Questions tagged [scrnaseq]

Use this tag for questions related to single-cell RNA-seq.

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61 views

Why am I getting empty expression data from GEO?

I am trying to analyze the scRNAseq data from this study. In their Method section they write: The accession number for the RNA and DNaseq data reported in this paper is GEO: GSE116237. When I go ...
13
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1answer
197 views

scRNA-seq multi-dataset integration for small datasets

There have been a few methods proposed for integration (or batch correction) of scRNA-seq datasets, such as Seurat CCA, MNN Correct, Scanorama, and Harmony. The concern is generally about the maximum ...
1
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1answer
51 views

Challenging benchmarks for supervised learning on sparse scRNA-seq data

One challenging aspect of modeling scRNA-seq data is data sparsity, that is, scRNA-seq measurements typically suffer from large fractions of observed zeros (i.e. dropouts), where a given gene in a ...
6
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2answers
183 views

Why are TPMs per 10k or 100k in many scRNA-seq studies?

I noticed that many scRNA-seq papers normalize TPMs to 10k or 100k as opposed to 1M (as the abbreviation defines them). It doesn't really matter since you are just moving the decimal point, so why ...
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1answer
321 views

Seurat Violin Plot: Why do dots align in one row?

I am analyzing a rather small scRNAseq dataset (39797 features across 3522 samples) and I have observed that dots in the violin plot are plotted in one line next to each other rather than in a cloud ...
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3answers
983 views

isSpike function in SingleCellExperiment package is deprecated?

I am trying to follow a tutorial from Sanger institute (from May 2019) on analysis of single cell RNA Seq data. They use isSpike function to filter out ERCC (control) and MT (mitochondrial RNA) reads, ...
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1answer
102 views

Seurat clusters

I'm trying to show cell cycle regression and input sample metadata to my umap. I've been following Satija Lab's tutorials and have generated my umap but now want to specify which sample corresponds ...
1
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1answer
32 views

Is there a difference between any scRNA-seq visualization and pseudotime?

Is there a good definition of pseudotime? Some tools are clearly labeled as pseudotime and produce values along a trajectory, but there are more complex approaches that involve branching and ...
3
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1answer
1k views

Percentage distribution of cells in all clusters based on their treatment condition?

I have 2151 cells, I clustered them by Seurat to 5 clusters. With the code below, I am able to have the number of cells per cluster and per condition: ...
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0answers
35 views

Why could I be having only NAs for p_val and p_val_adj in Seurat DGE analysis?

I am doing a differential gene expression analysis to find out genes that are differentially expressed in one cell type from the lung in two conditions, control and idipathic pulmonary fibrosis (IPF). ...
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0answers
41 views

10x Single-cell RNA-Seq - Updating gene symbol with latest ensembl release - Ensembl IDs missing

I am trying to update the gene symbols provided by the 10x human precompiled reference (2020-A), using the mapping Ensembl ID to gene symbol provided in release 102. However, I found out that 56 of ...
2
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2answers
748 views

VlnPlot problem

I was confused about the VlnPlot that it shows like the screenshot. It only shows the dot but no distribution of the data, is that normal?
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2answers
1k views

calculateQCMetrics defunct, how to calculate the quality metrics by perCellQCMetrics

I am trying to follow a tutorial from Sanger institute (from May 2019) on analysis of single cell RNA Seq data. They use calculateQCMetric function to calculate the quality metrics, but I am getting ...
0
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2answers
3k views

Subsetting from seurat object based on orig.ident?

I am pretty new to Seurat. I want to subset from my original seurat object (BC3) meta.data based on orig.ident. however, when i use subset(), it returns with Error. ...
0
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3answers
1k views

Which correlation method to compute the correlation score between different clusters of Sc-RNAseq data?

I'm analyzing single cell rna-seq data and trying to compute the correlation score between different clusters. Wondering how to choose the correlation method("pearson" (default), "kendall", or "...
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0answers
41 views

Clusters in scRNA of Glioblastoma

There is a scRNA dataset on Glioblastoma from 10xgenomics. Unfortunately, there seems to be no research paper related to that data. Question 0 : May be there is paper ? How to find it ? I am not ...
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0answers
60 views

Genes associated with digestion in single cell RNASeq

I am reading Saunders et al. (2018). In the methods' subsection "ICA based analysis and clustering" (independent component analysis), a set of independent components from ICA is labelled as &...
2
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1answer
152 views

shiny code with seurat object

I am trying to create a shiny board for my scRNA seq data. I have saved my seurat objects adn I use following code as below: ...
1
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1answer
372 views

Set new Idents based on gene expression in Seurat and mix n match identities to compare using FindAllMarkers

I am relatively new to Bioinformatics and scRNA-seq data analysis. I am using Seurat V3 to analyze a scRNA-seq dataset in R. Currently, I have merged three scRNA-seq samples from the same donor into ...
1
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1answer
106 views

umap highlighting two different models

I'm trying to create a umap for single cell data from human samples and ptx samples. I can get the umap to where it shows the umap with the different clusters but I want to show where the ptx samples ...
1
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1answer
54 views

Bootstrapping cell type identifications (scRNA-seq)

I have scRNA-seq data held in a SEURAT object, and cell-types were identified with SingleR (essentially a vector of strings - ...
5
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4answers
2k views

Pathway level analysis of single-cell gene expression

I'm looking for single-cell specific methods to construct (using gene expression data) new features that express pathway "level" or "activity", and then use these for clustering cells. One example ...
3
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2answers
2k views

10X Illumina demultiplexing sample sheet issue

Also posted on biostars. I am trying to use cellranger or bcl2fastq to convert the .bcl ...
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1answer
1k views

How to add cluster name to metadata in Seurat?

I'm working on a Seurat object and want to name the clusters according to 2 values alone (yes/no). So I want to add a new column to metadata and annotate the clusters (UMAP) with it. ...
1
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1answer
618 views

Seurat FindMarkers() output interpretation

I am using FindMarkers() between 2 groups of cells, my results are listed but i'm having hard time in choosing the right markers. Do I choose according to both the p-values or just one of them? If one ...
1
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1answer
33 views

Is there a computational method/method/way to predict if a cell population in a tissue immigrated or is enriched locally?

I have single cell data which I have analysed for differential expression. In my experiment, I subjected two groups of mice (a control and a treatment group) to treatment that will lead to the ...
5
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4answers
432 views

Specific cell type identification in Single Cell Sequencing

In order to define which cell is of which type we need to identify a set of rules, for instance neurons should express one of the following: Thy1, ...
5
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2answers
247 views

Assign cell types to groups of cells based on their gene expression profiles

I have large filtered, normalized dataset of scRNA-seq data of C.Elegans species. Rows are genes (10 000), columns are cells (66 ...
4
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0answers
51 views

Simulating 3' end tag-based scRNA-seq reads

Are there any tools that will simulate 3' end tag-based single-cell RNA-seq reads? For example, Drop-seq, 10X Chromium, ...
1
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1answer
3k views

Changing active.ident in Seurat

Im trying to change the active.ident to another column in metadata but this error keeps popping up! I recently upgraded to R version 4.0.2 from 3.6.1 The older version was working but the new one isn'...
0
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0answers
43 views

Alternatives to imputation for single-cell RNA-Seq analysis to increase statistical power of DE analysis

I have a small scRNA-Seq dataset (n = 357, inhibitory neurons). This set of cells is split almost evenly between two conditions (Case and Control). I would like to test for differential expression ...
0
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1answer
474 views

Adding treatment groups via metadata to Seurat object?

I'd like to add metadata to 6 individual Seurat objects so that after I merge the objects into one, I can later label or split by using these identifiers. For example, I'd like to append an age group ...
2
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0answers
90 views

scRNA-Seq: Account for sequencing depth and gene length?

Task: Normalize a single-cell RNA-Seq dataset to account for sequencing depth and gene-length. For UMI-count based protocols (like 10x) that don't suffer from gene-length biases, there are various ...
2
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0answers
54 views

gene set enrichment in single-cell data

Gene set or pathway analysis in scRNA-seq data has its own challenges. Since the data is sparse and many genes will be missing for any given cell, some people simply add up all the counts across the ...
6
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2answers
4k views

What are doublets in single cell RNA-seq data?

I am reading The Tabula Muris Consortium et al. (pp). In some organs, cells with more than 2 million reads were also excluded as a conservative measure to avoid doublets. How exactly is a “doublet”...
2
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3answers
370 views

How to reduce the occupied RAM when you are dealing with a very sparse matrix in a single-cell Experiment in R?

I'm dealing with a very large and sparse dataset and the first issues I met occurred when I tried to use quickCluster that reported me this error: ...
2
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2answers
84 views

Most scalable pseudotime ordering algorithm

What algorithms for linear Pseudotime trajectory construction (diffusion-based) are the most scalable to large datasets? I'm currently using Slingshot based on the recommendation in this manuscript: ...
0
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1answer
120 views

How do I perform a pathway related grouping of genes?

I have a list of gene names from the analysis of my mouse single seq data. I want to perform a simple grouping of the genes based on the pathways to which they belong or pathways through which they ...
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0answers
19 views

minimum number of neighbors for scRNA-seq analysis

Many scRNA-seq clustering/visualization methods are based on nearest neighbors. The maximum number of neighbors is limited by the neighborhood size. Is there a method to determine the minimum number ...
2
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0answers
72 views

Clustering issue about overlapping CD4, CD8, and Foxp3 genes in mouse tumor models

bioinformatics scientists. I hope your work goes smoothly well and be safe. I'm stuck in trouble at the step of clustering. Briefly, I downloaded fastq files from a published study (https://www.ncbi....
0
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1answer
1k views

Error: Insufficient values in manual scale. 24 needed but only 1 provided?

folks. Can you give me idea about an issue that I encountered? I ran single-cell RNA sequencing data using Suerat in R, and when I tried to draw violin plot, there is an error as below. VlnPlot(...
0
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1answer
387 views

PCA plot did not work in single cell RNA-seq

I run plotPCA for single cell RNA-seq data, while I get error message (I use R 4.0). I attached the code and error message here. Did anyone know the reason and how to deal with it. Thanks! ...
0
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1answer
442 views

How to remove zero value of gene on FeatureScatter plot using Seurat?

Nowadays, I am trying to calculate Pearson correlation values between two genes of my interest from single-cell RNA-data (features.tsv, barcodes.tsv, and matrix.mtx files) which are obtained from the ...
3
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1answer
587 views

Receptor-Ligand R-package for visualization

I was wondering if there was any R-package for receptor-ligand interaction similar to SingleCellSignal (https://github.com/SCA-IRCM/SingleCellSignalR_v1/blob/master/SingleCellSignalR/vignettes/...
0
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1answer
246 views

Is it important to filter out poor quality cells before performing an integration analysis on single cell RNA sequencing data?

In order to perform an integration analysis of single cell RNA seq data, is it important to check the percentage expression of mitochondrial genes of cells as well as the feature counts to exclude ...
0
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0answers
29 views

How do I generate a list of post-synaptic gene markers that will guide me in my search for these markers in intestinal stem cells?

I would like to generate a list of gene markers that represent post-synaptic related genes. I will then use this list to search through single cell sequencing data, in order to assess whether certain ...
3
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2answers
235 views

Differential gene expression bias due to effect of an individual sample

I am analysing a human single cell RNA seq experiment, where we have 4 groups, four samples each. Data has been analysed using Seurat, with the canonical workflow. I have tried DE using various ...
3
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3answers
2k views

How do I pull singe cell RNA sequencing data from GEO database?

I am new to R and computational biology. I am trying to look through a published data set to check for gene expression for my own project. I am having trouble finding materials to teach me how to ...
0
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1answer
50 views

Is there a way to get the code from "github.mit.edu"?

everyone. I am a postdoc who just begins to analyze single-cell RNA-seq data. Nowadays, I found a really interesting paper (https://doi.org/10.1016/j.celrep.2018.10.047, PMID: 30404002). So, I ...
0
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1answer
36 views

Is there a single cell RNAseq equivalent of GTEx or TCGA? [closed]

Or do I need to find individual studies and obtain data the long way!