10x genomics rna dual index protocol. 1 assay. Sample index sequences are incorporated as the i5 and i7 index reads. 0%. Once sorting is complete, proceed immediately Jun 4, 2023 · For additional information on preparation of specific sample types, consult 10x Genomics Demonstrated Protocols available on the 10x Genomics Support site support. Refer to step 1. pdf The sample index sets are either dual index or single index. These files can be loaded onto their respective thermal cyclers allowing for execution of each The Reagent Kits and User Guides are not cross-compatible between different versions. Answer: The 3' v3. Chromium Next GEM Chip N Single Cell Kit* 80 rxns PN-1000357 | 16 rxns Chromium Next GEM Single Cell 5' v2 (Dual Index) User Guide Rev E 4 TOC Table of Contents Introduction 6 Chromium Next GEM Single Cell 5 ' Reagent Kits v2 (Dual Index) 7 10x Genomics Accessories 12 Recommended Thermal Cyclers 12 Additional Kits, Reagents & Equipment 13 Protocol Steps & Timing 15 Stepwise Objectives 16 . The Visium CytAssist Spatial Gene and Protein Expression workflow assays RNA levels and protein detection in intact formalin-fixed paraffin embedded (FFPE) tissue sections. Question: What assays is the Dual Index Kit TS Set A (PN-1000251) compatible with? Answer: The Dual Index Kit TS Set A (PN-1000251) is compatible with Visium for FFPE, Visium CytAssist for FFPE, Visium HD, and Fixed RNA Profiling. This protocol also provides guidance for enriching labeled cells using Fluorescence Activated Cell Sorting (FACS). Key companion documents for use with this document: Visium HD Spatial Applications Protocol Planner (CG000698) Mar 11, 2024 · 10x Genomics Reagent Kit Requirements: See the Chromium GEM-X Single Cell User Guides for details; Third-party Items: See the Protocol Planner for a comprehensive list. 1 (Dual Index) with Feature Barcode technology for Cell Multiplexing Rev C31. Click to TOC. Cell loss during FACS is common. 1 (Dual Index) protocols with Feature Barcode technology for CRISPR Construct a 10x barcoded library using our reagent kits and a compatible Chromium instrument. Depending on your experimental goals, additional Library Construction Kits (PN-1000190 or PN-1000352) may be required. NO FILL. Chromium Next GEM Chip M Single Cell Kit* 80 rxns PN-1000349 | 16 rxns PN-1000371 (*Included with Chromium Next GEM Single Cell 3' HT Kit v3. 10x Genomics provides no warranty and hereby disclaims any and all warranties as to the use of any third-party products or protocols described herein. The i5 index read is used to capture the 10x barcode information. 1 (Dual Index) protocols with Feature Barcode technology for Cell Multiplexing and Single Cell 3' v3. 1 Single Index reagent kits, it is possible to upgrade to both Dual Indexing and Cell Multiplexing by following these instructions: Dec 7, 2022 · This protocol provides guidance for antibody-oligonucleotide conjugation and outlines cell surface protein labeling for use with Single Cell RNA sequencing protocols with Feature Barcoding technology. 1 Dual Index Gene Expression assay. 1 and 3' HT v3. If the incorrect index plate is used, library preparation will fail. PhiX Spike-In Recommendations: 1% for Singleplexed libraries, 5% for Multiplexed libraries, 10% for Multiplexed libraries on NovaSeq. After whole transcriptome probe panels are added to the sample, each probe pair hybridizes to its target gene and is then ligated. This protocol was demonstrated using primary cells (including peripheral blood mononuclear cells - PBMCs), dissociated tumor cells (DTCs), cell lines How do I pool 10x libraries for Illumina sequencing? Should I select Workflow A or Workflow B for the i5 index sequence? Can I sequence longer than 91-98bp on Read 2 for Single Cell 3' and 5' Gene Expression libraries? Can I decrease the sequencing length of Read 2 for Single Cell Gene Expression assays? Aug 26, 2022 · Chromium Single Cell 3' Reagent Kits User Guide (v3. Answer: Dual index plates are shared across certain 10x assays. These sequencing-ready dual index libraries can be pooled and sequenced based on recommendations outlined in Chromium Next GEM Single Cell 3’ user guide (Document CG000286). Dual-Indexed Sequencing Run: Fixed RNA Profiling libraries are dual-indexed. Profiling data analysis is also included in this document. Cell Surface Protein Labeling Protocol modifications (page 6) It is recommended to use the --barcode-mismatches argument set to the maximum 2 allowable mismatches to minimize the effect of sequencing errors on demultiplexing. com. Download the single index kit sample index reference files at Chromium Next GEM Single Cell 3ʹ - 10x Genomics • The total number of suspended nuclei used as input to 10x Genomics Single Cell protocols is determined by the nuclei recovery target. Chromium Single Cell Multiome ATAC + Gene Expression performs direct measurements of 3' gene expression and chromatin accessibility from the same cell. For example: There are product-specific pooling considerations. 1 (Dual Index) Rev A27. PhiX Spike-In Recommendations: 1%. 0 GEM Generation & Barcoding. Minimizing the presence of cellular aggregates, dead cells, noncellular nucleic acids, and potential biochemical inhibitors of reverse transcription is critical to obtaining high-quality data. 10x Genomics Support website Demonstrated protocols, 10x Genomics Support website Single-cell or single-nuclei suspension Prepare samples Chromium Single Cell Gene Expression Generate single-cell libraries NovaSeq 6000, NextSeq 2000, NextSeq 1000, or NextSeq 550 system Sequence Cell Ranger DRAGEN Single-Cell app Analyze data Loupe Browser DEMONSTRATED PROTOCOL CG00039 ev B 10xgenomics. 10xgenomics. The use of products described Aug 2, 2023 · Visium Spatial Gene Expression Libraries. Sep 9, 2022 · Depending on your experimental goals, additional Library Construction Kits (PN-1000190) may be required. 1 Chemistry Dual Index) with Feature Barcoding technology for CRISPR Screening and Cell Multiplexing (CG000389) Chromium Next GEM Single Cell 3' HT Reagent Kits v3. Chromium Single Cell V (D)J Amplification Kits. See the Supported Thermal Cyclers page for thermal cyclers we recommend using with this assay 951020460 951020362 Thermo Fisher Scientific MicroAmp 8-Tube Strip, 0. 1 protocol. 10x Genomics’ single cell RNA-seq (scRNA-seq) technology, the Chromium Single Cell 3’ solution, allows you to analyze transcriptomes on a cell-by-cell basis through the use of microfluidic partitioning to capture single cells and prepare barcoded, next-generation sequencing (NGS) cDNA libraries. It is done by partitioning thousands of cells into nanoliter-scale Gel Beads-in-emulsion (GEMs), where all generated cDNA molecules from an individual GEM share a common 10x Barcode. Please consult the corresponding User Guide, Sequencing Tech Notes, and the Sequencing Section on the 10x Support Page. It is critical to choose the User Guide that corresponds to your Reagent Kit version. If the incorrect dual index plate is used, library preparation will be unsuccessful. Demonstrated Protocol Thermal Cycler Protocol Download Guide for 10x Genomics Workflows CG000648 Rev B 10xgenomics. PhiX spike-in: None. Aug 9, 2021 · The Chromium Single Cell Gene Expression Solution provides a scalable microfluidic platform for gene expression profiling of 500-10,000 individual cells per sample. Aug 9, 2021 · 10x Genomics Support / Single Cell Gene Expression / Documentation / Library Construction / Chromium Next GEM Single Cell 3ʹ LT Reagent Kits (v3. Each member of the Chromium instrument family encapsulates each cell with a 10x barcoded Gel Bead in a single partition. Dual-Indexed Sequencing Run: Single Cell 5' v3 libraries are dual-indexed. If necessary, the collected cells may be concentrated by centrifugation at 350 rcf at 4oC and removing the supernatant. FFPE samples are NOT supported with Feature Barcode technology for Cell Surface Protein detection. The files below contain the mappings from the sample index names to the associated set of Index (i7) and Index 2 (i5 Each well of a dual index plate contains a mix of one unique i7 and one unique i5 sample index. R1 = 28bp, i7 = 8bp, R2 = 91bp. Additional Guidance. pdf Mar 15, 2023 · Dual index plates. 1. Map more cell types and pathways with 5,000 genes, use enhanced cell segmentation to guide your way, and get rapid insights with visualizable data at the end of a run. Contents 2 Chromium Fixed RNA Profiling Protocol Selection 3 Workflow Overviews and Document Resources 8 Protocol Steps & Timing 9 Kits, Reagents & Equipment – Sample Preparation Figure 1. Sep 12, 2023 · This protocol also optionally enables the detection of cell surface proteins in fixed single cell suspensions that are labeled with antibody-oligonucleotide conjugates prior to fixation. 3. Add 10 uL RT Enzyme Mix to the sorted nuclei in RT buffer, mix well but gently and load chip as per the Single Cell 3’ Reagents User Guide or Single Cell V(D)J 5’ Reagents User Guide. Mar 12, 2024 · 3’ v3. These libraries include 16 bp 10x GEM Barcodes (10x BC) encoded at the start of TruSeq Read 1 (Read 1T). Library Construction Kit C 16 rxns PN-1000694. For example: Suggested protocol for removing peaks below 200 bp: Add nuclease-free water to bring the library volume to 100 ul. 0 or higher is required in the Chromium Controller or the Chromium Single Cell Controller used for this Single Cell 3ʹ v3. 3 – cDNA QC & Quantification Protocol Step 7. Apr 3, 2024 · This document provides best practices for handling FFPE tissue samples and generating tissue slides. 1 Single Index and 3' v3. After removing chip from the sealed bag, use in ≤24 h. Chromium GEM-X Single Cell 5' Chip Kit 4 chips PN-1000698. Both the forward and reverse complement sequences are listed for the i5 sequence. The Tissue Preparation Guide provides guidance on: Selecting appropriate Visium Spatial slides specific to the Visium Spatial protocol being used. 1 User Guide with Feature Barcoding technology for Cell Surface Protein (CG000417) Answer: Each well of the Dual Index Plate TS Set A (PN-1000251) contains a mix of one unique i7 (Index 1) and one unique i5 (Index 2) sample index. Consult the applicable 10x Genomics Single Cell protocol to determine these relationships. 1_CellMultiplexing_RevC. 1, 48 rxns PN-1000348 | 8 rxns PN-1000370. Chromium Next GEM Single Cell 3 v3. Please refer to the appropriate User Guide and use only the specified index plate for your library type. Please click here for further guidance. Within each nanoliter-scale partition, cells undergo reverse transcription to generate cDNA, which shares a 10x Barcode with all Jun 23, 2023 · 10x Genomics Single Cell protocols require a suspension of viable single cells or nuclei as input. Dual Index Kit TS Set A Dual Index Plate TS Set A 1 3000511 # PN Dual Index Kit TS Set A, 96 rxns PN-1000251 (store at −20°C) for Fixed RNA Profiling Gene Expression library construction Dual Index Kit TT Set A Dual Index Plate TT Set A 1 3000431 # PN Dual Index Kit TT Set A, 96 rxns PN-1000215 (store at −20°C) Chromium Next GEM Single Cell 5' v2 (Dual Index) with Feature Barcode technology Rev C 4 TOC Table of Contents Introduction 6 Chromium Next GEM Single Cell 5 ʹ Reagent Kits v2 (Dual Index) 7 10x Genomics Accessories 12 Recommended Thermal Cyclers 12 Additional Kits, Reagents & Equipment 13 Protocol Steps & Timing 15 Consult Demonstrated Protocol Cell Surface Protein Labeling for Chromium Fixed RNA Profiling with Feature Barcode technology (CG000529) for guidance. • The optimal input nuclei concentration is 700-1,200 nuclei/µl. pdf Mar 12, 2024 · View and download. Repeat the SPRIselect cleanup in the "Post Sample Index PCR Double Sided Size Selection" step (this is step 3. We do not recommend sequencing these libraries with a single-index configuration. 2%. Each well of a dual index plate contains a mix of one unique i7 and one unique i5 sample index. Read key peer-reviewed research publications offering greater insights into biology, enabled by 10x Genomics products and technologies. Adapt fully constructed existing 10x Genomics libraries DEMONSTRATED PROTOCOL 10xGenomics. Do NOT use this protocol in conjunction with other Single Cell Gene Expression Flex Demonstrated Protocols. This Cell Preparation Handbook describes best Jun 13, 2022 · Dual index kit, TT, Set A is used with Chromium Single Cell Multiome Gene Expression to generate dual index libraries. Recommended Sequencing: Minimum 20,000 read pairs/cell*. For use with: Chromium Next GEM Single Cell Fixed RNA Sample Preparation Kit, 16 rxns PN-1000414 Jan 6, 2023 · Dual Index Kit TT Set A, 96 rxns PN-1000215; Dual Index Kit TN Set A, 96 rxns PN-1000250; Chromium Single Cell V(D)J Amplification Kits: Human 16 rxns TCR PN-1000252 / BCR PN-1000253; Mouse 16 rxns TCR PN-1000254 / BCR PN-1000255 Aug 2, 2023 · Dual-Indexed Sequencing Run: Single Cell Multiome ATAC libraries are dual-indexed. Please note, only the i7 index read is used for demultiplexing. The total number of cells required in the suspension used as input to 10x Genomics® Single Cell Solutions is determined by the cell recovery target defined by the user. See: Can single index and dual index libraries be pooled for Aug 9, 2021 · Chromium Single Cell 3' Reagent Kits User Guide (v3. 1 (Dual Index) with Feature Barcode technology for CRISPR Screening and Cell Multiplexing Rev B36. 0 or higher is required in the Chromium Controller or the Chromium Single Cell Controller used for this Single Cell 3’ v3. Best practices for handling tissue samples and Chromium Next GEM Single Cell 3' Reagent Kits v3. Chromium Single Cell Gene Expression provides single cell transcriptome 3' gene expression alongside the detection of surface protein expression or CRISPR edits in tens of thousands of cells. Chromium Single Cell Immune Profiling allows analysis of full-length, paired B-cell or T-cell receptor (BCR/TCR) sequences, surface protein expression, antigen specificity, and 5’ gene expression, all from a single cell. Caution: RNAse rich samples may need additional optimization. The upstream steps of the assay (GEM generation, reverse transcription, cDNA amplification) are identical between the Single Index and Dual Index assays. Publications. Sep 12, 2023 · Chromium Fixed RNA Profiling for Multiplexed Samples provides an efficient and cost-effective way to further increase experiment size and cell number by enabling up to 4 or 16 samples to be run within a single GEM reaction. Optimize the protocol steps accordingly. Fixation at the point of sample collection preserves fragile biology and greatly streamlines workflows to provide critical insights Sep 22, 2023 · This is critical for downstream library preparation and generation of high quality sequencing data using the Visium Spatial Gene Expression protocols. 4% to 8. Products: Visium for FFPE, Visium CytAssist for FFPE, Visium HD, Fixed RNA Profiling Gene Expression. DO NOT substitute any of the listed The Chromium Fixed RNA Profiling is designed to target 500-10,000 cells per Probe Barcode with a per sample undetected multiplet rate of 0. Results: We did not encounter any technical issues with sequencing 1:1 pools of single and dual index libraries using either single index or dual index run configurations. Dec 7, 2022 · This protocol provides guidance for antibody-oligonucleotide conjugation and outlines cell surface protein labeling for use with Single Cell RNA sequencing protocols with Feature Barcoding technology. 1 Chemistry Dual Index) with Feature Barcoding technology for Cell Multiplexing User Guide , CG000388 CG000388_ChromiumNextGEMSingleCell3-v3. Libraries are generated from the barcoded probes Click to TOC Chromium Next GEM Single Cell 3 v3. Indexing design Dual indexing design. Scale up with a high-throughput solution, or focus on key genes with targeted gene expression panels. Thus, as long as you choose a unique well from the 96-well plate for product. a. 1_Isothermal_Inc_60min GEM Incubation ATAC_GEM_Inc Aug 26, 2022 · Chromium Single Cell 3' Reagent Kits User Guide (v3. Download the dual index kit sample index reference files at these links: 3' Single Cell Gene Expression; 5' Single Cell Gene Expression; Fixed RNA Profiling; Single indexing design. View Publications. Please use only the dual index plate recommended in the User Guide for each assay and library type. As shown in Figure 1, the Fixed RNA Profiling Gene Expression libraries comprise standard Illumina paired-end constructs, which begin with P5 and end with P7. com Table 3: Chromium Next GEM Single Cell ATAC v1. In consideration that 10x has discontinued the single index assays, please note that BioLegend now offers an alternative dual index TSA protocol and TotalSeq-B or -C for use with 10x Genomics dual indexing kits that users Apr 27, 2022 · View and download. A list of 10x Genomics validated sequencers and expected data throughput can be found in the Sequencing Metrics for Illumina Sequencers Technical Note . RNA molecules are transcribed and processed Oct 9, 2023 · TotalSeq A libraries were generated according to manufacturer’s instructions and BioLegend’s TSA single index protocol. The 10x Genomics Chromium Fixed RNA Profiling Reagent Kits are not listed in this document. Multiplex-compatible Chromium Next GEM Single Cell Fixed RNA Human Transcriptome Probe kits (PN-1000420/1000456) contain 4 This protocol provides guidance for: • Labeling cells/nuclei with CellPlex reagents (see Cell Multiplexing Oligo Labeling) for use with Single Cell 3' v3. 1 (discontinued) Protocol Step(s) Protocol File Name Variable Cycle Number/Incubation Time Isothermal Incubation ATACv1. 8X (left side ratio) and Protocol steps correspond to the Chromium Next GEM Single Cell Multiome ATAC + GEX User Guide (CG000338). What happens if I forget to update my thermal cycler program for Pre-Amplification and I perform Barcode Oligo Capture protocols? Why is the annealing temperature during Pre-Amplification updated from 67°C to 63°C across all Fixed RNA Profiling protocols? Why isn’t TotalSeq™️-B compatible with Fixed RNA Profiling for Multiplexed Samples? Answer: The document LIT000154_Chromium-Connect-Kit-Selection-Guide outlines the Supported workflows for: Continuous or modular cells-to-libraries Single Cell 3’ and 5’ gene expression workflows from cell or nuclei suspensions. 1with Feature Barcode technology for CRISPR Screening (CG000418) Dec 7, 2022 · Please contact 10x Genomics support at [email protected] for TotalSeq-C antibody recommendations. com Cell Multiplexing Oligo Labeling for Single Cell RNA Sequencing Protocols with Feature Barcode technology Introduction The 10x Genomics 3’ CellPlex Kit provides a species agnostic sample multiplexing solution through the use of a set of 12 Feature Barcode oligonucleotides Step 1 GEM Generation & Barcoding. Click to TOCChromium Next GEM Single Cell 3 v3. 1_CellSurfaceProtein_RevD. Dilute cells using only compatible cell media/buffer (minimize carryover of media that can destabilize GEMs & impact GEM-RT reaction). We do not recommend sequencing 10x Single Cell 5’ v3 libraries with a single-index configuration. com CG000391 • Rev A Demonstrated Protocol Cell Multiplexing ligo Labeling for Single Cell RNA Sequencing Protocols • Rev A Overview The 10x Genomics 3’ CellPlex Kit provides a species agnostic sample multiplexing solution through the use of a set of 12 Feature Barcode oligonucleotides each conjugated to Proceed immediately with the 10x Genomics Single Cell Protocol and minimize the time between nuclei preparation/sorting and chip loading. 2 ml MicroAmp 8-Cap Strip, clear N8010580 4323032 Kits & Reagents Thermo Fisher Scientific Nuclease-free Water AM9937 10X Genomics Single Cell 3' (v3) RNA sequencing is a microdroplet-based method that permits the effective capture and sequencing of the mRNA and pre-mRNA molecules from single nuclei [1]. Chromium Single Cell Gene Expression provides single cell transcriptome 3' gene expression and multiomic capabilities to profile tens of thousands of cells. Click to TOCChromium Next GEM Single Cell 3 ʹ LT Reagent Kits v3. 1 Dual Index User Guide and uses a double-sided SPRIselect ratio of 0. To calculate this, first estimate the % of capture area covered by the tissue section based upon the H&E brightfield image. 1 (Dual Index) with Feature Barcode technology for CRISPR Screening • Rev D 6 Introduction Chromium Next GEM Single Cell 3 ’ Reagent Kits v3. The cell's a big place—now go eXplore it with the precision of high-plex single cell spatial imaging. For use with: Chromium Next GEM Single Cell 5' HT Kit v2, 48 rxns PN-1000356 / 8 rxns PN-1000374. Each sample index well in the 96-well plate is a unique set, and therefore the sample indexes can be used in any combination. Dual Index Kit TT Set A (PN-1000215 / PN-3000431): Dual Index Kit NT Set A (PN-1000242 / PN View Publications. The required changes are described below. 6 in the Single Cell 3' v3. 26. R1 = 28bp, i7 = 10bp, i5 = 10bp, R2 = 90bp. FOR USE WITH: Chromium Next GEM Single Cell 5' HT Reagent Kits v2 (Dual Index) with Feature Barcode technology for Cell Surface Protein & Immune Receptor Mapping User Guide (Document CG000424) This protocol provides guidance for: • Labeling cells/nuclei with CellPlex reagents (see Cell Multiplexing Oligo Labeling) for use with Single Cell 3' v3. For use with: Chromium GEM-X Single Cell 5' Kit v3 16 rxns PN-1000699 | 4 rxns PN-1000695. The 3’ CellPlex Kit is only supported for use with the Chromium Single Cell 3’ v. e. Then use the following formula to calculate the recommended sequencing depth: Jun 9, 2023 · This Technical Note highlights the consistency and reproducibility in generation of dual index Chromium Single Cell 3' Gene Expression libraries using the Chromium Connect automated workflow. It uses probes targeting the human whole transcriptome and oligonucleotide conjugated antibodies for detecting immune cell protein markers. The optimal cell loading concentration for this protocol is ~100-600 cells/ l. Use the filters below to find your specific user guide. The use of products described herein is subject to certain restrictions as set forth in the applicable terms and conditions of sale accompanying the purchase of such Single index run configuration. Sep 20, 2023 · Chromium Fixed RNA Kit, Human Transcriptome, 4 rxns x 4 BC PN-1000475 | 4 rxns x 16 BC PN-1000476; Chromium Fixed RNA Kit, Mouse Transcriptome, 4 rxns x 4 BC PN-1000496 | 4 rxns x 16 BC PN-1000497; Chromium Next GEM Chip Q Single Cell Kit, 48 rxns PN-1000418 / 16 rxns PN-1000422; Dual Index Kit TS Set A 96 rxns PN-1000251 Step 1 GEM Generation & Barcoding. 1 (Dual Index) protocols with Feature Barcode technology for CRISPR Answer : The Dual Index Kit Plate TS Set A, 96 rxns (PN-1000251) is the Sample Index Plate required for use with the Chromium Fixed RNA Profiling (i. Read Read 1 i7 Index i5 Index Read 2 Purpose 10x Barcode & UMI Sample Index N/A Insert Length** 26 8 0 91 Read Read 1 i7 Index i5 Index Read 2 Purpose 10x Barcode & UMI Sample Index N/A Insert Overview. The files below contain the mappings from the sample index names to the associated set of Index (i7) and Index 2 (i5) oligonucleotides. , Single Cell Gene Expression Flex) workflow. 1 Dual Index assays have different library preparation protocols, to incorporate single or dual indexes. When loading the chip, raising and depressing the pipette plunger should each take ~5 sec. in situ. For use with: Chromium Next GEM Single Cell Fixed RNA Sample Preparation Kit, 16 rxns PN-1000414 Chromium Fixed RNA Kit, Mouse Transcriptome, 4 rxns x 1 BC: 1000495: Chromium Next GEM Chip Q Single Cell Kit, 16 rxns: 1000422: Chromium Next GEM Chip Q Single Cell Kit, 48 rxns: 1000418: Fixed RNA Feature Barcode Multiplexing Kit, 64 rxn* 1000628: Dual Index Kit TS Set A, 96 rxns : 1000251: Dual Index Kit TN Set A, 96 rxns: 1000250 Aug 2, 2023 · Single Cell 5' v3 Gene Expression Libraries. 1; 16 rxn can also be ordered separately) Dual Index Kit TT Set A 96 rxns PN-1000215. Protocol Step 5. TruSeq Sep 13, 2023 · Recommended Sequencing: Minimum 10,000 read pairs/cell*. 1 (Dual Index) 10x Genomics Accessories Recommended Thermal Cyclers Additional Kits, Reagents & Equipment Protocol Steps & Timing Stepwise Objectives Dual Index Kit TT Set A, 96 rxns PN-1000215 (store at −20°C) Click to TOCChromium Next GEM Single Cell 5' v2 with Feature Barcode technology_CRISPR Screening Rev A13. 1 Dual Index Cell Multiplexing libraries are made using the following sample index plate: PN-1000243/PN-3000482: Dual Index Kit NN, Set A: CSV | JSON * Only Feature Barcode technology for Cell Surface Proteins is supported with the Single Cell 3' v4 and LT v3. Library construction or 5’ V (D)J amplification workflows (TCR/BCR profiling) from purified cDNA. Dual index plates are used with Chromium Single Cell 5' v3, 5' v2, and 5' HT v2 Dual Index libraries. What is Single Cell Gene Expression Flex? Single Cell Gene Expression Flex enables single cell gene expression on precious samples that were previously inaccessible due to logistical challenges in sample handling. Aug 22, 2023 · View and download. May 22, 2024 · This protocol is only compatible with FFPE samples and Chromium Fixed RNA Profiling reagent kits. For use with: Chromium Next GEM Single Cell 3' HT Kit v3. If you have already purchased Single Cell 3’ v3. Jan 6, 2023 · Chromium Single Cell 5' Reagent Kits User Guide (v2 Chemistry Dual Index) with Feature Barcoding technology for Cell Surface Protein and Immune Receptor Mapping User Guide , CG000330 CG000330_ChromiumNextGEMSingleCell5-v2_CellSurfaceProtein_UserGuide_RevF. multiplet rate of ~3. 1 Chemistry Dual Index) with Feature Barcoding technology for Cell Surface Protein User Guide , CG000317 CG000317_ChromiumNextGEMSingleCell3-v3. . Refer to the applicable 10x Genomics Single Cell Protocol to determine this relationship. The items in the table below have been validated by 10x Genomics and are highly recommended for the Single Cell 5' protocols. Protocols for performing deparaffinization, staining (H&E and IF), and decrosslinking of tissue sections. Oct 13, 2022 · View and download. View pricing. 1(Dual Index) User Guide Rev E. Dual index run configuration. 1 (Dual Index) User Guide with Feature Barcoding technology for Cell Surface Protein (CG000317) Chromium Next GEM Single Cell 3' HT Reagent Kits v3. Each well of the Dual Index Plate TS Set A (PN-1000251) contains a mix of one unique i7 (Index 1) and one unique i5 (Index 2) sample index. Run Cell Ranger mkfastq to demultiplex using only the i7 index with 2 allowed mismatches: cellranger mkfastq --id=FASTQ_output_directory \. The data can be analyzed and visualized using Cell Ranger and Loupe Cell Browser for assessing single cell gene expression. Answer: Pooling 10x libraries for sequencing will depend on the library type, target cell number, tissue covered spots (Visium), and desired read depth. The documents also presents a comparison of representative data derived from dual versus single index Chromium Single Cell 3’ libraries generated using and viability measured before proceeding to the 10x Genomics Single Cell protocols. Thermal Cycler Protocol Download Guide for 10x Genomics Workflows All thermal cycler protocols were manually programmed according to their respective 10x Genomics product-specific User Guides, resulting in instrument-generated protocol files. The recommended number of cells for this modified protocol is 0. To enable cell hashing, the Demonstrated Protocol for Cell Surface Protein Labeling for Single Cell RNA Sequencing Protocols must be modified. 10xgenomics. --use-bases-mask=Y28n*,I10,N10,Y90n* \. The items in the table below have been tested by 10x Genomics and are required for the Chromium Connect Automated Single Cell 5' protocol. The Single Cell Gene Expression Flex Fixed RNA Profiling (FRP) workflow measures RNA levels in fixed samples using probes targeting the whole transcriptome. 7 – Post Library Construction QC (Gene Expression Library) All traces are representative. Firmware Version 4. Cell Preparation for Single Cell Protocols Introduction 10x Genomics Single Cell protocols require a suspension of viable single cells or nuclei as input. Minimum Sequencing Depth: We recommend a minimum of 50k read pairs per spot covered with tissue. 2b. 5,000 ways to explore tissue biology. 2-2 x 10^ 6 cells per sample. A: You have three options to create compatible 10x Genomics libraries with the Singular Genomics G4 Sequencing Platform: Use Singular Genomics (SG) indexed or non-indexed t-primers during library preparation instead of the primers provided by 10x Genomics during the sample index PCR step. Explore cellular heterogeneity, novel targets, and biomarkers with combined gene expression, cell surface protein expression, or CRISPR edits in each cell. 3 – Post Library Construction QC (ATAC Library) Protocol Step 6. The solution can target up to 40,000 cells when multiplexing 4 samples and up to 128,000 cells when multiplexing 16 samples. FOR USE WITH: Chromium Next GEM Single Cell 5' Kit v2, 16 rxns PN-1000263 cell by cell. 1 Dual Index) with Feature Barcode technology for Cell Surface Protein Dual Index Plate TS Set A - contains Illumina TruSeq Read 1 and TruSeq Small RNA Read 2S sequences. See performance. Please see: Can I use RNase-rich tissue samples for single-cell gene expression or Multiome assays? product. nsjrabcyglqlcdiegezw