The rice landrace Horkuch, endemic to the southern saline coast of Bangladesh, is known to have salt tolerance traits and can therefore contribute to a high yielding recipient for breeding purposes. In this study, we reciprocally crossed Horkuch with high yielding but salt sensitive IR29 to detect the complement of genes that were responsible for conferring salt tolerance versus sensitivity at the seedling developmental stage. We looked at tolerant and sensitive F 3 families from individual F 2 segregating plants and analyzed them for differential gene expressions using RNAseq. In general, we observed higher numbers of genes differentially expressed in leaves compared to root tissues. This included both upregulation and downregulation of gene expression across our experimental factors. Gene expression decreased in sensitive leaf after stress exposure where tolerant plants showed the opposite trend. In root, tolerant plants expression decreased at higher time points of stress exposure. We also observed a strong maternal cytoplasmic effect on gene expression and this was most evident in roots where there was upregulation in functional enrichments related to phosphorylation, electron carriers, transporter and cation transmembrane activities. Stress groups (tolerant and sensitive) response in F 3 families were distinctive in both cytoplasmic backgrounds and involved uniquely upregulated genes in tolerant progenies including membrane sensor proteins, enzymes involved with signaling pathways, such as those producing trehalose and G-protein coupled receptor proteins, photosynthesis-related enzymes and golgi body recycling as well as prolamin precursor proteins involved in refolding of proteins. On the other hand, sensitivity was found to be associated with differential upregulation of only a few redox proteins and higher number of apoptosis related genes compared to the tolerant response. Overall, our highly replicated experimental design was powerful and allowed the detection of relatively subtle differential expression. Our future goal is to correlate these expression differences with QTLs in this population, which would help identify the relative importance of specific genetic loci and provide a direct avenue for combining higher levels of salt tolerance with better agronomic traits in rice.
Soil salinity is one of the most challenging problems that restricts the normal growth and production of rice worldwide. It has therefore become very important to produce more saline tolerant rice varieties. This study shows constitutive over-expression of the vacuolar Na+/H+ antiporter gene (OsNHX1) from the rice landrace (Pokkali) and attainment of enhanced level of salinity tolerance in transgenic rice plants. It also shows that inclusion of the complete un-translated regions (UTRs) of the alternatively spliced OsNHX1 gene provides a higher level of tolerance to the transgenic rice. Two separate transformation events of the OsNHX1 gene, one with 1.9 kb region containing the 5′ UTR with CDS and the other of 2.3 kb, including 5′ UTR, CDS, and the 3′ UTR regions were performed. The transgenic plants with these two different constructs were advanced to the T3 generation and physiological and molecular screening of homozygous plants was conducted at seedling and reproductive stages under salinity (NaCl) stress. Both transgenic lines were observed to be tolerant compared to WT plants at both physiological stages. However, the transgenic lines containing the CDS with both the 5′ and 3′ UTR were significantly more tolerant compared to the transgenic lines containing OsNHX1 gene without the 3′ UTR. At the seedling stage at 12 dS/m stress, the chlorophyll content was significantly higher (P < 0.05) and the electrolyte leakage significantly lower (P < 0.05) in the order 2.3 kb > 1.9 kb > and WT lines. Yield in g/plant in the best line from the 2.3 kb plants was significantly more (P < 0.01) compared, respectively, to the best 1.9 kb line and WT plants at stress of 6 dS/m. Transformation with the complete transcripts rather than the CDS may therefore provide more durable level of tolerance.
Proper management of nutrients in agricultural systems is critically important for maximizing crop yields while simultaneously minimizing the health and environmental impacts of pollution from fertilizers. These goals can be achieved by timely confirmatory diagnostics of nutrient deficiencies in plants, which enable precise administration of fertilizers and other supplementation in fields. Traditionally, nutrient diagnostics are performed by wet-laboratory analyses, which are both time-and labor-consuming. Unmanned aerial vehicle (UAV) and satellite imaging have offered a non-invasive alternative. However, these imaging approaches do not have sufficient specificity, and they are only capable of detecting symptomatic stages of nutrient deficiencies. Raman spectroscopy (RS) is a non-invasive and non-destructive technique that can be used for confirmatory detection and identification of both biotic and abiotic stresses on plants. Herein, we show the use of a hand-held Raman spectrometer for highly accurate presymptomatic diagnostics of nitrogen, phosphorus, and potassium deficiencies in rice (Oryza sativa). Moreover, we demonstrate that RS can also be used for pre symptomatic diagnostics of medium and high salinity stresses. A Raman-based analysis is fast (1 s required for spectral acquisition), portable (measurements can be taken directly in the field), and label-free (no chemicals are needed). These advantages will allow RS to transform agricultural practices, enabling precision agriculture in the near future.
Global increase in salinity levels has made it imperative to identify novel sources of genetic variation for tolerance traits, especially in rice. The rice landrace Horkuch, endemic to the saline coastal area of Bangladesh, was used in this study as the source of tolerance in reciprocal crosses with the sensitive but high-yielding IR29 variety for discovering transcriptional variation associated with salt tolerance in the resulting populations. The cytoplasmic effect of the Horkuch background in leaves under stress showed functional enrichment for signal transduction, DNA-dependent regulation and transport activities. In roots the enrichment was for cell wall organization and macromolecule biosynthesis. In contrast, the cytoplasmic effect of IR29 showed upregulation of apoptosis and downregulation of phosphorylation across tissues relative to Horkuch. Differential gene expression in leaves of the sensitive population showed downregulation of GO processes like photosynthesis, ATP biosynthesis and ion transport. Roots of the tolerant plants conversely showed upregulation of GO terms like G-protein coupled receptor pathway, membrane potential and cation transport. Furthermore, genes involved in regulating membrane potentials were constitutively expressed only in the roots of tolerant individuals. Overall our work has developed genetic resources and elucidated the likely mechanisms associated with the tolerance response of the Horkuch genotype.
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