Relationships between the expression of nutrient uptake genes and ambient ammonium and phosphate concentrations in the diatom, Skeletonema costatum

• Main Authors: Chih-Yu Chuang, 莊智宇
• Other Authors: Ching-Hua Hu
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/78524960735568871544

碩士 === 國立臺灣海洋大學 === 生物科技研究所 === 101 === Media with different concentrations of ammonium and phosphate were prepared to study how nitrogen (N) and phosphorus (P) stresses influence the expression of genes related to nutrient uptake in the diatom, Skeletonema costatum. In addition, genes sensitively regulated under nitrogen and phosphorus stresses may serve as candidates of molecular indicators in the sea. In experimental design, S. costatum was cultured in a combination matrix formed by ammonium concentrations at 500, 200, 100, 50 μM, and phosphate concentrations at 36.3, 18, 9, 2 μM, respectively. The mRNA expression of several genes were measured by real-time quantitative reverse transcription polymerase chain reaction. These genes were the nitrate transporter gene (Nrt2), the type III phosphate transporter gene (PHO）, the type II phosphate transporter gene (Npt2bL), and the alkaline phosphatase gene (AP1). The median of Nrt2 expression in nutrient sufficient, N-deficient, P-deficient media was 0.23, 52.5, and 0.12 μmol (mol 18S rRNA )-1, respectively. Nrt2 mRNA expression level rose on days 3 and 4, which was 1000-fold higher than low values on days 1 and 2. Nrt2 expression had a negative linear relationship (P &;lt;0.001) with the initial ammonium concentration. The median of PHO expression in nutrient sufficient, N-deficient, P-deficient media, was 143, 4.76, and 258 μmol (mol 18S rRNA )-1. PHO expression did not change much in N sufficient media, but declined 100-fold in N-deficient media. A positive linear relationship (P &;lt;0.001) was observed between PHO expression and the initial ammonium concentration. The median of Npt2bL expression in nutrient sufficient, N-deficient, and P-deficient media was 404, 151, and 1267 μmol (mol 18S rRNA )-1. Npt2bL expression increased with incubation time and reached high levels 30-times the low values on days 1 and 2. Npt2bL expression had a positive linear relationship (P &;lt;0.001) with N/P ratio. The median of AP1 expression in nutrient sufficient, N-deficient, and P-deficient media was 17.1, 2.49, and 39.3 μmol (mol 18S rRNA )-1. AP1 expression rose 10-times in N sufficient media. There was a positive linear relationship (P &;lt;0.001) between AP1 expression and N/P ratio. According to the results, Nrt2 expression at 1.98 μmol (mol 18S rRNA )-1 was apparently a good threshold for nitrogen starvation. However, there exist no suitable phosphorus indicator genes that possess a clear-cut expression pattern between P-stressed and nutrient replete conditions.