Accurately Determined Expression Levels of Nitrate and Phosphate Transporter Genes in the Diatom, Skeletonema subsalsum, at a Coastal Site in the East China Sea

Diatom cells possess a distinctive nitrate transporter (encoded by Nrt2) and a sodium/phosphate cotransporter (encoded by Npt2bl) to enable the uptake of these nutrients. The expression levels of Nrt2 and Npt2bl are sensitive to nutrient availability, and this relationship has been used to evaluate the intensities of nitrogen and phosphorous stress, respectively, in natural assemblages of diatoms. In this study, research cruises were conducted in the coastal region of southern East China Sea in June and July, 2017. During each cruise, a 24-h sampling program was established with additional nutrient-manipulating experiments performed at 2 time points. Meta-transcriptomes were constructed and analyzed to confirm that Skeletonema subsalsum was the dominant diatom in samples, and reads belong to this species were sorted out. These reads were aligned to reveal the precise sequences of Nrt2 and Npt2bl, which led to the design of specific primers. Next, quantitative PCR was applied to measure the mRNA abundances. Our results indicated that the gene expression of Nrt2 in S. subsalsum reached high levels in the nitrogen-free treatment, but decreased to low levels in the ammonium-addition treatment. Similarly, the high and low expressions of Npt2bl were observed in phosphate-free and phosphate-addition treatments, respectively. Comparing the results of nutrient-manipulating experiments to those of untreated samples, the in situ expressions of Nrt2 were similar to ammonium-addition treatments in June and July, indicating that S. subsalsum did not actively produce nitrate transporter. By contrast, gene expressions of Npt2bl revealed that S. subsalsum actively produced phosphate transporter in June, but the effort was moderate in July.