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Galaxy Training Network/galaxy-single-cell

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Björn Grüning

@bgruening

what do you see locally?

is it spawning 8 threads when you have 8 CPUs?

Mehmet Tekman

@mtekman

I believe so yes

Carolyn Nielsen

@carolynnielsen

Hi all - I'm looking to extract and analyse BCR sequences from my single cell SMART-Seq data. Are there tools for this in Galaxy? I've come across papers on Antigen Receptor Galaxy but this doesn't seem to be available on the instances I have checked. Are there alternatives? Or is there a way for me to import this (like a workflow) without having admin privileges? https://toolshed.g2.bx.psu.edu/repository?repository_id=2e457d63170a4b1c&changeset_revision=28fbbdfd7a87

Duaa Mohammad Alawad

@DuaaAlawad

how can I choose the root cell type from this dataset?

Mehmet Tekman

@mtekman

@DuaaAlawad which dataset are you referring to?

Duaa Mohammad Alawad

@DuaaAlawad

pbmc8k

Mehmet Tekman

@mtekman

so you don't typically choose the root/stem cell type, you infer it from the clustering. But you can rename the clusters to whatever you want by using the Rename Categories of Annotation option in the Manipulate Anndata tool

Carolyn Nielsen

@carolynnielsen

Any suggestions for workflows or tools in Galaxy for BCR sequence analysis from either SMART-Seq or 10X V(D)J data sets?

Mehmet Tekman

@mtekman

We're currently working on a SMART-Seq training -- but I think the following workflow should work: https://usegalaxy.eu/u/mehmet-tekman/w/smart-seq2

iMammal

@iMammal

When walking through the Alevin tutorial (https://training.galaxyproject.org/training-material/topics/transcriptomics/tutorials/droplet-quantification-preprocessing/tutorial.html) on the Human transcriptome "Comprehensive gene annotation" GTF file from Gencode (https://www.gencodegenes.org/human/) on the usegalaxy.eu server, the GTF2GeneList tools fails when the "Flag Mitochondrial Features" option is enabled (default parameters) with this error:

Error in [[<-(tmp, name, value = character(0)) :
0 elements in value to replace 258145 elements
Calls: $<- -> $<- -> [[<- -> [[<-
Execution halted

I tried the GTF with or without the "scaffolds, assembly patches and alternate loci" with the same error. Am I missing something or is this an actual bug? It seems to work fine on genes and transcripts but adding the mito features always makes it crash.

5 replies

Wendi Bacon

@nomadscientist

Is there a Plot with Seurat tool similar to Plot with Scanpy?

Pavankumar Videm

@pavanvidem:matrix.org

[m]

Not that I know. I can only see a plot tool for dimension reduction for seurat on Galaxy.

Helena

@hexylena:matrix.org

[m]

Hey GTN Teachers and Trainers! As many of you know, we're hosting the GTN Smörgåsbord 2022 Edition: Tapas! (Or GTN Tapas for short), happening March 14-18, 2021, all timezones (24/7 event)

Do you want to teach something at GTN Tapas? Are you curious what the draft schedule looks like? Want to help out? We've collected all of the information for presenters at this year's event and you might want to join the mailing list

Like last year, this event will be completely online, asynchronous, and based around discussions in Slack. Action Item for Presenters: We have a draft schedule in that page, please check it.

Let us (Saskia, myself, anyone from wg-GOAT) know if you have any questions!

psaunders1

@psaunders1

I am trying to run the Galaxy tutorial on processing single cell data with Alevin and am hitting a roadblock. The Alevin runs fine except that the quants_tier_mat has a metadata failure, but this file is not being used for the next step. When I try to do salmonKallistoMtxTo10x the program fails to initiate, reporting a "An error occured with this dataset. Conda dependency seemingly installed but failed to build job environment"

Pavankumar Videm

@pavanvidem:matrix.org

[m]

@psaunders1: if you are using EU Galaxy instance, please report the error from the Galaxy interface?

Wendi Bacon

@nomadscientist

Hello! Can you share your history please?

Carolyn Nielsen

@carolynnielsen

Hi all - I'm trying to get started with some single cell analysis of a 10X pilot and I've had the following error when trying to run RNA Star with the fastq files. Any thoughts?

Björn Grüning

@bgruening

@carolynnielsen can you please check if you find the parameter "soloBarcodeReadlength" in the tool?

Carolyn Nielsen

@carolynnielsen

Mentioned here - should this be disabled?

Pavankumar Videm

@pavanvidem:matrix.org

[m]

@carolynnielsen: also make sure that you are not switching the mate pairs in the input. Generally R1 is short in length. 26nt in 10x v2 chemistry and 28nt in v3 chemistry. The R2 is generally much longer and contains the actual sequenced RNA fragment. Checkout Figure 5 from this tutorial

Carolyn Nielsen

@carolynnielsen

Thanks @bgruening and @pavanvidem - pasting input below. I thought I had R1 and R2 the right way around- is this not the case? If looks ok, should I just disable checking the barcode read length and re-run? Error message says read length is 151bp.

Ebtihalmustafa

@Ebtihalmustafa

Hi,

I would like to know if there is any tutorial that can help me to analyse Feature barcode scRNA seq data ( CRISPR+scRNA ) by galaxy

Pavankumar Videm

@pavanvidem:matrix.org

[m]

Hi @Ebtihalmustafa Currently there is no tool that directly maps this data on Galaxy. Hopefully, STARSolo can do this in near future. I think it can be done on Galaxy with a little bit of effort. First, you can map your reads to the reference and then the unmapped to the guide RNAs fasta file. To map to the gRNAs, you have to manually create a GTF file of guide RNAs. There is a discussion on how to do this: alexdobin/STAR#1238

The downstream analysis workflow is in my TODO list. If you have any specific workflow that you're interested in, please let me know.

Ebtihalmustafa

@Ebtihalmustafa

Thank you very much for your response. For my feature barcode scRNA seq data workflow I would like to cluster the cells based on changes in the transcriptome and then identified in each cluster the enriched gRNA since in my experiment I knockout ~53 genes using 3 gRNA per gene

amini1573

@amini1573

Hello good time
I have a problem, in the Plot with scanpy tool the "Plot attributes" section does not work "Insert Component" option.Please help me

2 replies

Wendi Bacon

@nomadscientist

Hi folks! We’re having a meeting about wrapping tools for single cell analysis this Thursday at 3PM-London time. Direct message me if you’d like to join and are up for wrapping some tools! :)

bgruening

@bgruening:matrix.org

[m]

You might want to put this into the tools-wg and/or IUC channel

Wendi Bacon

@nomadscientist

Hey folks - I think this is probably something that applies to us (https://galaxyproject.org/events/2022-10-13-community-call/) as multiomics / single cell are friends

Pavankumar Videm

@pavanvidem:matrix.org

[m]

I guess this is physics muon, not single-cell muon

Wendi Bacon

@nomadscientist

oh damn, in the agenda it had ‘single cell’ so I was bamboozled

ah well, maybe we should have our own community call

Pavankumar Videm

@pavanvidem:matrix.org

[m]

https://www.youtube.com/watch?v=xhjyUBHlA_Y&ab_channel=GalaxyProject

deepika141296

@deepika141296

Hello All, I need help in performing trajectory analysis on single cell sequencing data. First I would like to combine data from different conditions what function should i use. Second do i have to label the clusters before combining the data?

1 reply

Wendi Bacon

@nomadscientist

Hi folks! I am getting an error using louvain clustering in a Jupyter Notebook in Galaxy (https://fd8602bf71bb0405-b77a49c1961e49cf974dfded0550ef6c.interactivetoolentrypoint.interactivetool.usegalaxy.eu/ipython/lab/tree/galaxy_inputs/Instructions/Trajectories_Instructions.ipynb). Can anyone figure out why this isn’t working? It’s bizarre because the louvain clustering works fine when performed as a Galaxy tool, just seemingly not when it’s in the Notebook/code form.

Pavankumar Videm

@pavanvidem:matrix.org

[m]

the error originated from here: ModuleNotFoundError: No module named 'leidenalg'

you need to install it

Wendi Bacon

@nomadscientist

Yeah thanks, that’s for that bit, but what about the Louvain step? sc.tl.louvain(adata,resolution=0.4)

Pavankumar Videm

@pavanvidem:matrix.org

[m]

sorry, I overlooked :)

1 reply

Pavankumar Videm

@pavanvidem:matrix.org

[m]

I tried to reproduce this and got the same error. Couldn't trace it back. Did you try with a different version?

Wendi Bacon

@nomadscientist

Yeah like a year ago :(

It worked then

maaryaam-gh

@maaryaam-gh

In the article that referenced the data that I am currently processing, it used the computeSumFactors method to normalize them.
And with my studies, I realized that this method is better for normalizing sparse count data
But I don't know if the tool introduced in the tutorial on Clustering 3K PBMCs with Scanpy for normalization (Normalize with scanpy) uses the same normalization method or not?
And if this is not the case, due to the fact that my data is a type of sparse data and normalization by the computeSumFactors method is mandatory for them.
Does Galaxy have a tool to normalize my data in this way?

1 reply

Pavankumar Videm

@pavanvidem:matrix.org

[m]

FYI, sran normalization is now available on EU Galaxy instance since a few weeks

Sheryuu

@Sheryuu

Guys, I ran into a problem I don't know how to solve: "cannot read workspace version 3 written by R 4.0.5; need R 3.5.0 or newer
Execution halted”. This is an error when I use the Seurat tool to convert 10X data into Seurat objects, and I would appreciate it if a friend could tell me how to solve it!

1 reply

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