Clear Cell Renal Cell Carcinoma by Integrated Bioinformatics Analysis

This article was originally published here Front Oncol. 2021 Sep 30;11:726655. doi: 10.3389/fonc.2021.726655. eCollection 2021. ABSTRACT BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is a common genitourinary cancer type with a high mortality rate. Due to a diverse range of biochemical alterations and a high level of tumor heterogeneity, it…

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Does hisat2 –rg flag eat the “/” character in multiline definitions?

Does hisat2 –rg flag eat the “/” character in multiline definitions? 0 Hello, I am trying to use hisat2, but I noticed something weird. When running it like so: hisat2 -p 8 –rg-id=UHR_Rep2 –rg SM:UHR –rg LB:UHR_Rep2_ERCC-Mix1 –rg PL:ILLUMINA –rg PU:CXX1234-TGACAC.1 -x $RNA_REF_INDEX –dta –rna-strandness RF -1 “$RNA_DATA_DIR/${SAMPLE}_1.fastq.gz” -2 “$RNA_DATA_DIR/${SAMPLE}_2.fastq.gz”…

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What is the difference between HISAT2 index “genome.tran” and “genome”

What is the difference between HISAT2 index “genome.tran” and “genome” 1 I try to use HISAT2 in order to map the reads to the genome, while HISAT2 pre-index the genome for users. I saw that alternative sequences will affect the mapping results, does genome.tran index includes the alternative splicing and…

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High counts of rRNA in RNA-Seq

Hi there, I have recently performed RNA-Seq on the total RNA of a mosquito tissue, where I have three biological replicates of the tissue at three different time points. The pipeline I used was HISAT2 –> featureCounts –> DESeq2. Looking at the normalized counts (output of DESeq2), the counts of…

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extract gene sequence from multiple bam files in order to do Ka/Ks test

extract gene sequence from multiple bam files in order to do Ka/Ks test 0 Hello, I have a dataset of RNA bulk rna-seq (single-end), mapped with hisat2 and now I want to extract the sequences of some genes of my interest in order to run a Ka/Ks test for them…

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Why my reads mapping rate extremely low

I am using a script to analyze a large amount of ribo-seq data, which comes from different studies. Because from different research, I use trim_galore to remove the adaptor. Use bowtie2 to remove rRNA and use STAR mapping. After mapping, I found that the mapping rate of some of the…

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Importing WGCNA edge and node files into Cytoscape

I’ve used the WGCNA packages (horvath.genetics.ucla.edu/html/CoexpressionNetwork/Rpackages/WGCNA/) to generate edge and node files for use in Cytoscape. cyt = exportNetworkToCytoscape(modTOM, edgeFile = file.path(“./environment”, paste(label, “_CytoscapeInput-edges-“, paste(modules, collapse=”-“), “.txt”, sep=””)), nodeFile = file.path(“./environment”, paste(label, “_CytoscapeInput-nodes-“, paste(modules, collapse=”-“), “.txt”, sep=””)), weighted = TRUE, threshold = 0.02, nodeNames = modProbes, altNodeNames = modGenes, nodeAttr…

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Combining different data types into a single matrix for WGCNA using DESeq2 normalization

Combining different data types into a single matrix for WGCNA using DESeq2 normalization 0 Hi, I have two different RNA-seq datasets for the sample set of samples, generated using mRNA-seq and smallRNA-seq. The goal here is to identify a set of coding-genes and smallRNAs (no known function/targets) that act together….

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TPMCalculator returns zero for some genes, which are non-zero with Cuffdiff (FPKM).

TPMCalculator returns zero for some genes, which are non-zero with Cuffdiff (FPKM). 0 I ran TPMCalculator on my RNA-seq data. This was my first time using this package. It seemed to have done without any problems, but I realized that counts of some genes are zero. I performed DEG analysis…

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Biocmanager Install Vs Install Packages

Introduction to RNAseq I Day 3 Nicolas Rochette (EEB/ISG, UCLA) Karolina Kaczor-Urbanowicz (Oral Biology & Medicine, UCLA) UCLA Institute for Quantitative and Computational BiologyOver-representation (or enrichment) analysis is a statistical method that determines whether genes from pre-defined sets (ex: those beloging to a specific GO term or KEGG pathway) are…

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WGCNA error during network construction

I am performing WGCNA analysis on my RNAseq dataset and getting this error message: >net = blockwiseModules(expression, power = 6, + TOMType = “unsigned”, minModuleSize = 30, + reassignThreshold = 0, mergeCutHeight = 0.25, + numericLabels = TRUE, pamRespectsDendro = FALSE, + saveTOMs = TRUE, + saveTOMFileBase = “SW_TOM”, +…

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PCA result and batch effect?

PCA result and batch effect? 0 Hello, I am processing a dataframe that consists of about 55000 genes(TPM values,no access to raw data) and 400 samples. After removing the zero variance genes, I am performing a PCA on the samples trying to detect outliers. I have noticed that there are…

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What to do with genes that appear as NA but have statistics

What to do with genes that appear as NA but have statistics 0 Hi I’m doing an RNA-seq analysis and I’m having problems identifying some of the genes in the output of the DESeq2 tool because the list of genes obtained are in Entrez id, for context the pipeline used…

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deepvariant is very slow on alignements produced by hisat2 and reports only half of expected SNPs/INDELs

Hi @lpryszcz With respect to your aligner question, we have evaluated DeepVariant with BWA-MEM/BWA-MEM2, minimap2, DRAGEN, and the graph mapper Giraffe. For mapping to a linear reference, we would recommend BWA-MEM/BWA-MEM2 or DRAGEN. DeepVariant is trained on BWA MEM data. Minimap2 will work, but has slightly lower accuracy. We have…

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WGCNA Labeled Heatmap

Hi. I’m trying to create a labelled heatmap of module trait relationships using the following code from the WGCNA tutorial: png(filename = “Module-Trait Relationship.png”, width = 20, height = 30, res=300, unit=”cm”) par(mar = c(6, 8.5, 3, 3)) labeledHeatmap(Matrix = moduleTraitCor, xLabels = names(stressdatTraits), yLabels = names(MEs), ySymbols = names(MEs),…

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WGCNA, what does it mean if no hub genes are identified?

WGCNA, what does it mean if no hub genes are identified? 0 I ran WGCNA for my genes following the tutorials by Horvath and Langfelder (horvath.genetics.ucla.edu/html/CoexpressionNetwork/Rpackages/WGCNA/Tutorials/). I’ve obtained my modules and the GS and MM for my list of genes – however I think my situation is a bit odd….

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iCOMIC: a graphical interface-driven bioinformatics pipeline for analyzing cancer omics data

Abstract Despite the tremendous increase in omics data generated by modern sequencing technologies, their analysis can be tricky and often requires substantial expertise in bioinformatics. To address this concern, we have developed a user-friendly pipeline to analyze (cancer) genomic data that takes in raw sequencing data (FASTQ format) as input…

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Genome indexing with hisat2 and samtools

Genome indexing with hisat2 and samtools 0 Hello, I found this code in Biostar handbook (RNA-Seq by Example) in section about genome indexing. # The reference genome. IDX=refs/genome.fa # Build the genome index. hisat2-build $IDX $IDX # Index the reference genome with samtools. samtools faidx $IDX I was wondering about…

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Differences in HISAT2 indexes for download

Differences in HISAT2 indexes for download 0 Hi all, hope you guys are doing fine in the pandemic. So I have just recently beginning to move into computational biology stuff (and found it amazing) and one of the first things Im trying to do is to map fastq files against…

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WGCNA

WGCNA 0 0 Entering edit mode 2 hours ago Nithya ▴ 10 Can any one help to solve this Error?? Installation WGCNA • 15 views ADD COMMENT • link 2 hours ago by Nithya ▴ 10 Login before adding your answer. Similar Posts Loading Similar Posts Traffic: 1630 users visited…

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Weird PCA plot based on WGCNA results

Weird PCA plot based on WGCNA results 0 Dear all, I used WGCNA to find the associated gene modules with the different subtypes of a given cancer as trait. I obtained multiple modules associated with some of the cancer subtypes as shown in . Next, I plotted PCA to visualize…

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How to install hisat2 on Mac OS Big Sur

How to install hisat2 on Mac OS Big Sur 0 I’d like to install HISAT2 on Mac OS Big Sur ver.11.5.2. I have tried to install HISAT2 by using bioconda (Hisate2 :: Anaconda), and the installation seemed to be successful, but when I tried to start hisat2, I’m getting the…

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hisat2-align exited with value 1″ in mapping long noncoding RNAs

what does mean this error ? “(ERR): hisat2-align exited with value 1” in mapping long noncoding RNAs 0 Hello everyone.what does mean this error ? “(ERR): hisat2-align exited with value 1” in mapping long noncoding RNAs. mapping long RNAs non-coding • 30 views • link updated 2 hours ago by…

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Does Hisat2 automatically align strandedness

Does Hisat2 automatically align strandedness 0 I’ve been using Hisat2 to align some RNA. RNA was prepped using NEB Directional library kit. When I originally aligned, I did not use the –rna-strandedness option. I then realised I do want stranded, as part of my reference (which is human genome plus…

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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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Methylkit (differential methylation analysis) error in data frame

Methylkit (differential methylation analysis) error in data frame 0 Hi Friends, I am performing the differential methylation analysis using methylkit but it is throwing error in data frame. ” Error in [.data.frame(data, , 6) : undefined columns selected”. Can anyone help me to solve this error. Following is my R…

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Hisat2 error

Hisat2 error 1 Hi all I am using this command. hisat2 -p 8 -q -X ${READDIR}/hisat2_index_Cm_genome -1 ${READDIR}/tr.p.UE-2955-CMLib1_S88_L004_R1_001.fastq.gz -2 ${READDIR}/tr.p.UE-2955-CMLib1_S88_L004_R2_001.fastq.gz -S ${Output}/UE-2955-CMLib1.sam Error: Encountered internal HISAT2 exception (#1) Need a help to fix this problem. thanks Hisat2 • 24 views It is a lowercase x, so -x for the index….

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hisat2 alignment showing 0% aligned but sam file is 6GB in size.

hisat2 alignment showing 0% aligned but sam file is 6GB in size. 0 Hello, I am trying to align my fastq files with the reference genome assembly (MRSA ATCC 33591). The sam file formed is some 5-6 GB in size whereas hisat2 output is showing 0.0% alignment rate. I am…

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How to select uniquely and concordantly reads from hisat2 alingment for raw read count

Ok, let’s go through it. The -f 0x2 bitwise flag selection keeps proper pairs, which are read pairs mapped in correct orientation (I suppose illumina reads facing each other) and within insert size (-X and -I) -> concordantly. Now, if you want the uniquely mapped, you should basically exclude all…

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How to align fastq files against a reference assembly

How to align fastq files against a reference assembly 0 Hello, I am trying to align fastq files against the bacterial MRSA ATCC 33591 reference genome. The problem I am facing is that I have the reference assembly in fasta format with multiple sequences and upon creating index with hisat2-build…

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samtools server vs cluster error

Using inspiration from this thread HISAT2 output direct to bam, I’m attempting to run this command. The shell variables in this case represent paths to files/locations that make sense and in fact this command runs fine on my Ubuntu 18.04 LTS server using hisat 2.1 and samtools 1.10 (this seems…

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Unable to locate package hisat2″

how to solve this error when I want to install HISAT2? “E: Unable to locate package hisat2” 1 Dear all, I need to install HISAT2 aligner in my study. My Linux version is 16.04 (Xenial Xerus). So I used the below command : sudo apt-get install -y hisat2 but I…

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Pybedtools error sans

Pybedtools error sans 20-08-2021 pysam – Error when I install samtools for python on windows – i trying install pysam, pybedtools modules on python got error: ($i=1; $i[email protected] temp]$ conda install pysam bedtools hisat2 [ snip. However,…

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Running featureCounts separately for each sample and merging results

Running featureCounts separately for each sample and merging results 3 I am running a differential expression analysis project using HISAT2 for alignment and featureCounts for assembly. I have 27 samples with paired-end reads, and the FASTQ files alone take up about 270 GB. After running alignment for a few samples,…

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Question regarding providing file path to indexed genome folder

HISAT2: Question regarding providing file path to indexed genome folder 0 hisat2 -p 1 -x hisatIndex/genome.fa –dta –rna-strandness RF -1 $R1 -2 $R2 -S genome.sam(ERR): “hisatIndex/genome.fa” does not exist Exiting now … this error showing up every time i run the code and I dont know what is the problem…

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align using file.ht2

align using file.ht2 1 now i downloaded in my terminal indexed file of UCSC hg19 and when i uncompress it , i found two files genome.5.ht2 genome.8.ht2 and every time i want to align my samples at indexed file this error show up [e::bwa_idx_load_from_disk] fail to locate the index files…

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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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