The 8 bp index is a component of the MULTI-seq barcode. There are two types of MULTIseq barcode oligos, one for 3’ capture and one for 5’ capture. Each barcode oligo contains an 8 bp variable region. The 8 bp sequence will be provided when ordered. The MULTIseq Additive Primer, NGS (Next Gen Sequencing) prep oligos and all oligos that will be needed to complete the scRNAseq process can also be custom ordered by email ([email protected]) or through SigmaAldrich.com/nextgenoligos.
LMO001
MULTI-seq Lipid-Modified Oligos
for Single Cell and Single Nucleus Multiplexing
Synonym(s):
LMO, Lipid modified oligos
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About This Item
UNSPSC Code:
12352211
NACRES:
NA.51
Pricing and availability is not currently available.
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Quality Level
form
liquid
usage
sufficient for 100 sample(s) (100RXN kit is adequate for labelling 100 samples when using 1 μL of each LMO per sample)
sufficient for 30 sample(s) (30RXN kit is adequate for labelling 30 samples when using 1 μL of each LMO per sample)
concentration
50 μM (each)
technique(s)
multiplexing: suitable (Single Cell and Single Nucleus)
shipped in
dry ice
storage temp.
−20°C
General description
MULTI-seq, multiplexing using lipid-tagged indices (lipid-modified oligonucleotides), is a novel tool that facilitates multiplex single-cell and single-nucleus RNA sequencing. This technology provides sample barcoding that is compatible with any cell type or nucleus containing an accessible plasma membrane and allows for streamlined, pooled single-cell sample processing. Learn more about the experiments and data by reviewing our technical article – (MULTI-seq Sample Multiplexing for Single Cell Analysis and Sequencing)
Features and Benefits
- Efficient. MULTI-seq increases the throughput and decreases the reagent cost of droplet-based single-cell RNA sequencing.
- Flexible. Compatible with any droplet generation platform, the indexing system enables users to run and analyze up to 96 barcoded samples simultaneously in an 8-lane droplet device.
- Clearer Results. The indices provide data quality advantages over non-indexed analysis methods in the form of doublet identification and retention of data from cells with low RNA content.
Components
The MULTI-seq Lipid Modified Oligos kit is comprised of a lignoceric amide-modified anchor DNA oligo solution and a palmitic amide-modified co-anchor DNA oligo solution. Together, these lipid-modified oligos embed into cell or nuclei membranes and provide a landing pad for DNA barcodes with a complementary 5’ sequence. The MULTI-seq Lipid Modified Oligos kit is intended for upstream sample preparation only, before pooling and single cell analysis.
Reagents Provided
Reagents Provided
- LMO001A Lignoceric Anchor with DNA Oligo
- LMO001B Palmitic Co-anchor with DNA Oligo
Other Notes
- This product is for R&D use only. Not for drug, household, or other uses.
- Unique barcode oligos are not included and must be purchased separately.
- Barcode 3’ oligo design with poly-A tail for mRNA enrichment: 5′-CCTTGGCACCCGAGAATTCCA-8-base index-A30-3′
- Barcode oligo design for 5’ cDNA library synthesis: 5’-CCTTGGCACCCGAGAATTCCA-8-base indexCCCATATAAGAAA-3’
- In addition, 96 x 5′-end and 96 x 3′-end unique barcode oligos are available to purchase separately. Details on our custom barcode product, Next-Gen Sequencing Oligos (NGSO), can be found at SigmaAldrich.com/nextgenoligos. For minimal cross-contamination of barcode primers, order NGSO-Silver or preferably, NGSO-Gold quality. Please review the specifications available and then submit a quote request for the MULTI-seq barcodes to [email protected]. Sequences are not provided prior to purchase but will be included on product documentation at the time of delivery.
related product
Product No.
Description
Pricing
Storage Class Code
12 - Non Combustible Liquids
Flash Point(F)
Not applicable
Flash Point(C)
Not applicable
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Daniel V Brown et al.
Genomics, 116(2), 110793-110793 (2024-01-15)
Single-cell RNA sequencing (scRNA-Seq) has emerged as a powerful tool for understanding cellular heterogeneity and function. However the choice of sample multiplexing reagents can impact data quality and experimental outcomes. In this study, we compared various multiplexing reagents, including MULTI-Seq
Christopher S McGinnis et al.
Nature methods, 16(7), 619-626 (2019-06-19)
Sample multiplexing facilitates scRNA-seq by reducing costs and identifying artifacts such as cell doublets. However, universal and scalable sample barcoding strategies have not been described. We therefore developed MULTI-seq: multiplexing using lipid-tagged indices for single-cell and single-nucleus RNA sequencing. MULTI-seq
Henrik Gezelius et al.
NAR genomics and bioinformatics, 6(1), lqae001-lqae001 (2024-01-30)
Functional precision medicine (FPM) aims to optimize patient-specific drug selection based on the unique characteristics of their cancer cells. Recent advancements in high throughput ex vivo drug profiling have accelerated interest in FPM. Here, we present a proof-of-concept study for
Articles
Use of MULTI-seq lipid-modified oligos, protocol, and troubleshooting guide for PCR Assays and Sequencing applications.
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How do I order the 8-bp index sample barcodes?
1 answer-
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What is needed to add MULTIseq to my scRNAseq workflow?
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Each sample that will be multiplexed will require the MULTIseq anchor oligos and a unique MULTIseq barcode. The common Anchor/Co-anchor Lipid Modified Oligos (LMOs) are components of the MULTIseq LMO001 kit. Each sample will also need a unique MULTIseq barcode oligo that can be custom ordered by sending an email to [email protected]. There are two types of MULTIseq barcode oligos, one for 3’ capture and one for 5’ capture. Each barcode oligo contains an 8bp variable region. The 8bp sequence will be provided when ordered. The MULTIseq Additive Primer, NGS (Next Gen Sequencing) prep oligos and all oligos that will be needed to complete the scRNAseq process can also be custom ordered by email or through SigmaAldrich.com/nextgenoligos.
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How will adding MULTIseq to my scRNAseq experiments impact my workflow?
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MULTIseq oligos are simple to use and have minimal impact on the overall scRNAseq process. During the harvest/cell prep, individual samples are labeled with the MULTIseq Anchor/Co-anchor LMOs and a unique MULTIseq barcode. The labeling process will add 30 min to an hour to the overall process. During the SPRI bead cDNA clean-up step, the supernatant is retained and will contain the MULTIseq barcodes. The supernatant will be subjected to its own separate clean-up and library prep.
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What cell types are MULTIseq compatible with?
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The MULTIseq Anchor/Co-anchor Lipid Modified Oligos can embed into any lipid bilayer. MULTIseq oligos are cell-type agnostic and can be implanted into any cell membrane. MULTIseq oligos are also compatible with purified nuclei with no changes to the labeling procedure.
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I will be submitting my MULTIseq labeled cells to our core facility for single cell processing and deep sequencing. How will MULTIseq labeled samples impact the workflow?
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First, if you wish to “superload” the lane(s), you will need to specify to have 40K cells loaded per lane instead of the normal 15K. Second, during the 0.6X SPRI Bead cDNA clean-up in Step 2.3, the supernatant MUST be saved. The supernatant is normally discarded but will contain the MULTIseq Barcodes. The barcodes must be cleaned up and library prepped separate from the cDNA. Once the MULTIseq barcode library has been generated, it can be sequenced together with the cDNA library at 2-3K MULTIseq barcode reads per cell.
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How can I ensure a high cell yield during labeling?
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Start with as healthy cells as possible. Process cells efficiently and store them on ice when feasible. Carefully pipette cells to ensure isolated cells/nuclei are free from clumps and aggregates during labeling/wash steps. Aspirate slowly with a pipette retaining 50-100 µl of supernatant to ensure the pellet is not disrupted. Free floating DNA from dead cells can cause stickiness that could lead to straining/filtration cell loss.
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How will the raw deep sequencing files (FASTQ) be processed for the gene expression library vs the MULTIseq barcode library?
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The FASTQ files generated from the gene expression library will be processed through the 10X Cell Ranger pipeline, as usual. The MULTIseq barcode library FASTQ files will be processed deMULTIplex2 (https://github.com/Gartner-Lab/deMULTIplex2).
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How many samples can we multiplex together using MULTIseq?
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MULTIseq can multiplex up to 96 separate samples together and are dependent on the unique MULTIseq barcodes that are used. The MULTIseq LMOs are available in 30 and 100 reaction sizes. The MULTIseq barcodes are available with 96 unique sequences.
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What 10X kits is MULTIseq compatible with?
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MULTI-seq is compatible with all 10X kits that profile gene expression. MULTI-seq is not compatible with kits that do not capture the mRNA.
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What scRNAseq platforms is MULTIseq compatible with?
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The MULTIseq LMOs are compatible with 10X Genomics, BD Rhapsody, Drop-Seq, iCell8, etc.
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