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Comparative Study
. 2024 Nov 11;25(1):1069.
doi: 10.1186/s12864-024-10976-x.

Comparative transcriptomic analyses of thymocytes using 10x Genomics and Parse scRNA-seq technologies

Igor Filippov #  1   2 , Chinna Susan Philip #  3 , Leif Schauser  4 , Pärt Peterson  5
Affiliations
Comparative Study

Comparative transcriptomic analyses of thymocytes using 10x Genomics and Parse scRNA-seq technologies

Igor Filippov et al. BMC Genomics. .

Abstract

Background: Single-cell RNA sequencing experiments commonly use 10x Genomics (10x) kits due to their high-throughput capacity and standardized protocols. Recently, Parse Biosciences (Parse) introduced an alternative technology that uses multiple in-situ barcoding rounds within standard 96-well plates. Parse enables the analysis of more cells from multiple samples in a single run without the need for additional reagents or specialized microfluidics equipment. To evaluate the performance of both platforms, we conducted a benchmark study using biological and technical replicates of mouse thymus as a complex immune tissue.

Results: We found that Parse detected nearly twice the number of genes compared to 10x, with each platform detecting a distinct set of genes. The comparison of multiplexed samples generated from 10x and Parse techniques showed 10x data to have lower technical variability and more precise annotation of biological states in the thymus compared to Parse.

Conclusion: Our results provide a comprehensive comparison of the suitability of both single-cell platforms for immunological studies.

Keywords: 10x; Parse; Thymus; Transcriptomics; scRNA-seq.

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Conflict of interest statement

Declarations Ethics approval and consent to participate This study was conducted in accordance to the permission from the Ministry of Regional Affairs and Agriculture (Estonia) and approved by the Animal Experiments Ethics Committee at the Ministry (Protocol No. 224). All methods were performed in accordance with relevant guidelines and regulations. This study was carried out in compliance with the ARRIVE guidelines (Animal Research: Reporting of In Vivo Experiments). Consent for publication Not applicable. Competing interests The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
The experiment design. Thymi from two mice were divided into two technical replicates each. The resulting four samples were further divided between 10x and Parse workflows
Fig. 2
Fig. 2
Cell recovery and gene detection. A Number of cells recovered from each replicate, (B) UMI counts/cell for each sample, (C) Genes detected/cell in each sample, (D) Percentage of expression mapped to mitochondrial genes, (E) Percentage of expression mapped to ribosomal genes, (F) Percentage of expression mapped to long non-coding RNA genes, (G) Top expressed genes in 10x library, (H) Top expressed genes in Parse library, (I) Overlap of all genes detected in both libraries, (J) Overlap of top 1000 expressed genes in both libraries
Fig. 3
Fig. 3
Batch effect exploration. (A) UMAP representation of 10x data without batch effect correction, (B) UMAP representation of Parse data without batch effect correction, (C) UMAP representation of Parse data with batch effect correction, (D) Overlap in HVGs between samples, (E) KEGG pathways enriched in HVGs for each sample
Fig. 4
Fig. 4
Cell type annotation. UMAP (A) and marker genes (B) of 10 × data, UMAP (C) and marker genes (D) of Parse data, Proportion of cells in 10x (blue) and Parse (purple) in replicates (E) A_1, (F) A_2, (G) B_1, and (H) B_2
Fig. 5
Fig. 5
Workflow and time frame required for each kit. Key differences between the steps and time spent on 10x and Parse protocols have been highlighted
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