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High‐affinity nitrate transporters are considered to be the major transporter system for nitrate uptake in diatoms. In the diatom genus Skeletonema, three forms of genes encoding high‐affinity nitrate transporters (NRT2) were newly identified from transcriptomes generated as part of the marine microbial eukaryote transcriptome sequencing project. To examine the expression of each form of NRT2 under different nitrogen environments, laboratory experiments were conducted under nitrate‐sufficient, ammonium‐sufficient, and nitrate‐limited conditions using three ecologically important Skeletonema species: S. dohrnii, S. menzelii, and S. marinoi. Primers were developed for each NRT2 form and species and Q‐RT‐PCR was performed. For each NRT2 form, the three Skeletonema species had similar transcriptional patterns. The transcript levels of NRT2:1 were significantly elevated under nitrogen‐limited conditions, but strongly repressed in the presence of ammonium. The transcript levels of NRT2:2 were also repressed by ammonium, but increased 5‐ to 10‐fold under nitrate‐sufficient and nitrogen‐limited conditions. Finally, the transcript levels of NRT2:3 did not vary significantly under various nitrogen conditions, and behaved more like a constitutively expressed gene. Based on the observed transcript variation among NRT2 forms, we propose a revised model describing nitrate uptake kinetics regulated by multiple forms of nitrate transporter genes in response to various nitrogen conditions in Skeletonema. The differential NRT2 transcriptional responses among species suggest that species‐specific adaptive strategies exist within this genus to cope with environmental changes.