### LoupeR Setup Prompt Source: https://www.10xgenomics.com/support/software/loupe-browser/latest/tutorials/introduction/lb-louper Example of the prompt displayed when setup is required. ```text Please call `setup()` to agree to the EULA before continuing ``` -------------------------------- ### Run setup() command Source: https://www.10xgenomics.com/support/software/loupe-browser/latest/tutorials/introduction/lb-louper Execute the setup function to proceed with the installation or configuration. Type 'y' and ENTER to agree to the 10x EULA. ```bash setup() ``` -------------------------------- ### Install LoupeR via GitHub Source: https://www.10xgenomics.com/support/software/loupe-browser/latest/tutorials/introduction/lb-louper Commands to install LoupeR directly from GitHub and perform initial setup. ```R remotes::install_github("10xGenomics/loupeR") loupeR::setup() ``` -------------------------------- ### Example SSO Configuration Inputs Source: https://www.10xgenomics.com/support/software/xenium-explorer/latest/tutorials/open-file-aws Example inputs for the `aws configure sso` command, including SSO start URL and region. ```bash SSO session name (Recommended): WARNING: Configuring using legacy format (e.g. without an SSO session). Consider re-running "configure sso" command and providing a session name. SSO start URL [None]: https://your-organization.awsapps.com/start SSO Region [None]: us-west-2 ``` -------------------------------- ### Set Up Authentication Source: https://www.10xgenomics.com/support/software/cloud-analysis/latest/tutorials/CA-cloud-cli-documentation-for-linux Initiate the authentication process by running `txg auth setup`. This command guides you through obtaining and saving an access token for your 10x Genomics Cloud account. ```bash txg auth setup ``` -------------------------------- ### Download Example FASTQ Files Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/tutorials/cr-tutorial-multi-beam-t Download the compressed FASTQ files for the example BEAM-T dataset using curl. ```bash curl -O https://cf.10xgenomics.com/samples/cell-vdj/7.1.0/5k_BEAM-T_Human_A0201_B0702_PBMC_5pv2_Multiplex/5k_BEAM-T_Human_A0201_B0702_PBMC_5pv2_Multiplex_fastqs.tar ``` -------------------------------- ### Create and navigate to the working directory Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/tutorials/cr-tutorial-vdj Initializes the project folder structure for the tutorial. ```bash mkdir dataset-vdj-practice/ cd dataset-vdj-practice/ ``` -------------------------------- ### Create and enter working directory Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/tutorials/cr-tutorial-multi Initializes the project workspace by creating a directory and navigating into it. ```bash mkdir dataset-multi-practice cd dataset-multi-practice ``` -------------------------------- ### Install and Initialize Claude Code Source: https://www.10xgenomics.com/support/software/cloud-analysis/latest/tutorials/cloud-mcp-server-code Install the Claude Code CLI tool globally via npm and launch the interactive setup process. ```bash # Install Claude Code npm install -g @anthropic-ai/claude-code # Open Claude Code and follow prompts to set up and authorize account claude ``` -------------------------------- ### Create a new project Source: https://www.10xgenomics.com/support/software/cloud-analysis/latest/tutorials/CA-cloud-cli-documentation-for-linux Initializes a new project with a unique name and optional description. ```bash txg projects create --name NAME [--description DESC] ``` -------------------------------- ### Create and navigate to project directory Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/tutorials/cr-tutorial-multi-beam-ab Initializes a new directory for the analysis project. ```bash mkdir beam-ab cd beam-ab ``` -------------------------------- ### 5' CRISPR Guide Capture Feature Reference CSV Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/inputs/cr-feature-ref-csv Example Feature Reference CSV for a 5' CRISPR Guide Capture library. The 'pattern' must be reverse complemented to match the guide RNA sequence in the read. ```csv id,name,read,pattern,sequence,feature_type,target_gene_id,target_gene_name ACTR8-1,ACTR8-1,R2,TTCCAGCTTAGCTCTTAAAC(BC),TCTCCTTCTCGCCGCCCTTC,CRISPR Guide Capture,ENSG00000113812,ACTR8 ACTR8-2,ACTR8-2,R2,TTCCAGCTTAGCTCTTAAAC(BC),CCCTTCTCCTTTCCGTTCTC,CRISPR Guide Capture,ENSG00000113812,ACTR8 BCL2-1,BCL2-1,R2,TTCCAGCTTAGCTCTTAAAC(BC),CACCGGGCATCTTCTCCTCC,CRISPR Guide Capture,ENSG00000171791,BCL2 BCL2-2,BCL2-2,R2,TTCCAGCTTAGCTCTTAAAC(BC),GGAGATAGTGATGAAGTACA,CRISPR Guide Capture,ENSG00000171791,BCL2 NEG_CTRL-1,NEG_CTRL-1,R2,TTCCAGCTTAGCTCTTAAAC(BC),TACATCGCACCCCCCCGGTC,CRISPR Guide Capture,Non-Targeting,Non-Targeting NEG_CTRL-2,NEG_CTRL-2,R2,TTCCAGCTTAGCTCTTAAAC(BC),TATAGTCGTCACTAGTACAC,CRISPR Guide Capture,Non-Targeting,Non-Targeting ``` -------------------------------- ### 3' CRISPR Guide Capture Feature Reference CSV Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/inputs/cr-feature-ref-csv Example Feature Reference CSV for a 3' CRISPR Guide Capture library. The 'pattern' column is used directly. ```csv id,name,read,pattern,sequence,feature_type,target_gene_id,target_gene_name ACTR8-1,ACTR8-1,R2,(BC)GTTTAAGAGCTAAGCTGGAA,GAAGGGCGGCGAGAAGGAGA,CRISPR Guide Capture,ENSG00000113812,ACTR8 ACTR8-2,ACTR8-2,R2,(BC)GTTTAAGAGCTAAGCTGGAA,GAGAACGGAAAGGAGAAGGG,CRISPR Guide Capture,ENSG00000113812,ACTR8 BCL2-1,BCL2-1,R2,(BC)GTTTAAGAGCTAAGCTGGAA,GGAGGAGAAGATGCCCGGTG,CRISPR Guide Capture,ENSG00000171791,BCL2 BCL2-2,BCL2-2,R2,(BC)GTTTAAGAGCTAAGCTGGAA,TGTACTTCATCACTATCTCC,CRISPR Guide Capture,ENSG00000171791,BCL2 NEG_CTRL-1,NEG_CTRL-1,R2,(BC)GTTTAAGAGCTAAGCTGGAA,GACCGGGGGGGTGCGATGTA,CRISPR Guide Capture,Non-Targeting,Non-Targeting NEG_CTRL-2,NEG_CTRL-2,R2,(BC)GTTTAAGAGCTAAGCTGGAA,GTGTACTAGTGACGACTATA,CRISPR Guide Capture,Non-Targeting,Non-Targeting ``` -------------------------------- ### GEX + CRISPR Guide Capture Libraries CSV Example Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/inputs/cr-libraries-csv Configure libraries for Gene Expression and CRISPR Guide Capture experiments. Ensure FASTQ file paths are absolute. ```csv fastqs,sample,library_type, /opt/foo/,GEX_sample1,Gene Expression, /opt/foo/,CRISPR_sample1,CRISPR Guide Capture, ``` -------------------------------- ### Human and Mouse Reference Setup Source: https://www.10xgenomics.com/support/software/cell-ranger/downloads/cr-ref-build-steps Initializes environment variables and directories for human and mouse reference builds. ```bash #################### SETUP #################### human_genome="GRCh38" mouse_genome="GRCm39" version="2024-A" build="GRCh38_and_GRCm39_GENCODEv44-GENCODEvm33" mkdir -p "$build" # Download source files if they do not exist in reference_sources/ folder source="reference_sources" mkdir -p "$source" ``` -------------------------------- ### GEX + Antibody Capture + CRISPR Guide Capture Libraries CSV Example Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/inputs/cr-libraries-csv Set up libraries for experiments combining Gene Expression, Antibody Capture, and CRISPR Guide Capture. Verify FASTQ file paths. ```csv fastqs,sample,library_type, /opt/foo/,GEX_sample3,Gene Expression, /opt/foo/,Ab_sample3,Antibody Capture, /opt/foo/,CRISPR_sample3,CRISPR Guide Capture, ``` -------------------------------- ### Create and navigate to working directory Source: https://www.10xgenomics.com/support/software/space-ranger/latest/tutorials/setup-spaceranger Initializes a dedicated directory for the Space Ranger tutorial. ```bash # Create working directory mkdir spaceranger_tutorial # Change directory cd spaceranger_tutorial ``` -------------------------------- ### Spaceranger testrun expected output Source: https://www.10xgenomics.com/support/software/space-ranger/latest/tutorials/setup-spaceranger This is an example of the expected output when `spaceranger testrun` completes successfully, indicating a valid installation. ```text Martian Runtime - v4.0.5 Running preflight checks (please wait)... Checking sample info... Checking FASTQ folder... Checking reference... Checking reference_path Checking optional arguments... ... Pipestance completed successfully! ``` -------------------------------- ### Create and Navigate to Directory Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/tutorials/cr-tutorial-multi-beam-t Use these commands to create a new directory for your analysis and change into it. ```bash mkdir beam-t cd beam-t ``` -------------------------------- ### Cell Ranger vdj Pipeline Output Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/running-pipelines/cr-5p-vdj Example output indicating the start of the cellranger vdj pipeline stages after preflight checks. ```bash Martian Runtime - v4.0.8 Running preflight checks (please wait).... yyyy-mm-dd hh:mm:ss [runtime] (ready) ID.sample345.SC_VDJ_ASSEMBLER_CS.VDJ_PREFLIGHT yyyy-mm-dd hh:mm:ss [runtime] (run:local) ID.sample345.SC_VDJ_ASSEMBLER_CS.VDJ_PREFLIGHT.fork0.chnk0.main yyyy-mm-dd hh:mm:ss [runtime] (ready) ID.sample345.SC_VDJ_ASSEMBLER_CS.VDJ_PREFLIGHT_LOCAL ... ``` -------------------------------- ### Prompting for a detailed multi analysis setup Source: https://www.10xgenomics.com/support/software/cloud-analysis/latest/tutorials/cloud-mcp-server Provide specific project details, sample information, and protocol references to help Claude generate an accurate configuration. ```text The analysis I want to run should be called: "Flex-GEX-multi", and I want it added to an existing project called "first-claude-mcp-CR-9-flex". I have 1 sample of single nuclei Flex Gene Expression human data with 2 lanes of sequencing. The library and chemistry type is GEM-X Flex Gene Expression. Fresh frozen human kidney tissue was obtained from Avaden BioSciences by 10x Genomics. The tissue was sectioned into two 44 mg samples and nuclei were isolated using the Chromium Nuclei Isolation Kit (CG000505). A total of 4,180,000 nuclei were obtained, which were aliquoted into a sample containing approximately 2 million nuclei. The sample was fixed for one hour at room temperature using the Fixation of Cells & Nuclei for GEM-X Flex Gene Expression protocol (CG000782). Following fixation, probe hybridization was performed according to the user guide. 300,000 nuclei aliquots were used for hybridization. After hybridization, the samples were washed and filtered according to the protocol. The sample was then resuspended, counted, and loaded into a Chromium GEM-X Chip. 4,000 nuclei were targeted. Gene Expression libraries for GEM-X Flex were generated using the GEM-X Flex Gene Expression Reagent Kit for Singleplex samples (CG000786). Libraries were sequenced on an Illumina NovaSeq 6000, with a mean read depth of approximately 20,000 reads per cell using a paired-end, dual indexing sequencing scheme. The FASTQ files are already uploaded to this project: "first-claude-mcp-CR-9-flex". I want to run the Cell Ranger multi pipeline. Use the support documentation from these websites to help me set up an analysis: https://www.10xgenomics.com/support/software/cloud-analysis/latest and https://www.10xgenomics.com/support/software/cell-ranger/latest. Are there any parameters for the multi pipeline that I should consider for my analysis? ``` -------------------------------- ### Verify Installation with Test Run Source: https://www.10xgenomics.com/support/software/cell-ranger-atac/downloads/installation Execute the testrun command to validate the installation, which may take up to 60 minutes. ```bash $ export PATH=/opt/cellranger-atac-2.1.0:$PATH $ cellranger-atac testrun --id=tiny cellranger-atac testrun 2.1.0 Copyright (c) 2018 10x Genomics, Inc. All rights reserved. ------------------------------------------------------------------------------- Running Cell Ranger ATAC in test mode... Martian Runtime - 4.0.7 Running preflight checks (please wait)... 2018-09-17 20:44:33 [runtime] (ready) ID.tiny.SC_ATAC_COUNTER_CS.SC_ATAC_COUNTER._BASIC_SC_ATAC_COUNTER._ALIGNER.SETUP_CHUNKS 2018-09-17 20:44:33 [runtime] (run:local) ID.tiny.SC_ATAC_COUNTER_CS.SC_ATAC_COUNTER._BASIC_SC_ATAC_COUNTER._ALIGNER.SETUP_CHUNKS.fork0.chnk0.main ... Pipestance completed successfully! Saving diagnostics to tiny/tiny.mri.tgz ``` -------------------------------- ### Sample Data: perturbation_efficiencies_by_feature.csv Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/outputs/cr-outputs-crispr-overview Example data from the perturbation_efficiencies_by_feature.csv file. This file details guide RNA combinations, target genes, log2 fold change, p-values, and cell counts. ```csv Perturbation, Target Guide, Log2 Fold Change, p Value, Log2 Fold Change Lower Bound, Log2 Fold Change Upper Bound, Cells with Perturbation, Mean UMI Count Among Cells with Perturbation, Cells with Non-Targeting Guides, Mean UMI Count Among Cells with Non-Targeting Guides Guide1|Guide2 Guide1 -2.487458049, 0.684604341, -3.8938847, -1.591207405, 37, 0.189189189, 3760, 1.199734043 Guide1|Guide2 Guide2 1.46887148, 0.273153378, -0.206506878, 2.354657695, 37, 0.054054054, 3760, 0.028723404 ``` -------------------------------- ### CRISPR Guide Capture cells_per_protospacer.json Example Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/outputs/cr-outputs-crispr-overview A JSON object mapping protospacer names to lists of cell barcodes where they were detected. Useful for understanding protospacer distribution across cells. ```json { "GFP": [ "AAACCCAAGCCGTTAT-1", "AAACCCATCTCAAAGC-1", "AAACGAACATTGCTTT-1", "AAACGCTAGCCGATCC-1", "AAAGAACAGAACAGGA-1", ... ], "Control_1": [ "AAACCCAAGCCGTTAT-1", "AAACGAACACCCTGAG-1", "AAACGAACAGTTGTCA-1", "AAACGAAGTATTCCTT-1", "AAACGAAGTCTGTCAA-1", ... ``` -------------------------------- ### Run bamtofastq with threads Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/miscellaneous/cr-bamtofastq Example command to convert a BAM file to FASTQ using eight threads. ```bash bamtofastq --nthreads=8 /path/to/mydata.bam /path/to/home/directory ``` -------------------------------- ### Create Analysis with Cell Ranger Arc Source: https://www.10xgenomics.com/support/software/cloud-analysis/latest/tutorials/CA-cloud-cli-documentation-for-linux Start a Cell Ranger Arc analysis by using the `analyses create cellranger-arc` command. This command handles both analysis setup and file uploads if required. ```bash analyses create cellranger-arc ``` -------------------------------- ### TotalSeq™-A Feature Reference CSV Example Source: https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/inputs/cr-feature-ref-csv Use this CSV format for TotalSeq™-A antibodies with Single Cell 3' v2 and v3 kits. The pattern '^(BC)' indicates the barcode is at the start of Read 2. ```csv id,name,read,pattern,sequence,feature_type TIGIT,TIGIT_TotalA,R2,^(BC),TTGCTTACCGCCAGA,Antibody Capture CD279,CD279_TotalA,R2,^(BC),ACAGCGCCGTATTTA,Antibody Capture CD127,CD127_TotalA,R2,^(BC),GGCACCCATGTCTTT,Antibody Capture CD56,CD56_TotalA,R2,^(BC),TTCGCCGCATTGAGT,Antibody Capture CD45RO,CD45RO_TotalA,R2,^(BC),CTCCGAATCATGTTG,Antibody Capture CD45RA,CD45RA_TotalA,R2,^(BC),TCAATCCTTCCGCTT,Antibody Capture CD45,CD45_TotalA,R2,^(BC),TAGCGTGGAGTAGTG,Antibody Capture CD25,CD25_TotalA,R2,^(BC),TTTGTCCTGTACGCC,Antibody Capture CD19,CD19_TotalA,R2,^(BC),CTGGGCAATTACTCG,Antibody Capture CD16,CD16_TotalA,R2,^(BC),AAGTTCACTCTTTGC,Antibody Capture CD15,CD15_TotalA,R2,^(BC),TTGGACGTGCGATCT,Antibody Capture CD14,CD14_TotalA,R2,^(BC),TCTCAGACCTCCGTA,Antibody Capture CD8A,CD8A_TotalA,R2,^(BC),GCTGCGCTTTCCATT,Antibody Capture CD4,CD4_TotalA,R2,^(BC),TGTTCCCGCTCAACT,Antibody Capture CD3,CD3_TotalA,R2,^(BC),CTCATTGTAACTCCT,Antibody Capture ``` -------------------------------- ### Download and Prepare Reference Sources Source: https://www.10xgenomics.com/support/software/cell-ranger-arc/downloads/ref-build-steps Downloads FASTA, GTF, and motif files if they do not already exist. Ensures the reference source directory is created. ```bash source="${genome}-${version}-reference-sources" mkdir -p "$source" fasta_url="http://ftp.ensembl.org/pub/release-98/fasta/mus_musculus/dna/Mus_musculus.GRCm38.dna.primary_assembly.fa.gz" fasta_in="${source}/Mus_musculus.GRCm38.dna.primary_assembly.fa" gtf_url="http://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_mouse/release_M23/gencode.vM23.primary_assembly.annotation.gtf.gz" gtf_in="${source}/gencode.vM23.primary_assembly.annotation.gtf" motifs_url="https://jaspar.genereg.net/download/data/2018/CORE/JASPAR2018_CORE_vertebrates_non-redundant_pfms_jaspar.txt" motifs_in="${source}/JASPAR2018_CORE_vertebrates_non-redundant_pfms_jaspar.txt" if [ ! -f "$fasta_in" ]; then curl -sS "$fasta_url" | zcat > "$fasta_in" fi if [ ! -f "$gtf_in" ]; then curl -sS "$gtf_url" | zcat > "$gtf_in" fi if [ ! -f "$motifs_in" ]; then curl -sS "$motifs_url" > "$motifs_in" fi ``` -------------------------------- ### Open and Inspect Zarr Files with Python Source: https://www.10xgenomics.com/support/software/xenium-onboard-analysis/latest/advanced/example-code Read Zarr files into NumPy N-dimensional arrays using the zarr Python library. This example demonstrates how to open a Zarr file stored in a zip archive and inspect its structure. Ensure zarr is installed. ```python # Import Python libraries # This script was tested with zarr v2.13.6, v2.18.3, and v3.0.5 import zarr import numpy as np # Function to open a Zarr file def open_zarr(path: str) -> zarr.Group: store = ( zarr.storage.ZipStore(path, mode="r") if path.endswith(".zip") else zarr.storage.LocalStore(path) ) return zarr.open_group(store=store, mode="r") # For example, use the above function to open the cells Zarr file, which contains segmentation mask Zarr arrays root = open_zarr("cells.zarr.zip") ``` -------------------------------- ### Setup Reference Genome Build Directories Source: https://www.10xgenomics.com/support/software/cell-ranger/downloads/cr-ref-build-steps Sets up build directories and defines variables for human and mouse reference genomes, including version and build path. ```bash human_genome="GRCh38" mouse_genome="mm10" version="2020-A" build="GRCh38_and_mm10-2020-A_build" mkdir -p "$build" ```