Single-cell RNA sequencing
Full-length isoform information for your single-cell transcriptome studies
Reveal hidden isoform diversity at single-cell resolution
Understanding cell heterogeneity at the isoform level is critical for basic and disease research. Short reads can only capture gene-level information, while other long-read technologies lack the accuracy for accurate UMI/barcode identification.
Only HiFi reads can provide accurate detection of isoforms in your single-cell study. No complicated algorithms for barcode correction or orthogonal sequencing data are required. With the MAS-Seq method for single-cell isoform sequencing, you can:
- Achieve 16-fold throughput increase compared to a regular single-cell Iso-Seq library
- Move beyond gene counting to get full-length isoform information
- Characterize the full diversity of transcript isoforms at the single-cell level
- Reveal cell type-specific spliced isoforms and expressed variants
Long reads reveal isoform differences across cell types. Reads cover the entire isoform with single-cell tag information.
Video
Single-cell RNA sequencing with MAS-Seq
How do the MAS-Seq kit + informatics work to make single cell isoform sequencing cost effective and easy to analyze? Come learn how to make MAS-Seq libraries from 10x Genomics single-cell cDNA, sequence on PacBio systems, and analyze with a click-button SMRT Link software solution.
HIFI SEQUENCING ADVANTAGE IN SINGLE-CELL RNA SEQUENCING
|
Long-read sequencing performance |
Short reads | Other long reads | PacBio |
|---|---|---|---|
| Single technology | ✓ | X | ✓ |
| Isoform information | X | ✓ | ✓ |
| Variant information | X | ✓ | ✓ |
| Official software | ✓ | X | ✓ |
Common questions about single-cell RNA sequencing and the MAS-Seq kit
Single-cell RNA sequencing (scRNA-seq) emerged to characterize gene expression differences between individual cells derived from a complex tissue, allowing a higher-resolution look at the transcriptome. A typical single-cell library takes RNA as input and outputs cDNA molecules that have been tagged with single-cell barcodes and unique molecular indices (UMIs) so that transcripts can be traced back to their “single cell” origins. This is in contrast with, bulk RNA-Seq, where there are no single-cell barcodes to attribute transcripts back to cells.
Single-cell RNA sequencing using PacBio HiFi reads offers full-length isoform information at the single-cell level. Short-read sequencing only sequences a short fragment of one end of the transcript, providing only gene-level information. In contrast, PacBio HiFi reads can cover the entire transcript, revealing isoform diversity at the single-cell level. Isoforms — not genes — are often the driver of disease and biology.
The MAS-Seq method was developed by Al’Khafaji et al. as a concatenation method for increasing throughput by joining cDNA molecules generated from single-cell platforms into longer concatenated fragments. HiFi reads generated from sequencing the concatenated molecules can then be bioinformatically broken up to retrieve the original molecules. The result is higher throughput and reduced sequencing needs. The authors showed that, applying the MAS-Seq method to 10x single-cell libraries, unique isoforms associated with specific cell types relevant to cancer can be distinguished which are otherwise undetectable using short-read single-cell RNA-Seq.
Yes! With MAS-Seq, you can obtain full-length isoform information for your single-cell libraries, whereas with other sequencing methods, such as short reads, you can only obtain gene-level information. Multiple publications, using MAS-Seq (Al’Khafaji et al.) or regular HiFi sequencing (Palmer et al., Joglekar et al.) have demonstrated the use of PacBio for identifying cell-type isoform specificity.
No. With the MAS-Seq kit, PacBio HiFi sequencing offers sufficient coverage for a typical single-cell experiment with 1 SMRT Cell on the PacBio long-read systems. No orthogonal sequencing technology is required.
The MAS-Seq kit is compatible with cDNA generated using the 10x Chromium Next GEM Single Cell 3’ kit (v3.1).
Hear how MAS-Seq is transforming the single-cell RNA sequencing world
Video
Multiplexed arrays sequencing
Dr. Aziz Al’Khafaji shares a new method, developed at the Broad Institute, called Multiplexed Arrays sequencing (MAS-Seq), which maximizes the sequencing potential of full-length mRNA isoforms on the PacBio platform.
Single-cell transcriptomics for cancer research
A more complete transcriptome for cancer research – now at single-cell level. With MAS-Seq, upgrade your single-cell cancer research to:
- Discover isoforms as potential biomarkers and help enable therapeutic targets
- Identify fusion isoforms
- Detect expression mutations
Workflow
Empower your single-cell transcriptome study with MAS-Seq
- 16-fold throughput increase using the MAS-seq method
- From cDNA to sequencing-ready library in 2 days
- 1 SMRT Cell 8M sufficient to characterize a typical single-cell (3,000–10,000 cell) library
- SMRT Link workflow to output gene- and isoform- matrix, compatible with popular tertiary software