Questions tagged [single-cell]

Questions about techniques in which the genetic material of a single cell is analysed individually

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56 views

semantic similarity measurement for cell line ontologies

I have a set of cell line pairs and I want to know to what extent the pairs are similar based on their ontologies. The problem I have is that I have found a Python library called Fastsemsim, but it ...
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1answer
404 views

Cluster is split in 2-3 locations on tsne plot - Suerat

I am running a single cell dataset (count data - exon) through Seurat. After running tsne I see a cluster (13) split in 3 different locations on the plot. Here are the commands I am running: ...
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2answers
1k views

How to create Seurat object while RNA expression and ADT combined into one matrix

I got an input matrix which RNA expression and ADT capture are combined into one file. I loaded the file into Seurat successfully, however, when I tried to create Seurat object, it threw out an error ...
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1answer
1k views

How to highlight specific cells in Seurat 2.4

I used Seurat 2.4 on our scRNA dataset to obtain the following tSNE plot.I was able to successfully extract cell IDs from the different clusters, and generate gene expression profiles. The analysis, ...
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3answers
1k views

Import gene list to Seurat to define cell types

Is there a way to import gene list into Seurat to define cell type? The default cell types in Seurat is not enough for our research. For example, we want to mark a subtype of B cells in Seurat, but ...
5
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1answer
783 views

PCA vs tSNE in single cell RNA-seq

What makes tSNE being the preferred dimensional reduction for visualization in single cell RNA-seq over PCA? I am aware that tSNE works better at showing local structures and fails to capture global ...
1
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1answer
407 views

Which values from Seurat::FetchData function are to be used for correlation analysis between genes?

I want to perform a correlation test between genes in on my single cell RNA seq data set. I perfomed the differential expression analysis using the Seurat version 2 package, after performing stages of ...
0
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1answer
53 views

how to write the script to know each sample's cluster number?

I have followed the Seurat Tutorial:Integrating stimulated vs. control PBMC datasets to learn cell-type specific responses by using my data. And I am confused about how to write the script to ...
2
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2answers
223 views

Why are the clusters in the left side of a tSNE plot and not in the centre?

I have no idea why I used the subsetdata in Seurat after using CCA the TSNEPlot shows the cluster all in the left side and merge, did I miss something? Here are the code and figure I got. ...
2
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1answer
217 views

Why does my SplitDotPlotGG plot show differential expression of a gene but Seurat::FindMarkers() function does not return it?

I have analysed and clustered my single cell rna seq data with methods in the Seurat package. After clustering, I obtained two main clusters: 0 and 1. I used the Seurat SplitDotPlotGG function to ...
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1answer
531 views

GEOquery errors due to list input

I want to extract the expression matrix from the GSE object in GEOquery. ...
0
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1answer
2k views

how to obtain only one cluster's data (.csv file) in Seurat

I was using Seurat to analysis single-cell RNA Seq. And I was interested in only one cluster by using the Seurat. Does anyone know how to achieve the cluster's data(.csv file) by using Seurat or any ...
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2answers
110 views

scRNA-seq differential transcript usage

Many of the modern gene-quantification tools (Salmon/Kallisto) output transcript-level (as opposed to gene-level) data. All of the scRNA-seq analysis I have seen just uses the gene-level values. I ...
0
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1answer
528 views

how to write italic script in Rstudio [closed]

I have got the problem on how to write the italic script in RStudio. I used the script in the screenshot and got the error "Error in grobs[[i]]: subscript out of bounds."
5
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1answer
690 views

How to normalise scRNASeq data for differential expression analysis

I wish to perform differential expression analysis for cluster-specific gene expression in single-cell data (with a tool such as MAST or SCDE). I have data for 3 biological replicates. I performed ...
2
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3answers
1k views

scRNA-seq,10x cellranger pipelines,low custom tdtomato gene content! looking for help!

I have a set of scRNA-seq samples expressing TdTomato, which has high content in microscope. I followed the 10x cellranger pipelines to finsh the work, my procedures are as follows: added TdTomato on ...
4
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0answers
63 views

Database of Cell Volumes by Cell Ontology? [closed]

Are there any databases that have aggregated information on cell volumes? Most useful would be a database keyed by cell ontology (e.g., CL:0000236 for B cell). ...
5
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1answer
531 views

Derive a GTF containing protein coding genes from a GTF file with Exons and CDS

Why I need a compatible file I’m trying to run velocyto with the R package to analyse RNA velocity (cell trajectories) with single cell RNASeq data. I have performed single cell analysis from 10x ...
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2answers
1k views

Using Seurat to compare mutant vs.wt

I am interested in using Seurat to compare wild type vs Mutant. I don't know how to use the package. How can I test whether mutant mice, that have deleted gene, cluster together?
2
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2answers
214 views

Can I get the graph generated by cellranger

I’ve run the cellranger analysis pipeline on single cell RNASeq datasets. I can import the matrix and graph-based clusters into R. Doing this I can optimise the dimension reduction and plot cells with ...
2
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1answer
44 views

What does that mean if there is no bifurcations in my time series single cell data?

Sorry, I have analysed my time series single cell data with Scuba algorithm but that says that there is no bifurcations in my data and outputted a single chain of 4 states with no branching like ...
4
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2answers
499 views

Collapse cell barcodes distribution within 1 Hamming distance

I have a barcode distribution from single-cell data, e.g: ...
3
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2answers
769 views

Changing a wide range of colours to a limited gradient

I returned a FeaturePlot from Seurat to ggplot. My plot has a weird range of colours as below I produced this plot by this code ...
3
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1answer
721 views

about dotplot legend meaning

Here is the code I used: ...
3
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0answers
51 views

What is the correct way of dealing with the analysis of data from flow cytometry?

I would like to detect a change in expression of a molecule present on a cell type by flow cytometry. Assuming I am able to detect, using an antibody, a signal that represents the amount of the ...
3
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1answer
52 views

Why is cell count stated relative to animal size?

Why do some single-cell RNA-seq papers give the number of cells sampled as a fraction of the size of the organ, animal, or embryo, as opposed to just saying how many cells they sampled? For instance,...
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1answer
93 views

How is PCR duplication rate computed in scATAC-seq?

Reading Cusanovich et al. (2015) I encountered the sentence: We mixed pairs of cell lines (HEK293T or HL-60 versus GM12878), performed combinatorial cellular indexing, and sequenced the resulting ...
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1answer
2k views

Violinplot of gene expression

I have plotted the log normalized expression of two genes by violonplot for 4 clusters. I have links to my pictures and Seurat object too. My problem is this; in violin plot I can not see the mean or ...
3
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3answers
4k views

Mapping a list of cells in seurat featureplot

I have 209 cells, I clustered them by Seurat to 4 clusters. By Featureplot I am able to track a gene in clusters: Higher color shows higher expression. Now, for some genes I want to highlight some ...
7
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1answer
95 views

Network comparison of single cells (from sequencing data)

What methods can we use to compare networks (PPI, gene regulatory etc) created from single-cell gene expression (or proteomics) data? There are methods that construct networks by comparing two ...
5
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3answers
9k views

Resolution parameter in Seurat's FindClusters function for larger cell numbers

In Seurats' documentation for FindClusters() function it is written that for around 3000 ...
5
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1answer
168 views

Which are the use cases for the methods for DE in Seurat

In Seurat we can specify multiple methods for finding DE genes. I am wondering when should we use which one, the use cases. As ...
4
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2answers
2k views

What is cellranger doing in comparison to other methods?

I've recently started working with the 10X-Genomics platform with Illumina (MiSeq and HiSeq) for single-cell RNA-Seq. I've been recommended the "cellranger" (version 2.1.0) which I understand handles ...
5
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2answers
9k views

Manually define clusters in Seurat and determine marker genes

I want to define two clusters of cells in my dataset and find marker genes that are specific to one and the other. Is there a way to do this in Seurat? Say, if I ...
4
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1answer
2k views

Subset on multiple genes in Seurat

I know that I can do subsetting on just one gene in Seurat: ...
5
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3answers
1k views

Regressing out unwanted sources of variation in single cell RNA-seq data

I have a dataset of single cell count data and I want to regress out the variation caused by the number of UMI's and the percentage of mitochondrial genes. I know that count data is discrete data, ...
11
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2answers
3k views

determining doublets in single-cell RNA-seq

Doublets are a known problem with scRNA-seq experiments, where 2 or more cells are sometimes captured instead. To determine their presence, there are studies that mix multiple species (such as human ...
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”...
6
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1answer
209 views

Scaling by linear regression against the number of reads

I am trying to build the preprocessing pipeline presented in The Tabula Muris Consortium et al. (pp). It is a pipeline to preprocess single-cell sequencing data. There is one step that is not clear: ...
5
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1answer
471 views

Single-cell RNA sequencing (scRNA-seq): filtering cells by transcript counts, how to choose cutoffs?

I am running a notebook with example for the MAGIC algorithm. In the data preprocessing step, a filtering operation is required to filter out cells with a small count of transcripts. My question is ...
5
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1answer
76 views

How to compute the chance of failing to detect a gene given the detection limit of a protocol

In Shapiro et al., when discussing about loss of molecules as source of error in single-cell sequencing, it is written that: Another source of error is losses, which can be severe. The detection ...
2
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1answer
224 views

Mini-bulk RNA-seq and scRNA-seq analysis differences?

We have RNA-seq data from libraries prepared using the Smartseq2 single-cell protocol on 500 cells (mini-bulk) / library. The complexity is much better than with single-cells (~14k genes for 1.5M ...
7
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1answer
817 views

How to trim adapter sequences from GSE65360 in order to map the reads?

I am trying to map single-cell chromatin accessibility data from doi:10.1038/nature14590, obtained using scATAC-seq, to the reference genome. Example paired-end reads are here. The reads contain ...
19
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3answers
2k views

What is the actual cause of excessive zeroes in single cell RNA-seq data? Is it PCR?

First, sorry if I am missing something basic - I am a programmer recently turned bioinformatician so I still don't know a lot of stuff. This is a cross post with a Biostars question hope that's not ...
2
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3answers
93 views

Getting hierarchy of cell populations with Drop-seq data

I plan on collecting some drop-seq data from brain tissue (from mice and humans). Using only the 75-85% of genes that have 1:1 orthology between mice and humans, I'd like to cluster the cells by ...
2
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1answer
2k views

Cellranger aggr and Seurat merge difference?

I have multiple libraries of 10x Chromium single-cell RNA-seq data, which I'd like to combine. One option is using cellranger aggr which by default does a depth ...

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