Description

QIAscout and NGS applications for identification of subpopulation-specific variants

Ruth Kläver, Andrea Janosch, Charline Bemmann, Claudia Kappmeier, Dorothee Unt, Frank Reinecke, Divya Vijay Pratheek, Norbert Hochstein

QIAGEN GmbH, QIAGEN Straße 1, 40724 Hilden, Germany

Although single cell research is currently gaining momentum, it remains challenged by the lack of affordable methods to precisely isolate a single cell from a heterogeneous cell population without manipulating the cellular status. QIAscout is an effective and fast method to isolate viable single cells ensuring minimal disturbance of the cell. For isolation, cells are seeded on an array consisting of 12.000 microrafts which can be pierced, dislodged and transferred to reaction tubes for further processing. This novel single cell isolation method works in conjunction with inverted microscopes and is the ideal method to separate single cells for further downstream analysis or cultivation of clonal subpopulations.

Due to the heterogeneity of a cell population, methods are needed to detect cell type specific variants and to reliably identify isolated single cells using these specific variants. In this study, we used the QIAscout technology to isolate 44 single cells of two different cell lines (22 LoVo and 22 SW48) and to detect cell type specific variants using whole genome amplification and low pass sequencing followed by census-based variant identification. In a second experiment, we were able to selectively isolate single cells of one cell type from a heterogeneous cell population using the QIAscout. Pyrosequencing technology was used to prove the successful and specific isolation of single cells by detecting cell line specific SNPs that were identified in the initial experiment.

We demonstrated that the QIAscout technology allows efficient and reliable isolation of single targeted cells. We further showed that QIAscout isolated single cells can be used to detect subpopulation specific SNPs using whole genome amplification and low pass sequencing followed by the census-based variant calling method.