Questions tagged [rna-seq]

Questions should include this tag if they pertain to issues related to bioinformatics analysis of RNA-seq data, e.g. normalization, differential expression analysis, sequencing and experimental design or transcriptome assembly...

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What do the numbers mean in these RNA-Seq gene/transcript TPM files?

From the link https://gtexportal.org/home/datasets, under V7, I'm trying to do R/Python analyses on the Gene TPM and Transcript TPM files. But in these files (and to open them I had to use Universal ...
Macromind101's user avatar
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Getting VCF file that contain common SNPs from 6 VCF file using isec

I have 6 VCF files, where I would like to obtain the SNPs that are common (by position) in all the 6 files. I have tried this command ...
Mohamed Samir's user avatar
1 vote
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How can I delete these lines in my fastq file?

Hi guys =) I'm sorry if this is a repeat but I haven't been able to find an answer using the search field or google. I am trying to edit a fastq file that has a corrupt line. The read in question is ...
Sebastian Quezada's user avatar
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What statistical test to apply for DE after CibersortX deconvolution?

This question was also asked on Biostars I am running CibersortX in high-resolution mode (which yields estimates of gene values per sample). After that, I want to perform DE between two conditions on ...
Sam's user avatar
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Salmon Pseudo count when dealing with male and female RNA-seq data

I've generated a quant seq data that I intend to use to compare male and female gene expresion, with a focus on sexual chromosome. For my species (three-spined stickleback), it is a classic XY sex ...
Florent Sylvestre's user avatar
1 vote
1 answer
48 views

Gene rank scores from LINCS database: Evangelista et al. 2022

The website https://maayanlab.cloud/sigcom-lincs/#/SignatureSearch/UpDown can be used to "Identify reversers and mimickers from over 1 million signatures by entering up and down gene sets .. &...
c00kieRaptor's user avatar
1 vote
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30 views

How to use hashsolo for demultiplexing hashtags?

I am trying to use hashsolo and I want to make sure that I have done things correctly. I did the below: ...
pythonbeginner's user avatar
2 votes
1 answer
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Why is there antisense sequence in RNAseq data

I'm looking at RNAseq data from CCLE. The data is paired-end. Take the cell line Hs578T and the gene HRAS as an example. The cell line carries a G12D mutation (c.35G>A), so the change in cds is: <...
geom_na's user avatar
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5 votes
2 answers
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Integrating bulk RNA-Seq data with different sequencing depths and from different sources

I am attempting to integrate different bulk RNA-Seq datasets. While this is not ideal, I'm trying to reduce the technical variability in these datasets by using data generated by similar protocols (...
CodingSquirrel's user avatar
1 vote
1 answer
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Test for differences between groups of samples

Sorry if the answer to this should be obvious. I have RNA-expression results from 24 samples which can be divided into 6 groups, (wildtype and two different mutants at two different ages) with a total ...
Sethzard's user avatar
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How to remove orig ident. already data normalized, scaled assigned cluster/

How to remove orig ident. already data normalized, scaled assigned cluster/ Example, I have 4 sample data. like 1-control, 2- treatment with A, 3- treatment with B, and 4- treatment with C, I want to ...
Sandi's user avatar
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How to differentiate DNA fastq and RNA fastq files?

I have 2 sets of fastq files from another collaborator. The first is contains exome data and the second contains RNAseq data. But both have the RNA in the name but a different ID. How do I ...
SNanda's user avatar
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2 votes
2 answers
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RNAseq alignment: best practices for aligning to multiple isoforms?

I have Illumina RNAseq data and would like to maximize my power to find candidate genes that are differentially expressed genes between experimental conditions. Many of my (de novo assembled and ...
DavidR's user avatar
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How would you define a set of active genes from a bulk RNA-seq experiment in one biological sample? [duplicate]

I am trying to identify genes that are overall considered to be "actively transcribed" in my cell line of interest for some downstream analysis. What would the approach be for defining this ...
ricardo3889's user avatar
3 votes
1 answer
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Nextflow Error: failed to read header from "-"

I am trying to run my nextflow pipeline, and have gotten this error: samtools sort: failed to read header from "-" I'm not sure why this error is ...
anne's user avatar
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1 answer
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RNAseq: Why would using a more complete scaffold reduce DEGs?

I have a set of RNAseq data for a couple of relatively under-studied species. They're fungi, of the order Hypocreales. The assemblies we're working with are de novo - reference seqs don't actually ...
HKI_PJTB's user avatar
1 vote
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Generating gene signature for sample classification and survival analysis patient cohort

This is from this paper The LSC17 score is calculated for each patient as a linear combination of GE of these 17 genes weighted by regression coefficients that were estimated from the training data as ...
PesKchan's user avatar
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Nextflow Pipeline: Unexpected Input "{"

I'm attempting to run a nextflow pipeline and have begun getting unexpected output for a few of the processes in my pipeline. I believe this to be a syntax error, but I am not sure where to correct it....
anne's user avatar
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To find genes that don't change in RNA-seq, Deseq2 has altHypothesis="lessAbs". Is there a way to make limma do the same thing for proteomics?

I love that Deseq2 has altHypothesis="lessAbs" !!!!!!! I've used it a ton for RNA-seq. However, now I'm working with mass spect data using limma. Is there a way to make limma do the same ...
Mary Allen's user avatar
1 vote
1 answer
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Gene symbol list for all protein coding genes in mice

How can I get a csv or a list of the gene symbol names for all the protein coding genes in mice? I have RNA sequencing data and I'm not interested in the non-coding stuff. I'm worried it could mess ...
Angus Campbell's user avatar
2 votes
1 answer
29 views

Inconsistent replicate numbers in RNA-seq

I have 3 groups for the RNA-seq analysis (Control, treatment A and treatment B). There are 2 replicates for control and treatment A and 3 replicates for treatment B (lost 2 replicates due to a mistake ...
Wang Ming's user avatar
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3 votes
2 answers
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Which candida albicans fasta and gff file should I use for alignment?

I downloaded the gff and fasta files for candida albicans from http://www.candidagenome.org. I want to use hisat2 to align some fastq files against it. There are ...
irritable_phd_syndrome's user avatar
1 vote
2 answers
96 views

How to calculate cell type percentage in every sample

I have a Seurat object (metadata) with the single R samples consisting of cell types and sample types columns. I am trying to make a table that has a sample and percentage of cell types for each ...
Yogesh Budhathoki's user avatar
1 vote
1 answer
215 views

How much does Nanopore cDNA Sequencing Cost?

[this question is based on a question that was asked on Reddit] We're interested in doing whole-transcriptome cDNA sequencing of 30 mouse cell lines, and are deciding between Illumina and Nanopore ...
gringer's user avatar
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1 vote
1 answer
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blood contamination rna seq

I have tumor rna seq samples. I want to find out percentage of blood contamination within these samples. What is the best possible way to find % of blood contamination within the rna seq samples?
user3138373's user avatar
1 vote
1 answer
33 views

Error while using SummarizedExperiment() in R

I'm tryig to perform RKM normalization on raw counts for RNA-Seq Data: ...
Félix Agosto's user avatar
2 votes
0 answers
31 views

Does the contrast matrix I have made address the question I am trying to answer?

I have the following design matrix: mm_noreps.interactions <- model.matrix(~condition*TRAPed) Both variables are factors condition has 4 levels and TRAPed has 2 ...
Angus Campbell's user avatar
2 votes
1 answer
62 views

Is it possible to do homology inference across species using different kinds of NGS data?

Background: I have a list of species that I want to put through homology inference. The goal of homology inference is to investigate the evolution of a trait on a species tree. I want to use the ...
Sudoh's user avatar
  • 107
2 votes
1 answer
50 views

Examples of machine learning approaches to validate results where ground truth is lacking?

I am currently looking into some of the published methods for deconvolution of spatial transcriptomics data where each spot does not have single-cell resolution. These methods all rely on cell type ...
Alos's user avatar
  • 21
2 votes
2 answers
60 views

Display row and column names in R

I have plotted a heat-map with 235 rows and 570 columns which is below: how do we make clear names of row and columns on this graph using R?
Mahendra Singh's user avatar
4 votes
1 answer
141 views

Kallisto error: index input file could not be opened!

I am utilizing Kallisto in Anaconda/miniconda for RNA sequencing. I have successfully made ...
Tristan's user avatar
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3 votes
1 answer
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How to remove double '/' from file path, bash script

I'm using the following script to detect strandedness of my paired end RNA-seq data. ...
pubsurfted's user avatar
1 vote
1 answer
104 views

How to run how_are_we_stranded_here?

I'm trying to run the python library called how_are_we_stranded_here. I have installed with pip install how_are_we_stranded_here. I have also installed its ...
pubsurfted's user avatar
0 votes
2 answers
111 views

How do I determine read length from a user-inputted fastq file and then input that information into a tool's command line?

Can you guide me on how could take an input fastq file, read the length from read and then feed it into the findtail's command? I have suggested concerns about documentation and program defaults to my ...
pubsurfted's user avatar
2 votes
1 answer
56 views

How to make unrooted tree for Likelihood mapping result by using IQ2-tree?

I am a biologist, and I do not fully understand the tree topology of the experimental species. I used four-taxon set (4 sequences) to identify the Four-cluster Likelihood-Mapping by using ...
Adi's user avatar
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2 votes
1 answer
39 views

What are the conditions to perform Gene Expression Analysis on data you've performed RNA Sequence Analysis on?

I want to perform Differential Gene Expression Analysis. I recently completed RNA Sequence Analysis using the Seraut Tutorial up to this point: Finding Differentially Expressed Features (Cluster ...
Antonio's user avatar
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1 vote
1 answer
69 views

Can I perform Expression Analysis on this 10x dataset?

10x Genomics: 20k Human PBMCs is the dataset. Description of the dataset: Inputs/Libraries Human peripheral blood mononuclear cells (PBMCs) of a healthy male donor aged 30-35 were obtained by 10x ...
Antonio's user avatar
  • 141
2 votes
2 answers
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Calling isoforms from long read data generated from partially degraded RNA

What will be the best tool to call isoforms from long read data generated from partially degraded RNA. By mistake we processed some samples with poor quality RNA to generate long read. Now we are ...
user3377241's user avatar
0 votes
1 answer
83 views

How to find closely related genes for a specific gene from WGCNA modules?

I have used WGCNA by integrating several datasets which are processed in a similar way. Identified 17 modules, followed by performed pathway analysis with the genes present in those modules. Among all ...
user9114's user avatar
1 vote
0 answers
44 views

Benchmarking for variant identification using RNA-seq data

I am in need to benchmark the variant identification pipeline which uses RNA seq data alone without any matched-normal. I would like to know the reference dataset (and the pipeline on which the ...
Ahkam's user avatar
  • 11
5 votes
3 answers
89 views

Ubiquitous regulation of highly specific marker genes

I am fairly new to scRNAseq analysis and keep running into the same problem in the two datasets I am currently working with. We work with kidney inflammation and have two new mouse models, which we ...
Smeerlap's user avatar
1 vote
1 answer
49 views

Snakemake MissingRuleException

I'm trying to run a snakefile for the first time with limited coding experience using salmon to index a reference genome. I'm not too sure what I'm doing wrong based on this error message. ...
Hua Yuan's user avatar
1 vote
0 answers
22 views

Number of Spots in a Spatial Seurat object

I have got an integrated seurat object of 21 Spatial samples. I want to know how many spots are in all the samples together, Is there a way I can do that? Also, Please suggest any R Packages that do ...
David's user avatar
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1 vote
0 answers
72 views

Stringtie/DEprep.py gene/transcript IDs are wrongly formatted

Hi my RNAseq workflow is ending up with wrongly formatted gene IDs (and separately transcript _IDs) after a Hisat2 ->samtools sort -> stringtie -> DEprep.py workflow. DEprep.py outputs a ...
RichardBJ's user avatar
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2 answers
93 views

Is it possible to integrate multiple gene expression datasets and use it for WGCNA?

I have 8 RNA-seq datasets and am interested in looking at genes co-expressed with a specific gene. Among 8 RNA-seq datasets, 6 have less than 20 samples. Rather than working on each dataset ...
user9114's user avatar
0 votes
1 answer
130 views

select highly variable genes out of dataframe

lets say I have the following dataframe: ...
user16197's user avatar
2 votes
2 answers
53 views

WCGNA - Relate modules with Y features when the % of variance explained of each eigengen is low

I'm doing a WCGNA analysis (signed network) on microbiome 16S data. I have transformed counts to centeres log-ratio transformed data (CLR) to address the compositional characteristics of the data and ...
Julieta's user avatar
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2 votes
0 answers
23 views

Avoiding false anticorrelations between individual genes per cell in single-cell RNAseq

When analysing a single-cell RNAseq dataset, one question I like to ask is: At a single-cell level, are these two genes correlated (expressed together) or anticorrelated (only one or the other is ...
D Greenwood's user avatar
2 votes
0 answers
42 views

What codes represent what genes?

I want to run experiments on the data used in PatternMarkers & GWCoGAPS for novel data-driven biomarkers via whole transcriptome NMF link So far, the paper has reduced the dimension to ammon's ...
A.Dumas's user avatar
  • 487
0 votes
1 answer
40 views

Is loss/gain of function reflected in RNA-seq transcript counts?

Do LoF/GoF transcripts count toward the RNA-seq TPM count? Or would these LoF/GoF transcripts only be detected by isoform quantification?
Kermit's user avatar
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