Single-cell transcriptomics with Oxford Nanopore
During this two-part series on single-cell transcriptomics, members of the Oxford Nanopore team shared how you could use nanopore sequencing for single-cell analysis. Starting with sample and library prep, then moving onto data analysis.
Getting started
Carly Tyer provided an overview of sample and library preparation for single-cell transcriptome sequencing with nanopore technology. Carly focused on the benefits of single-cell analysis using the technology, including the ability to generate full-length cDNA sequences for exploring splice isoforms, obtaining quantitative expression data for comparing transcriptome variation at the single-cell level, and the identification of novel transcripts. Single-cell nanopore sequencing has been used in applications ranging from characterising aberrant splicing in cancer, to immune system profiling of full-length antibody sequences.
Viewers will learn:
- Considerations for wet lab and library prep for 10x Genomics single-cell transcriptome sequencing with Oxford Nanopore
- What to expect from a single-cell library run on PromethION
- How to identify and quantify full-length transcripts from long nanopore reads
- How to explore splice variants and isoforms with single-cell data
Meet the speaker
Carly Tyer is an Applications Scientist on the Genomic Applications team at Oxford Nanopore Technologies, where she specialises in wet lab sequencing protocol development, including chromatin conformation capture using Pore-C, genomic characterisation in cancer and immunology, and single-cell sequencing of full-length human transcriptomes.
Data analysis
John Beaulaurier guided users through the process of demultiplexing and conducting preliminary analyses on sequencing data produced from the Oxford Nanopore Technologies “Single-cell transcriptomics with cDNA prepared using 10X Genomics Protocol”. These steps are performed using the Sockeye open-source bioinformatic pipeline, which can now be executed via the wf-single-cell workflow in EPI2ME Labs. This workflow is designed to recover cell barcode and UMI sequences from the nanopore reads, as well as generate some basic outputs describing the composition of the single-cell sequencing data.
Viewers will learn:
- What kind of sequencing data to expect from the single-cell sequencing protocol
- How to install, configure, and run the EPI2ME Labs wf-single-cell workflow
- What primary and intermediate output files are generated by various steps during the pipeline run
- FAQs and considerations when examining output files
Meet the speaker
John Beaulaurier is a Genomic Applications Bioinformatics Manager at Oxford Nanopore, where he leads a team of bioinformatics scientists focused on collaborative projects designed to highlight the unique capabilities of nanopore sequencing technology.