This dataset contains the amoA genes on three size-fractionated (> 3.0 μm, 0.45‒3.0 μm, and 0.22‒0.45 μm) ammonia oxidizers and relevant environmental parameters observed along a salinity gradient in the Pearl River estuary, China during July 2014. The temperature and salinity were determined using the CTD system. Gene-related data was obtained using qPCR analysis. Nutrients were determined by an automatic analysis system (Bran-Lube, GmbH). All data were aimed to distinguish the distribution and bioactivity of ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) in the Pearl River estuary.