Tag: RPKM

DESeq2 input from GDAC firehose

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…

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How to perform deconvolution with RPKM values

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…

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r – RNA-Seq Data Heatmap: Is it necessary to do a log2 transformation of RPKM values before doing the Z-score standardisation?

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…

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New bioinformatics method to analyze viral sgRNA

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…

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New bioinformatics method for viral sgRNA analysis

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…

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Comparing RPKM (calculated from raw counts rnaseq file) between a few genes across 2 experimental conditions with a t-test : bioinformatics

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…

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How to define the gene length for RPKM calculation

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:…

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Frontiers | Metagenomic Analysis Reveals New Microbiota Related to Fiber Digestion in Pigs

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…

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CD19 expression is maintained in DLBCL patients after treatment with tafasitamab plus lenalidomide in the L-MIND study

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,…

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Using TCGA RSEM data to calculate isoform expression

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…

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Is it conventional to use a threshold RPKM value to determine if a gene of interest is expressed in a sample? : bioinformatics

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…

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Using dgelist function in EdgeR to calculate RPKM

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 <-…

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Differential enrichment of H3K9me3 at annotated satellite DNA repeats in human cell lines and during fetal development in mouse

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…

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What is bigwig file?

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…

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Analyzing gene expression in different RNAseq datasets

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…

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Up-to-date RNA-Seq Analysis Training/Courses/Papers (Dec 2017)

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…

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Convert FPKM and RPKM to TPM values

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…

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ATAC-seq sample normalization

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…

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Misuse of RPKM or TPM normalization when comparing across samples and sequencing protocols

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…

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Calculating fpkm from raw counts

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…

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Statistics on RNAseq data

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…

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TMM followed by inverse normal transform

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…

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RPKM on TSS using DiffBind

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…

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RPKM and differential expression analysis

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…

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pre-proccessing of RNAseq data for WGCNA

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…

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DESeq2 with a small number 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…

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