title: ESTABLISHMENT OF MINIMAL AND MAXIMAL TRANSCRIPT LEVELS FOR NITRATE TRANSPORTER GENES FOR DETECTING NITROGEN DEFICIENCY IN THE MARINE PHYTOPLANKTON ISOCHRYSIS GALBANA (PRYMNESIOPHYCEAE) AND THALASSIOSIRA PSEUDONANA (BACILLARIOPHYCEAE)(1).
abstract: Nitrate transporter genes (Nrt2) encode high-affinity nitrate transporters in marine phytoplankton, and their transcript levels are potential markers of nitrogen deficiency in eukaryotic phytoplankton. For the proper interpretation of measured Nrt2 transcript abundances, a relative expression assay was proposed and tested in Isochrysis galbana Parke (Prymnesiophyceae) and Thalassiosira pseudonana (Hust.) Hasle et Heimdal (Bacillariophyceae). The minimal transcript levels of Nrt2 genes were achieved by the addition of 100 μM ammonium, which led to a rapid decline in Nrt2 transcripts in 10-30 min. Experiments using a concentration series revealed that the effective dosage of ammonium to create a minimal transcript level of ∼1 μmol · mol(-1) 18S rRNA was ≥25 μM in both species. On the other hand, the addition of l-methionine sulfoximine (MSX), an inhibitor of glutamine synthetase, enhanced the Nrt2 transcript level in I. galbana but did not affect that in T. pseudonana. Nitrogen deprivation was used as an alternative means to create maximal Nrt2 transcript levels. By transferring cells into N-free medium for 24 h, Nrt2 transcript levels increased to ∼90 μmol · mol(-1) 18S rRNA in I. galbana, and to ∼800 μmol · mol(-1) 18S rRNA in T. pseudonana. The degree of nitrogen deficiency thus can be determined by comparing original Nrt2 transcript levels with the minimal and maximal levels.
authors.affiliation: Department of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Cellular and Organismic Biology, Academia Sinica, Taipei 11529, TaiwanDepartment of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Clinical Nutrition, Hung Kuang University of Technology, Taichung 43302, TaiwanInstitute of Marine Biology, Institute of Marine Environmental Chemistry and Ecology, and Center for Marine Bioscience and Biotechnology, National Taiwan Ocean University, Keelung 20224, Taiwan.
authors.forename: Lee-Kuo
authors.initials: LK
authors.lastname: Kang
authors.affiliation: Department of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Cellular and Organismic Biology, Academia Sinica, Taipei 11529, TaiwanDepartment of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Clinical Nutrition, Hung Kuang University of Technology, Taichung 43302, TaiwanInstitute of Marine Biology, Institute of Marine Environmental Chemistry and Ecology, and Center for Marine Bioscience and Biotechnology, National Taiwan Ocean University, Keelung 20224, Taiwan.
authors.forename: Sheng-Ping L
authors.initials: SP
authors.lastname: Hwang
authors.affiliation: Department of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Cellular and Organismic Biology, Academia Sinica, Taipei 11529, TaiwanDepartment of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Clinical Nutrition, Hung Kuang University of Technology, Taichung 43302, TaiwanInstitute of Marine Biology, Institute of Marine Environmental Chemistry and Ecology, and Center for Marine Bioscience and Biotechnology, National Taiwan Ocean University, Keelung 20224, Taiwan.
authors.forename: Hsing-Juh
authors.initials: HJ
authors.lastname: Lin
authors.affiliation: Department of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Cellular and Organismic Biology, Academia Sinica, Taipei 11529, TaiwanDepartment of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Clinical Nutrition, Hung Kuang University of Technology, Taichung 43302, TaiwanInstitute of Marine Biology, Institute of Marine Environmental Chemistry and Ecology, and Center for Marine Bioscience and Biotechnology, National Taiwan Ocean University, Keelung 20224, Taiwan.
authors.forename: Pei-Chung
authors.initials: PC
authors.lastname: Chen
authors.affiliation: Department of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Cellular and Organismic Biology, Academia Sinica, Taipei 11529, TaiwanDepartment of Life Sciences, National Chung Hsing University, Taichung 40227, TaiwanInstitute of Clinical Nutrition, Hung Kuang University of Technology, Taichung 43302, TaiwanInstitute of Marine Biology, Institute of Marine Environmental Chemistry and Ecology, and Center for Marine Bioscience and Biotechnology, National Taiwan Ocean University, Keelung 20224, Taiwan.
authors.forename: Jeng
authors.initials: J
authors.lastname: Chang
keywords: Isochrysis galbana; Thalassiosira pseudonana; high-affinity nitrate transporter genes; nitrogen deficiency; relative expression assay