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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Surrogate variable analysis for paired RNA seq experiment

I have an RNAseq experiment with 1000s of samples. Each sample comes from a patient and was treated either with a drug or DMSO (as control). I am currently deciding between SVA and RUVseq to account ...
nhaus's user avatar
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Abnormal representation of differentially expressed genes in volcano plots located at the extremities

This question was also asked on reddit Does anyone know how to correct the abnormal genes that seemed to be located on the curves near the extremities? The analysis was done using R package edgeR on ...
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Is it normal to have low mapping to bacterial genome in a total RNA sample of plant root-colonized bacterial cells?

I had isolated the total RNA from a sample of plant roots with colonized bacteria by excising the roots (harboring the bacterial cells) and crushing them into a powder using liquid nitrogen. The RNA ...
K_081's user avatar
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missing additional visits in GSE data

When downloading GSE data, often I encounter that the authors refer to additional visits in the data description, yet, they are not found in the data. For example, the following study. They mention ...
Kozolovska's user avatar
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Generating synthetic RNA-Seq data for performance assessment of new algorithm

I want to generate synthetic miRNA data for the performance assessment of the newly developed algorithm designed for miRNA data analysis. There are several synthetic sequence simulators available in ...
Lot_to_learn's user avatar
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1 answer
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rna seq matrix design hybrids

I am struggling to get my head around the correct design to use for my experiment. I have RNAseq of 2 pure species (s1, s2) and of the hybrid (h). I aligned all three genotypes to the s1 genomes. I am ...
RomainL.'s user avatar
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GSEA vs GSVA: Pros and cons of each method

Trying to understand when GSEA is more appropriate than GSVA and vice versa. I have seen cases when running GSEA and GSVA on the same task - compare enrichment of a geneset between two groups - gives ...
Tomas Bencomo's user avatar
5 votes
2 answers
89 views

Multiple genome assemblies of the same bacterial species

I have some RNA-seq data where there are reads from the "host" as well as from several bacteria species. In this experimental context, I am interested in the host associated reads and the ...
haci's user avatar
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Bedtools genomecov gives me coverage files showing zeros at all nucleotide positions

Also posted on biostars I have bam files that I've obtained through STAR alignment. Here is the first 10 lines of one of my bam files: ...
gcabebe's user avatar
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1 answer
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Salmon Multiple File SCRIPT giving error

Hi I am using Salmon quantitation for multiple fastq paired files using following code ...
Malik Saad's user avatar
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1 answer
61 views

After running nf-core, is there a way to map a gene_id to a specific gene's DNA sequence?

I have been running nf-core in Python and it works great! But I have a seemingly simple question that I'm struggling to find an answer for online. After running the nf-core pipeline on my RNA ...
user18959's user avatar
2 votes
1 answer
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WGCNA: Extract genes within the different modules to create a dataframe with two columns

My inquiry pertains to WGCNA. After identifying the modules, I would like to extract genes within the different modules to create a dataframe with two columns: ...
nann's user avatar
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How to normalise Bulk RNA-seq data to account for transcript length, coverage depth, library size and batch effects?

I'm currently working on a project where I'm comparing aggregate measurements (mean, median, etc.) of expression data (RNA-seq) from different groups of genes across various samples with different ...
Duhlky's user avatar
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Continuous predictor variable to identify responder genes using DESeq2

I have samples from 45 individuals at 2 time points and these individuals were given a specific diet. The BMI of these individuals were recorded at both time points. The responders to the diet are the ...
Angelo's user avatar
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Featurecount output .txt file from bam file

After running this command: featureCounts -p -s 2 -a $genome -o $dir"/"$specie"/count_table/"$value".txt" $output_loc -T 8 on the BAM output of ...
ToTheMoon's user avatar
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RNAseq and TTseq results

I'm working with two sets of data. One of two sets of data, one contains the results of a RNAseq analysis and the other contains the result of TTseq analysis. I want to check which genes overlap in ...
Adrián Coto's user avatar
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Suggestion for way foward with upregulated genes

I am working with single cell RNA data- control and treated. I analyzed and got a list of upregulated genes. Upon doing pathway analysis all the genes were of apoptotic pathway. But these genes can ...
subhiksha sundaram's user avatar
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problem with feature counts of RNA bulk seq paired data- in Rsubread

I did manage to bild index file (from NCBI transcriptome) and perform alingment in Rsubread in R with my fq files. I did get BAM files as a result and no error. However i am having trouble with next ...
MKE1508's user avatar
2 votes
1 answer
78 views

Classical alignment with HISAT2

This question was also asked on Biostars I have this script ...
Emmanuel Aroma's user avatar
1 vote
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What method uses "diverge probability" in RNA-Seq differential expression analysis?

I am replicating differential expression analysis from a paper (here) on RNA-Seq data. I extracted the regulated genes using DESeq2::results with a threshold of 1 ...
Hadeer Ibrahiem's user avatar
1 vote
1 answer
120 views

Kallisto RNA-seq errors because of incompatible indices when running <kallisto quant> using downloaded mouse index

I'm trying to run RNA-seq on single-end mouse RNA data (50bp reads) but have very limited RAM (<4 gb) - I don't think I really have a way to work around this memory restriction, and wanted to try ...
mz.'s user avatar
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issue with rna seq analysis

I am working on RNA seq analysis and I would like to know the following things: Current Methods: downloaded genome fasta file for non-coding rna from ensembl and got the gtf file for hg38 performed ...
subhiksha sundaram's user avatar
2 votes
1 answer
94 views

Mitochondrial genes - TPM calculation bulk RNA-Seq

This question was also asked on Biostars I was wondering if any of you have encountered a situation for bulk RNA-Seq where, possibly due to low sample quality or presence of dead cells, mitochondrial ...
nick_b55's user avatar
2 votes
0 answers
50 views

How to incorporate negative controls in DESeq2

We are doing a comparison between two outcomes (positive and negative). We could not have any positive controls as we do not have any "control" data to set as baseline, either from ...
Karthik Nair's user avatar
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99 views

how to use CIBERSORT

I have seen in several articles, the use of CIBERSORT to calculate the proportion of immune system cells using gene expression data, I would like to try it but I have not found how to install it for R,...
María José's user avatar
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1 answer
125 views

HISAT2: RNA strandedness

My library is unstranded and the code I'm trying to use is this: ...
Neophytos Kouphou's user avatar
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29 views

Calculating p-value and adjusted p-value for pre-normalized microarray data with fold change precalculated

I have a microarray dataset with two mutants dataset that has already been normalised, and the fold change values for each gene in each mutant versus the wild type have been calculated. I'm interested ...
shaimaa Hassan's user avatar
3 votes
0 answers
75 views

how to install cibersort in R, to do deconvolution using my RNA-seq Data with public single cell data

I have RNA-seq Data from lung cancer immunotherapy patients but i don't have any single cell data from my sample, so i wanted to do a deconvolution with a public single cell data. I thought of doing ...
Rita Soares's user avatar
3 votes
1 answer
47 views

Discrepancy with featurecounts analysis using a forward stranded and reverse stranded protocol

My RNAseq analysis pipeline is as follows: fastqc (read quality is good, some overrepresentation of adaptor sequence) → trimmomatic (trimmed adaptor sequence, qc report after trimming suggests the ...
xtian's user avatar
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3 votes
2 answers
107 views

Cannot obtain alignment summary after running Bowtie2

I am aligning my Small RNA Seq data with Bowtie 2. Although the alignment performs well, the only information I obtain after finishing running the alignment is the following: ...
ALEJANDRA PANDO CACIANO's user avatar
1 vote
0 answers
95 views

Working with Smart-seq3 data, have some questions regarding UMI and bar coding

Most of my experience has been working with genomic data, this time I am working on data from Smart-seq3. RNA excreted by cells in culture medium were processed with Smart-seq3 (From the looks of it, ...
Karthik Nair's user avatar
4 votes
2 answers
189 views

Question about umap using different numbers of pca components as initialization

I am new to the scRNA-seq field and I have been doing some experiments of visualization of UMAP using different numbers of PCA components for initialization. The process involves projecting scRNA-seq ...
Zack's user avatar
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1 answer
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scRNA: What are good dimensionality reduction/clustering parameters to get biologically plausible groupings?

I've got a moderately large set of PBMCs, over 1M cells. That means I can't easily do a grid search of dimensionality reduction/clustering parameters/methods. Some examples results I'm getting with ...
Henry Gong's user avatar
0 votes
1 answer
61 views

Paired end reads with different read lengths

I downloaded some raw RNA-seq reads from NCBI using the sra-toolkit (Accession numbers: SRR2053159-64). The reads were generated using ABI SOLiD, and in colour space. I downloaded them in base space ...
Arkajyoti Banerjee's user avatar
-1 votes
1 answer
170 views

Using CSV files continaing scRNA-seq count data from GEO [closed]

I'm completely new to scRNA-seq analysis (and to most of Bioinformatics as well) so I apologize if this was asked a million times before. I am trying to get the single cell expression data of lymph ...
Newbie Bioinformatician's user avatar
1 vote
1 answer
69 views

What are the output files of RNA-Seq from facility?

This question was also asked on Reddit I am new in our lab and I am going to do bulk RNA-Seq. What type of files will we get from the company (Genewiz)? Will it be a bunch of Fastq files? or they give ...
Sina Asadi's user avatar
-1 votes
1 answer
55 views

Is there a data analysis software (free) or code that I can use to view normalized CDR3 amino acid length distributions?

Is there a data analysis software (free) or code that I can use to view normalized CDR3 amino acid length distributions? Regarding code, preferably using Python or R. EDIT: Added a picture of the non-...
SData11's user avatar
3 votes
1 answer
125 views

Applying glmnet to identify predictors for subtypes

I am new to glmnet and other ML techniques, so apologise in advance if this sounds a trivial question. However, your guidance is very much appreciated. So, I have nearly 700 genes with their ...
Angelo's user avatar
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3 votes
1 answer
121 views

Issue with Differential Gene expression analysis with Deseq2

I am using the Deseq2 package to perform a DE analysis on multiple factors. So, basically to perform the analysis I consider patients characterized by the Brugada Syndrome type 1 and other patients ...
Alessia Avon's user avatar
2 votes
1 answer
58 views

DE analysis tool and method for non-coding features

I am currently working with the non-coding features of A. thaliana, and trying to get the DE features. Among the three DE test methods in edgeR such as ...
Arkajyoti Banerjee's user avatar
2 votes
0 answers
24 views

Feasible to find genetic variations of two samples using RNAseq data?

I have bulk RNAseq data from two strains of mice from Jackson Lab: C57BL/6 and B6.SJL. The former expresses a Ptprc-b allele and the latter expresses a ...
geom_na's user avatar
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0 votes
1 answer
112 views

is it possible to count cell in Violin plot in seurat?

I tried to quantify the number of cells of Fzd9+ Macrophage. how can I know the cell count? thank you
kiseoking's user avatar
1 vote
1 answer
59 views

Filtering criteria for non-coding features with very low counts

I am trying to do DE analysis of non-coding features of A. thaliana. I find in the miRNA and lncRNA counts file that they are abundant in zero counts, and most of the non-zero counts are very low. Now,...
Arkajyoti Banerjee's user avatar
1 vote
1 answer
40 views

Statistical methods suitable for DE analysis of non coding RNAs

I am currently working on DE analysis of coding as well as non-coding features of A. thaliana using the edgeR package. Is the negative binomial method that is ...
Arkajyoti Banerjee's user avatar
1 vote
0 answers
78 views

building custom gtf file with entrez ID for any organism

How do I generate a custom gtf file format for a organism of my interest similar to human gtf file, in the custom gtf file I want to add entrez id to the new custom gtf. My organism is ...
kcm's user avatar
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23 views

Gene Expression Database for Psychological Disorders

Is there a comprehensive database available online of gene expression across the brain taken from individuals with psychological conditions? Resources like the Allen Institute's Human Brain Map (link ...
Lambda's user avatar
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2 votes
1 answer
228 views

Correlation heatmap of RNA-seq clusters all samples together leading to very low no. of DEGs

I am writing to you to take an input or may be you can provide a different perspective. I am at wits end :( So I have nearly 200 samples. These are separated into two groups (group I treated with ...
Angelo's user avatar
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0 votes
1 answer
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Looking for datasets of RNA-Seq (or scRNA-Seq) for L1 larvae C. elegans

I was wondering if anyone know of a repository for C. elegans data sets, specifically in L1 larvae stage. Are there any papers or GEO-sets for this organism at this stage? thanks Assa
Assa Yeroslaviz's user avatar
2 votes
2 answers
291 views

Integrate bulk RNA and ATAC-seq genes

I have up and down-regulated genes from bulk RNA result from DeSeq2. Genes that are differential in chromatin accessibility between two conditions from DiffBind. I would like to find the genes overlap ...
Chris's user avatar
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0 votes
1 answer
59 views

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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