Galacto-Light Plus™ β-Galactosidase Reporter Gene Assay System

Protocol tips

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	Add diluted Galacton-Plus substrate 1:100 with Reaction Buffer Diluent. Add 70µL of Reaction Buffer, mix, and incubate for 30-60 min

Protocol tips
Add diluted Galacton-Plus substrate 1:100 with Reaction Buffer Diluent. Add 70µL of Reaction Buffer, mix, and incubate for 30-60 min

Publication protocol

The cells were transfected with DGKα siRNAs for 4 hours using oligofectamine or treated with DGKα inhibitor R59022 10 µM, PDE4 inhibitor rolipram 40 µM, exogenous cAMP 20 or 80 pmol for 3 days and subsequently transfected with β-galactosidase (2 ng/ul), mTOR promoter luciferase reporter or empty promoter vector (Switchgear Genomics Inc., Menlo Park, CA) following the manufacturer’s instructions for 48 hours. Luciferase assays for mTOR activity were performed using the LightSwitch Assay System (Switchgear Genomics Inc.) and for β-galactosidase activity using Galacto-Light Plus™ beta-Galactosidase Reporter Gene Assay System (Applied Biosystems, Carlsbad, CA). Luminescence was measured on a Promega GloMax 20/20 luminometer and normalized as described previously (6). mTOR luciferase activities were double-normalized by dividing each well by both β-galactosidase activity and the average luciferase/β-galactosidase value in a parallel set done with constitutively-expressed luciferase expression vector.

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Manufacturer protocol

Download the product protocol from Thermo Fisher Scientific for Galacto-Light Plus™ β-Galactosidase Reporter Gene Assay System below.

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