MIRALCS – microwell full-length mRNA amplification and library construction from a single-cell

Viral infection causes multiple forms of human cancer, and HPV infection is the primary factor in cervical carcinomas. Recent single-cell RNA-seq studies highlight the tumor heterogeneity present in most cancers, but virally induced tumors have not been studied. HeLa is a well characterized HPV+ cervical cancer cell line.

Researchers from BGI-Shenzhen have developed a new high throughput platform to prepare single-cell RNA on a nanoliter scale based on a customized microwell chip.


The schematic diagram of MIRALCS. a The flowchart of the MIRALCS. b The box plot of Ct (left) and Tm (right) value of the 20 % Percoll solution (negative control) and 10 pg total RNA (positive control), respectively. c The Ct value and Tm value distribution of 20 % Percoll solution, 10 pg total RNA, non-target well and target well during cDNA amplification process in microwells. The target wells (well with cell) and the non-target wells (without cells) were validated by Agilent 2100 Bioanlyzer. The line denotes Ct median. Horizontal bars denote ± 0.5

Using this method, the research team successfully amplified full-length transcripts of 669 single HeLa S3 cells and 40 of them were randomly selected to perform single-cell RNA sequencing. Based on these data, they obtained a comprehensive understanding of the heterogeneity of HeLa S3 cells in gene expression, alternative splicing and fusions. Furthermore, the researchers identified a high diversity of HPV-18 expression and splicing at the single-cell level.

These results reveal the heterogeneity of a virus-infected cell line. It not only provides a transcriptome characterization of HeLa S3 cells at the single cell level, but is a demonstration of the power of single cell RNA-seq analysis of virally infected cells and cancers.

Wu L et al. (2015) Full-length single-cell RNA-seq applied to a viral human cancer: applications to HPV expression and splicing analysis in HeLa S3 cells. Gigascience 4:51. [article]

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