Abstract
Single cell multi-omics analysis has the potential to yield a comprehensive understanding of the cellular events that underlie the basis of human diseases. The cardinal feature to access this information is the technology used for single-cell isolation, barcoding, and sequencing. Most currently used single-cell RNA-sequencing platforms have limitations in several areas including cell selection, documentation and library chemistry. In this study, we describe a novel high-throughput, full-length, single-cell RNA-sequencing approach that combines the CellenONE isolation and sorting system with the ICELL8 processing instrument. This method offers substantial improvements in single cell selection, documentation and capturing rate. Moreover, it allows the use of flexible chemistry for library preparations and the analysis of living or fixed cells, whole cells independent of sizing and morphology, as well as of nuclei. We applied this method to dermal fibroblasts derived from six patients with different segmental progeria syndromes and defined phenotype associated pathway signatures with variant associated expression modifiers. These results validate the applicability of our method to highlight genotype-expression relationships for molecular phenotyping of individual cells derived from human patients.
Highlights
IntroductionSingle cell multi-omics analysis has the potential to yield a comprehensive understanding of the cellular events that underlie the basis of human diseases
The most critical step for obtaining transcriptome and genome information from individual cells is the single cell isolation, and it is necessary to distinguish between methods with low- or high-throughput for the collection of single cells
The advantages of ICELL8 include flexible chemistry for transcript coverage (3’ end fragment or full-length approaches), flexible sequencing mode selection choosing between single-end (SE) or paired-end (PE) mode, compatible for both living as well as fixed cells with a large range of sizes (3 μm to 500 μm in diameter) and dispensing of cells with abnormal morphology e.g., Schwann cells that exhibit axonic regrowing forms and sizing of 200–500 μm, as we show in this paper for the first time (Fig. 1b and Supplementary Fig. 1)
Summary
Single cell multi-omics analysis has the potential to yield a comprehensive understanding of the cellular events that underlie the basis of human diseases. We describe a novel high-throughput, full-length, single-cell RNA-sequencing approach that combines the CellenONE isolation and sorting system with the ICELL8 processing instrument This method offers substantial improvements in single cell selection, documentation and capturing rate. It allows the use of flexible chemistry for library preparations and the analysis of living or fixed cells, whole cells independent of sizing and morphology, as well as of nuclei. We applied this method to dermal fibroblasts derived from six patients with different segmental progeria syndromes and defined phenotype associated pathway signatures with variant associated expression modifiers. Several studies explored the utility of this platform for single-cell sequencing in the cardiac field, due to the fact that cardiomyocyte size limits the use of drop-seq and other p latforms[7,8]
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