High-throughput functional analysis of regulatory variants using a massively parallel reporter assay

Kate Delfosse; Chiara Gerhardinger; John L Rinn; Philipp G Maass

doi:10.1016/j.xpro.2023.102731

High-throughput functional analysis of regulatory variants using a massively parallel reporter assay

STAR Protoc. 2023 Dec 15;4(4):102731. doi: 10.1016/j.xpro.2023.102731. Epub 2023 Nov 18.

Authors

Kate Delfosse¹, Chiara Gerhardinger², John L Rinn³, Philipp G Maass⁴

Affiliations

¹ Genetics & Genome Biology Program, The Hospital for Sick Children, Toronto, ON M5G 0A4, Canada.
² Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA.
³ BioFrontiers Institute, University of Colorado Boulder, Boulder, CO 80303, USA; Department of Biochemistry, University of Colorado Boulder, Boulder, CO 80303, USA.
⁴ Genetics & Genome Biology Program, The Hospital for Sick Children, Toronto, ON M5G 0A4, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada. Electronic address: philipp.maass@sickkids.ca.

Abstract

Association studies describe genetic associations between noncoding variants and disease susceptibility; however, they do not provide functional insight into the underlying molecular mechanisms of these variants. We present a protocol to assay the regulatory potential of thousands of noncoding variants using massively parallel reporter assays. We describe steps for oligo design, generating a plasmid pool, and extracting tag-seq libraries from cells to quantify the tested sequences. For complete details on the use and execution of this protocol, please refer to Oliveros and Delfosse et al.¹.

Keywords: Gene Expression; Genetics; Genomics; High Throughput Screening; Systems biology.

MeSH terms

Plasmids* / genetics