Tag: RPKM

Appropriate RPKM cutoff 0 Hey, I’m using multiple previously published RNA-Seq studies as validation and to search for similar “signatures” as in our data. For these other studies I have their final read counts, and statistically significant filtered data that includes RPKM, FPKM, or other normalized read values as per…

Dubilier N, Bergin C, Lott C. Symbiotic diversity in marine animals: the art of harnessing chemosynthesis. Nat Rev Microbiol. 2008;6:725–40. Article  CAS  PubMed  Google Scholar  Sogin EM, Kleiner M, Borowski C, Gruber-Vodicka HR, Dubilier N. Life in the dark: phylogenetic and physiological diversity of chemosynthetic symbioses. Annu Rev Microbiol. 2021;75:695–718….

ATAC peak-calling: same peak ID across multiple experimental conditions? 0 I’m trying to reproduce the ATAC analysis here. I have a few questions about alignment and peak-calling. In the “ATAC-seq alignment” section in Method Details, they use Trimmomatic to remove adapters. However, I don’t know the adapter sequence. Would it…

v + T – normal size Emirates Airlines carried 43.6 million passengers, a growth of 123%, in the fiscal year 2022/2023, with a seat capacity increase of 78%, and seat occupancy of 79.5%, compared to 58.6% in the previous fiscal year. RPKM increased by 7% to 37.5 fils (10.2 US…

Average bigwig files (not sum) 1 Hello, I have bigwig (RPKM) files of a chip-seq experiment for treatment and control conditions which I am trying to compare. I have 3 replicates for control and 5 replicates for treatment condition. To show the average difference in signal, I merged the replicates…

In this section, we derive a simplified expression for the proportionality constant \(\alpha\) that enables quantitative ChIP-seq and we introduce some consequences for track building. This new expression is more intuitive to understand, easier to evaluate, and more accurate to sequencing outcomes than the previous expression. While values derived from…

Hello! I have a table of counts that I got by aligning rna seq samples with STAR and using featureCounts, and my goal is to get TPM values for each gene of the table. As a first step, I imported my table into R and modified it a bit to…

RPKM normalization 1 I have strand specific RNASeq data. I have mapped sequences of negative strand on Negative strand bam file and positive strand sequences on positive strand bam file. I got the read counts. Now, I have to RPKM normalize it, but I am confused weather I should take…

Hello, I’m currently following this paper Myc Regulates Chromatin Decompaction and Nuclear Architecture during B Cell Activation. It is a ChIP-Seq to generate genome-wide maps of 34 chromatin modifications (17 acetylation marks, and 17 methylation marks) and the histone variant H2AZ. B cells were either naïve (G0) or activated for…

Abstract It is generally assumed that viruses outnumber cells on Earth by at least tenfold. Virus-to-microbe ratios (VMR) are largely based on counts of fluorescently labelled virus-like particles. However, these exclude intracellular viruses and potentially include false positives (DNA-containing vesicles, gene-transfer agents, unspecifically stained inert particles). Here, we develop a…

What is used to measure transcript abundance? a variety of units, which have different requirements in order to ensure comparisons are meaningful number of reads that align to a given feature What unit does differential expression often use? What do counts depend on? sequencing depth/library size and on feature length,…

Hoe calculate RPKM from RUVr? 0 I have an RNA-seq count table which is generated by integrating several studies. I want to calculate RPKM but first I run RUVr to remove the unwanted variables. I want to know if I use normCounts(RUVr) to calculate RPKM, would it be correct? RUV…

Novel Roseobacter strains isolated from marine biofilms We isolated and cultured bacterial strains from biofilms on natural rocks immersed in coastal waters. After preliminary analyses of 16S rRNA gene sequences generated by Sanger sequencing, more than 500 non-redundant strains were identified, including 54 distant strains affiliated with Roseobacter (hereafter referred…

How to normalise ChIP-Seq samples using deeptools 0 I was wondering what is the best approach to normalising Chip-seq samples using the deeptools software. In our data set we’re using an antibody to pull down specific regions of the genome. But when I do the coverage plot I already get…

Microfluidic probe-based scRNA-seq method and validation. a, Cells were fixed and permeabilized to allow the penetration of thousands of unique, genome-specific oligonucleotide probes. Hybridized probes retrofitted transcripts with a poly-A tail and UMI, whereas unhybridized probes were washed away. b, Permeabilized cells with hybridized probes were flowed through a commercial…

How to “guess” the transformation based on already-transformed, “normalized count data”? 2 Thanks for your attention, TLDR: The minimum value in a transformed count matrix is -2.57. How can I guess what transformation was applied? Any good advice on performing differential gene analysis on such transformed data? Details: I would…

Question about mixing TPMs/RPKMs in deconvolution analysis 0 Hi all, I have a rather silly question that I’m fairly certain I know the answer to, but since I have never performed cellular deconvolution myself, nor do I have experience in typical workflows for the analysis/using the tools needed, I’ll ask…

. 2023 Mar 22;18(3):e0282428. doi: 10.1371/journal.pone.0282428. eCollection 2023. Sarah K Highlander  1 , Jason M Wood  2 , John D Gillece  1   3 , Megan Folkerts  1 , Viacheslav Fofanov  3   4 , Tara Furstenau  3 , Nitin K Singh  2 , Lisa Guan  2 , Arman Seuylemezian  2 , James N Benardini  2 , David M Engelthaler …

I was trained in cell biology and biophysics and, as I was often reminded by study section reviewers, I am “not a cancer biologist”. My closest collaborator, the cell biologist Gunter Blobel6, with whom I had worked for 25 years, encouraged me to leap off in this new direction, building upon…

Calculate RPKM 0 Hi bioinformaticians, would anyone calculate RPKM from the count matrix with edgeR or DESeq2? I found some resources guide on this but there are one or two steps I don’t know such as this: y <- DGEList(counts=counts,genes=data.frame(Length=GeneLength)) y <- calcNormFactors(y) RPKM <- rpkm(y) How to get GeneLength?…

Calculating RPKM for DESeq 1 Hi friends DESeq works via count table and create DE genes by count fold change, but it’s better to know about gene RPKM values. Is there any possible way to use / show RPKM for DESeq’s output? And how can i normalized those RPKM values?…

Human subjects Research involving human subjects was approved by the Institutional Review Board of the University of California, Irvine (#2003-3131). All enrolled patients provided written, informed consent. Clinical investigations were performed based on the ethical principles of the Declaration of Helsinki26. Cybrid generation Patient blood was collected in tubes containing…

A gain-of-function screen for regulators of gene silencing We utilized the native Arabidopsis gene FWA as a reporter to screen for regulators of gene silencing. FWA encodes a transcription factor that causes a late flowering phenotype when overexpressed, resulting in a greater number of leaves produced before flowering. In Col-0…

Transcribed image text: 2) Consider the following comparisons corresponding to genes ( 3 pts) Consider the following experimental comparisons (in all cases Y axis vs X axis in that order) in experiments to detect transcript levels (RPKM or counts for each gene, using RNA Seq) for specific zygotic-only products WT…

MERVL exhibits distinct localization in mouse embryos To understand the dynamics of MERVL expression, we first analyzed publicly available single-cell RNA-sequencing (scRNA-seq) datasets from each blastomere at eight representative stages of preimplantation development18 (Fig. 1a). To define regions of nonredundant MERVLs in mouse genome, we used RepeatMasker to annotate the…

By Nuvama research SpiceJet’s (SJ) reported a positive EBITDAR of Rs 5.9bn as demand increased during the festive season. The return of aircraft and a waiver on lease rental also contributed to the growth of EBITDAR. Highlights: SJ’s lack of transparency in sharing its key operational data (RPKM, CASK) since…

Get TPM from RNA counts and gene length? 1 Hello, I am working with an RNA-seq FeatureCounts output file that supplies the counts for a given ENSG gene ID, as well as the gene length(according to documentation this is in base pairs, not kilobases). Is there a way to obtain…

The JMP Genomics has a few normalization methods for RNAseq data, including KDMM, RPM scaling, TMM, TPM and upper quartile scaling. The JMP Pro 17 is missing such important tools. The purpose of normalization methods for RNAseq or other large scale data, such as metabolomics, is to reduce systematic experimental bias…

Chromosomal-scale genome assembly and full spidroin gene set of T. clavata To explore dragline silk production in T. clavata, we sought to assemble a high-quality genome of this species. Thus, we first performed a cytogenetic analysis of T. clavata captured from the wild in Dali City, Yunnan Province, China, and…

Revenue grew 61% YoY on 34% volume growth. Yield (revenue/RPKM) increased 22% YoY and 6% QoQ to Rs5.38. Ebitdar margin improved 21ppt QoQ and 80bp YoY to 21.3% (250bp beat). If we exclude the MTM hit on Forex liabilities, Ebitdar margin came in at 25.3%, a 14-quarter high. Absolute Ebitdar…

The abundance of metabolic marker genes is shown on the basis of the metagenomic short reads across the seawater sampled from the three study sites (left; n = 14), metagenomic short reads from the Tara Oceans dataset (middle; n = 213; replicates averaged) and metatranscriptomic short reads from the Tara Oceans dataset (right; n = 89;…

How to choose Normalization methods (TPM/RPKM/FPKM) for mRNA expression Why do mRNA expression values need to be normalized? The unification of mRNA expression value measurements across studies, or the normalization of mRNA data, is a significant problem in biomedical and life science research. The abundance of transcripts is measured digitally…

Even as the recovery in the aviation sector has been highly impressive with stable traffic at around 90 per cent of pre-Covid levels, Brokerage firm Prabhdas Lilladhar has said the domestic passenger traffic is likely to decline nearly 8-9 per cent QoQ, led by the seasonal nature of Indian aviation…

Bedtools Bam To Bed With Code Examples With this article, we’ll look at some examples of how to address the Bedtools Bam To Bed problem . bedtools bamtobed [OPTIONS] -i <BAM> As we have seen, a large number of examples were utilised in order to solve the Bedtools Bam To…

TPM and RPKM normalization from counts dataframe 2 Folks: I have two dataframes for counts information from two RNAseq data… is there a quick way to get from counts to TPM or RPKM or both efficiently? Thanks RNA-Seq • 3.5k views • link updated 5.8 years ago by Ron &starf;…

Hello everyone I have multiple bulk RNA-seq datasets that I need to apply the same pipe line on. I want to normalize them from counts data to TPM. In all datasets, I have the genes as rows, and samples as columns. Unfortunately, I don’t have the fastq files, all I…

Warning – longer object length is not a multiple of shorter object length 0 I have a counts dataframe of RNA-seq dataset, and got the gene lengths using this code: exons = exonsBy(EnsDb.Hsapiens.v86, by=”gene”) exons = reduce(exons) len = sum(width(exons)) INDEX = intersect(rownames(counts),names(len)) geneLengths = len[INDEX ] counts = counts[INDEX…

Hi, I have multiple .txt files (each file contains 4 columns; an identifier Gene column, a raw_counts and other columns). I would like to merge those files into a combined dataframe using the common gene column. I was able to import multiple .txt files together, merge based on identifier column,…

Normalization for comparing gene coverage values. RPKM corrects differences in both: sample sequencing depth and gene length. RPKM – Reads per kilo base per million mapped reads Formula RPKM = numReads / ( geneLength/1000 * totalNumReads/1,000,000 ) numReads – number of reads mapped to a gene sequence geneLength – length…

Problem with the output of Deeptools PlotProfile: a strange pattern of repetitive summits 0 Hi! I am trying to plot DNA binding profiles of my ChIP-seq bw files using Deeptools plotProfile. I generated the matrix using the computeMatrix reference-point. I used some publicly available bed files as my regions of…

Dear All, I have a doubt in the calculation of False postitive rate while checking for FPKM threshold in a RNAseq experiment. Following the method previously published (www.ploscompbiol.org/article/…l.pcbi.1000598). I am not getting desired results. I followed the method as mentioned the publication Reads were mapped to Ensembl genes (blue) and…

RPKM, how to normalize for library size 1 Hi everybody! I’m working on several metagenomes (so not trascriptomes), and I mapped high quality reads to a database made of nucleotide sequences through bowtie2. I want to convert raw mapping counts of each gene in RPKM. I know this method might…

Infection with L. donovani amastigotes or promastigotes promotes early changes in the mRNA pool of the host cell To compare the early effects of the two life stages L. donovani in the mature mRNA pool of the host cell, total cytosolic mRNA extracts from bone marrow-derived macrophage (BMDM) cultures infected…

Can I convert HTSeq count into RPKM or TPM value or standard unit of RNA-Seq 0 Now, I’m comparing RNA expressions that have RNA-Seq and HTSeq count How can I interpret it together with different unit or Can I convert HTSeq count equivalent RNA-Seq? or if you have other suggestions,…

TCGA Data download in terms of ease of use ,RTCGA The bag should be better , And because it’s already downloaded data , The use is relatively stable . But also because of the downloaded data , There is no guarantee that the data is new .TCGAbiolinks The package is…

How to do functional analysis on differentially expressed gene list from RNA-seq data? 0 @06f08eeb Last seen 1 day ago Canada Hi all, I am a complete beginner in terms of bioinformatics analysis and I am hoping to complete some functional analysis on some differentially expressed gene lists of some…

Hi guys, I hope you are fine. I’m not good in English so if you couldn’t understand my question, please feel free to reply. I’m a beginner of bioinformatics. I want to practice differential expressed gene (DEG) analysis in R. The RNA seq data I used was downloaded from broad…

Hi Xuran, I tried to apply MuSiC to RNAseq bulk data with RPKM as the input. According to your paper(Discussion), “MuSiC can utilize RPKM if estimates of cell type-specific total RNA abundance can be provided.” I am wondering how I can incorporate cell-type-specific total RNA abundance into your function? Or…

I am making a heatmap using RNA-Seq data in R. The heatmap shows gene expression values (RPKM) in different brain regions. I have the following code: library(tidyverse) library(pheatmap) library(matrixStats) read_csv(“prenatal_heatmap_data.csv”) -> all_data all_data %>% column_to_rownames(“Brain Region”) -> heatmap_data heatmap_data %>% pheatmap() Which generates the following heatmap: I want to do…

Single guide ribonucleic acid (sgRNA) molecules are produced by discontinuous transcription, in which viral RNA-dependant RNA polymerase pauses early negative-sense RNA synthesis and then jumps to the other end of the genome. The specifics of this process are still not fully understood. Since sgRNAs can play an important role in…

Single guide ribonucleic acid (sgRNA) molecules are produced by discontinuous transcription, in which viral RNA-dependant RNA polymerase pauses early negative-sense RNA synthesis and then jumps to the other end of the genome. The specifics of this process are still not fully understood. Since sgRNAs can play an important role in…

Hello everyone! Very new to analyzing RNAseq data and trying to learn the process behind it. This is completely not my area, but I would like to learn and was hoping someone could give me an explanation to my question. Thank you! I’d like to know if a few genes…

How to define the gene length for RPKM calculation 4 Hi guys, I would like to calculate the RPKM of my RNA seq experiment. To do this, as from the formula, I need to know the gene length. My starting point are the row reads (single end) counts resulting from:…

Introduction Corn and soybean meal are the main components of high energy and high protein diets for pigs and are also the main raw materials of food products for human consumption, fermentation, and bioenergy industry (Sevillano et al., 2018). As the arable land of food crops was limited whereas the…

Introduction CD19 is an important target for novel anti-lymphoma treatments as it is broadly and homogenously expressed across many B-cell malignancies [1,2]. Approximately 30–50% of patients with diffuse large B-cell lymphoma (DLBCL) who do not respond to first-line therapy with R-CHOP have a poor prognosis and need effective treatment options,…

Using TCGA RSEM data to calculate isoform expression 0 Hello everyone, I have download the TCGA RNAseq RSEM data about isoform expression. I would like to check which one of the isoforms of my gene is the one expressed the most. Can I directly use the RSEM values to conclude…

Hello, I am interested in finding out at what stages of development a gene of interest is expressed in the human brain. I am using the Developmental Transcriptome tool from the Allen Brain Atlas to find this information. The heatmap showing the gene expression levels across the different brain samples…

Using dgelist function in EdgeR to calculate RPKM 0 Hi, I’m new to RNAseq and I want to calculate rpkm values from my raw counts but I am unsure if my understanding of how to do so is correct. From what I understand, I should do the following: df <-…

The removal of problematic regions The removal of problematic genomic regions is considered essential for the accurate analysis of data obtained by chromatin immunoprecipitation followed by genome sequencing (ChIP-Seq) [27, 35]. Repetitive regions including satellite DNA arrays comprise a majority of such problematic regions, mainly because they reside in the…

Asked by: Vada Ratke Score: 4.7/5 (25 votes) BigWig is a file format for display of dense, continuous data in a genome browser track, created by conversion from Wiggle (WIG) format. BigWig format is described at the UCSC Genome Bioinformatics web site, and the Broad Institute file format guide provides…

Analyzing gene expression in different RNAseq datasets 0 Hello! I really need some assistance here, I came up with an analysis of my own that makes sense to me but I really new in this (started studying bioinformatics on my own with the pandemics) and I’m not sure if I…

Hi all, I am a PhD student with biology background. I recently inherit a RNA Sequencing project from another PhD student in my lab. We already have paired-ended RNA-Seq data generated from Illumina HiSeq but haven’t started analysis yet. I have basic Linux command line training but have no idea about how…

Convert FPKM and RPKM to TPM values 0 Hi all, I’m trying to collect the datasets from public datasets from GEO. I found some data in fpkm and some in rpkm,tpm. My goal is to feed these data into a machine learning model, but my question is “ALL THESE DATA…

What you describe seems to be a difference in signal-to-noise ratio which is not uncommon. This is where more elaborate normalization methods such as TMM from edgeR or RLE from DESeq2 come into play. See the following links on why these methods are superior. The videos talk about RNA-seq but…

Forum:Misuse of RPKM or TPM normalization when comparing across samples and sequencing protocols 2 Published in the RNA Journal in 2020 – this paper argues that if the original RNA amount in the different samples is different, TPM should not be used to find differentially expressed genes. www.ncbi.nlm.nih.gov/pmc/articles/PMC7373998/ Seems like…

Calculating fpkm from raw counts 2 Hello, I have a file consisting of raw counts. For my analysis, I want to normalize the counts by fpkm. The question is, how can I calculate fpkm from raw counts? Also, upon a little reading I found that DESeq2 could do i, but…

Statistics on RNAseq data 2 Hi I would like to know whether you can do statistical tests (e.g. ANOVAS etc.) on the TPM/RPKM counts of RNAseq data? Thanks on Statistics data RNAseq • 58 views This is not recommended due to a few underlying problems with RNA-seq data that include…

TMM followed by inverse normal transform 0 Hey all, I am following a protocol from a paper that uses the following pre-processing procedure: a. Read counts were normalized between samples using TMM (Robinson, M. D. & Oshlack, A. A scaling normalization method for differential expression analysis of RNA-seq data. Genome…

RPKM on TSS using DiffBind 1 Hi everyone, I want to extrapolate RPKM value from Diffbind. Investigated regions are TSS (± 2.5kb) but around 3500 TSS are “lost” because merged. I have read that in the new version of DiffBind is possible to use dbObj_region$config$mergeOverlap with negative value to avoid…

RPKM and differential expression analysis 1 Hello all, I am trying to analyse differential expression in a dataset for which I only have RPKM values available to me. I usually use the R/BioC environment for RNA-seq analysis, and have read in various BioC documentation that using RPKM values in packages…

pre-proccessing of RNAseq data for WGCNA 0 Hi everyone, i wanted to create an expression matrix for WGCNA input. however, i has been said that use RPKM/FPKM data instead of CPM, how can i change my TCGA data to RPKM/FPKM in GDCquery and how to filter expression set of genes…

DESeq2 with a small number of genes 1 Dear all, I am writing a program in order to study the coverage of only one sequence. To sum up the pipeline: Detect ORFs in the input sequence Align all reads on the sequence (bowtie), reads come from RNA-seq Count the number…