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Item Molecular cloning and characterization of a membrane associated NAC family gene, SiNAC from foxtail millet [Setaria italica (L.) P. Beauv.](Springer Science, 2011) Puranik, Swati; Bahadur, Ranjit Prasad; Srivastava, Prem S.; Prasad, ManojThe plant-specific NAC (NAM, ATAF, and CUC) transcription factors have diverse role in development and stress regulation. A transcript encoding NAC protein, termed SiNAC was identified from a salt stress subtractive cDNA library of S. italica seedling (Puranik et al., J Plant Physiol 168:280-287, 2011). This single/low copy gene containing four exons and four introns within the genomic-sequence encoded a protein of 462 amino acids. Structural analysis revealed that highly divergent C terminus contains a transmembrane domain. The NAC domain consisted of a twisted antiparallel beta-sheet packing against N terminal alpha helix on one side and a shorter helix on the other side. The domain was predicted to homodimerize and control DNA-binding specificity. The physicochemical features of the SiNAC homodimer interface justified the dimeric form of the predicted model. A 1539 bp fragment upstream to the start codon of SiNAC gene was cloned and in silico analysis revealed several putative cis-acting regulatory elements within the promoter sequence. Transactivation analysis indicated that SiNAC activated expression of reporter gene and the activation domain lied at the C terminal. The SiNAC:GFP was detected in the nucleus and cytoplasm while SiNAC ΔC(1-158):GFP was nuclear localized in onion epidermal cells. SiNAC transcripts mostly accumulated in young spikes and were strongly induced by dehydration, salinity, ethephon, and methyl jasmonate. These results suggest that SiNAC encodes a membrane associated NAC-domain protein that may function as a transcriptional activator in response to stress and developmental regulation in plants.Item Comparative transcriptome analysis of contrasting foxtail millet cultivars in response to short-term salinity stress(Elsevier, 2011) Puranik, Swati; Jha, Sarita; Srivastava, Prem S.; Sreenivasulu, Nese; Prasad, ManojSoil salinity represents a major abiotic stress that adversely affects crop growth and productivity. In this study, 21-day-old seedlings of two foxtail millet (Setaria italica) cultivars differing in salt tolerance were found to also differ in lipid peroxidation, ion balance and activity of antioxidative enzymes (glutathione reductase and catalase) under short-term salinity stress (250 mM NaCl for 1-48 h). With the aim of better understanding the molecular mechanisms underlying plant responses to short-term salinity stress, two suppression subtractive hybridization cDNA libraries (forward and reverse) were constructed of these cultivars. A total of 249 non-redundant ESTs was identified by random EST sequencing and grouped into 11 functional categories. Macroarray analysis of these clones showed that 159 (63.9%) were differentially expressed (≥ 2-fold) in response to salinity stress, with 115 (72.3%) up and 44 (27.7%) down-regulated. A data search of transcriptional profiling under salinity stress in other species revealed that 81 (51%) of the 159 differentially expressed transcripts found in foxtail millet have not been reported in previous studies. Hence, these new transcripts may represent untapped gene sources allowing specific responses to short-term salt-stress in an orphan crop known to possess a natural adaptation capacity to abiotic stress. Quantitative real-time PCR of 21 highly up-regulated (≥2.5-fold) transcripts showed temporal variation in expression in both cultivars under salinity. Among them, several transcription factors and signalling genes were preferentially expressed in the tolerant cultivar. These results suggest that the tolerant cultivar possesses more effective signal-perception mechanisms for metabolic adjustments in plants under harsh saline conditions. Our findings provide evidence that the unknown genes identified in this study, in addition to several known genes, may play important roles in stress tolerance mechanisms present in foxtail millet.
