Luciferase Reporter Assay

Overview

Luciferase reporter assays measure the transcriptional or enhancer activity of a DNA element by cloning it upstream of the firefly or Renilla luciferase gene and transfecting the construct into cells. Luciferase enzyme activity (measured by bioluminescence) is proportional to transcriptional output from the inserted element. Allele-specific reporter assays compare constructs carrying the reference and alternate alleles of a candidate causal SNP to quantify the effect of the variant on regulatory activity. These assays are routinely used to validate GWAS functional candidates and FGFR 3’UTR regulatory hypotheses.

Used by

Allele-specific luciferase reporters in LNCaP, DU145, 22Rv1, PC3, and VCaP cells (±DHT) gave significantly higher activity for the rs4519489 A allele than the T allele across all cell lines and conditions, confirming allele-specific enhancer activity at the 2p25 NOL10/USF1 prostate cancer risk locus PMID:28927585
Luciferase reporter assays in HEK293T and H69 (cholangiocyte) cells confirmed that the intact FGFR2 3’UTR represses FGFR2 expression; truncating rearrangements removing the 3’UTR in CCA Cluster 4 tumors explain elevated FGFR2 transcript levels PMID:28667006

Notes

Allele-specific assays use paired constructs differing only at the SNP position to isolate the effect of the variant.
DHT (dihydrotestosterone) treatment used to assess androgen-dependent vs androgen-independent enhancer activity at the rs4519489 locus.

Sources

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