Nitrophenyl β-D-galactopyranoside, ONPG

The chromogenic substrate Nitrophenyl β-D-galactopyranoside is utilized in quantitative investigations and the kinetics of the beta-galactosidase enzyme.
In molecular biology, it is employed in tests to measure the activity of β-galactosidase reporter gene constructs. This test is frequently used in diagnostics to identify Enterobacteriaceae family lactose-fermenting bacteria.
Two enzymes are required for lactose utilization: β-galactosidase, which breaks down lactose into galactose and glucose, and β-galactoside permease, which catalyzes the transport of lactose into the cell. Without specificity towards lactose, β-Galactosidase can function on basic galactosides, such as the ONPG substrate. Galactose and o-nitrophenol, a yellow chromogenic molecule, are released upon ONPG hydrolysis. ONPG, does not require an induced or constitutive permease enzyme to enter the cell.