10x Genomics single-cell transcriptomics human cell lines

This post details the release of 10x Genomics single cell cDNA sequencing of two cell lines: 293T and Jurkat. For each cell line two biological replicates were prepared, with each replicate being sequences with two PromethION Flow Cells (for a total of 8 Flow Cells). The 293T data were prepared using the Chromium GEM-X Universal 3’ Gene Expression v4 kit, while Jurkat data were prepared using the Chromium GEM-X Universal 5’ Gene Expression v3 kit.

Dataset

The dataset is available for anonymous download, without login credentials, from a public Amazon Web Services S3 bucket. The bucket is part of the Open Data on AWS project enabling sharing and analysis of a wide range of data. The data is located in the bucket at:

s3://ont-open-data/sc_gemx_2025.02

See the tutorials page for information on downloading the dataset.

You can also browse and download the files in your web browser courtesy of 42basepairs.

Sample

Preparation

Sample preparation was performed according to protocols published on the main Oxford Nanopore Technologies’ website.

Further information such as sample storage suggestions can be found at https://nanoporetech.com/documentation/prepare.

Sequencing

Sequence data was generated using the PromethION sequencer, with the following configuration:

using the the following 10x Genomics chemistries to prepare initial cDNA libraries:

Basecalling was performed off device using dorado (v0.9.1).

Analyses

To perform cell demulltiplexing and UMI deduplication we used our wf-single-cell (v2.4.1) Nextflow workflow. The reference used in the analyses and provided to the workflows --ref_genome_dir parameter is available to download from 10x Genomics.

The analysis results are located in the S3 bucket under the prefix:

s3://ont-open-data/sc_gemx_2025.02/analysis

Related Materials

• Obtaining full-length isoforms from single cells with nanopore sequencing whitepaper