Nitrate signals determine the sensing of nitrogen through differential expression of genes involved in nitrogen uptake a
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ORIGINAL PAPER
Nitrate signals determine the sensing of nitrogen through differential expression of genes involved in nitrogen uptake and assimilation in finger millet Alok Kumar Gupta & Vikram Singh Gaur & Sanjay Gupta & Anil Kumar
Received: 12 July 2012 / Revised: 30 January 2013 / Accepted: 4 February 2013 # Springer-Verlag Berlin Heidelberg 2013
Abstract In order to understand the molecular basis of high nitrogen use efficiency of finger millet, five genes (EcHNRT2, EcLNRT1, EcNADH-NR, EcGS, and EcFdGOGAT) involved in nitrate uptake and assimilation were isolated using conserved primer approaches. Expression profiles of these five genes along with the previously isolated EcDof1 was studied under increased KNO3 concentrations (0.15 to 1,500 μM) for 2 h as well as at 1.5 μM for 24 h in the roots and shoots of 25 days old nitrogen deprived two contrasting finger millet genotypes (GE-3885 and GE1437) differing in grain protein content (13.76 and 6.15 %, respectively). Time kinetics experiment revealed that, all the five genes except EcHNRT2 in the leaves of GE-3885 were induced within 30 min of nitrate exposure indicating that there might be a greater nitrogen deficit in leaves and therefore quick transportation of nitrate signals to the leaves. Exposing the plants to increasing nitrate concentrations for 2 h showed that in roots of GE-3885, NR was strongly induced while GS was repressed; however, the pattern was found to be reversed in leaves of GE-1437 indicating that in
Electronic supplementary material The online version of this article (doi:10.1007/s10142-013-0311-x) contains supplementary material, which is available to authorized users. A. K. Gupta : V. S. Gaur : A. Kumar (*) Department of Molecular Biology and Genetic Engineering, College of Basic Sciences and Humanities, GB Pant University of Agriculture and Technology, Pantnagar, Uttarakhand 263145, India e-mail: [email protected] S. Gupta Department of Biotechnology, SBSPGI, Walawala, Dehradun, Uttarakhand, India
GE-3885, most of the nitrate might be reduced in the roots but assimilated in leaves and vice-versa. Furthermore, compared with the low-protein genotype, expression of HNRT2 was strongly induced in both roots and shoots of highprotein genotype at the least nitrate concentration supplied. This further indicates that GE-3885 is a quick sensor of nitrogen compared with the low-protein genotype. Furthermore, expression of EcDof1 was also found to overlap the expression of NR, GS, and GOGAT indicating that Dof1 probably regulates the expression of these genes under different conditions by sensing the nitrogen fluctuations around the root zone. Keywords Gene expression . Real time PCR . Finger millet . Nitrogen responsiveness Abbreviations NRT Nitrate transporter GS Glutamine synthetase GOGAT Glutamine oxoglutarate aminotransferase Dof DNA binding with one finger
Introduction Nitrogen (N) is a crucial plant macronutrient by being the constituent of numerous important compounds, including amino acids, proteins (enzymes), nucleic acids, chl
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