Identification Of The Germplasm With Low Phosphorus Stress Tolerance And Studies On The Tolerant Mechanisms In Tibetan Wild Barley

Low phosphorus (LP) in agricultural soils is a main constraint of crop growth and yield formation worldwide. Overcoming the problem caused by LP stress is quite important for better utilization land resource and increasing crop production in the world. The best and most effective approach in this regard is to develop the crop cultivars with LP stress tolerance or high P use efficeincy. It is therefore important to identify the genetic resources that have high tolerance and to understand the mechanisms of LP stress tolerance in plants. Barley(Hordeum vulgare L) is the fourth important cereal crops in terms of planting areas in the world. Annual wild barley from Qinghai-Tibet Plateau is one of the progenitors of cultivated barley and rich in genetic diversity for the use in barley and other crop improvement. Accordingly, the present study was carried out to elucidate the physiological mechanism in the tolerance to LP stress in Tibetan wild barley, and to identify specific proteins relevant to LP stress tolerance. The main results were summarized as follows:1. Identification of Tibetan wild barley genotypes with high tolerance to low phosphorus stressThe experiments were carried out to evaluate the genotypic difference in low phosphorus (LP,10μM Pi) stress tolerance using104Tibetan wild and20cultivated barley genotypes. The results showed that plant growth and P accumulation were significantly reduced in the plants exposed to LP stress relative to normal P (500μ,M Pi) condition, while some genotypes of wild and cultivated barley increased root length and dry weight under LP stress. There were significant differences among the104Tibetan wild barley genotypes in the growth parameters examined under LP stress, suggesting a wide genetic diversity among the tested genotypes. Significantly genotypic differences were also observed in the relative values of the examined physiological parameters and mineral nutrients in responses to LP stress. Three Tibetan wild barley genotypes, XZ77, XZ99and XZ167, and two cultivated barley genotypes, ZD9and B1034were identified as high tolerance or moderate tolerance to LP.2. The changes in physiological and biochemical traits of Tibetan wild and cultivated barleys in response to LP stressA hydroponic experiment was conducted to investigate the changes in physiological and biochemical traits of Tibetan wild barleys(Hordeum vulgare L. ssp. spontaneum) XZ99(LP tolerant), XZ100(LP sensitive), and cultivated barley ZD9(moderately LP tolerant) under two phosphorus (P) levels during vegetative stage. These genotypes showed considerable differences in the change of biomass accumulation, root/shoot dry weight (DW) ratio, root morphology, organic acid secretion, carbohydrate metabolism, ATPase (Adenosine triphosphatase) activity, P concentration and accumulation under LP in comparison with the control (normal P level). The higher LP stress tolerance of XZ99is associated with more developed roots, enhanced sucrose biosynthesis and hydrolysis of carbohydrate metabolism pathway, higher APase (Acid phosphatase) and ATPase activities, and more secretion of citrate and succinate in roots when plants are exposed to LP condition. In addition, lower reactive oxygen species (ROS) on lateral roots of XZ99indicates the role of ROS in regulating the formation of lateral roots and root hairs. Furthermore, maintaining of the normal root cell structure was considered as an adaptative strategy of XZ99to LP stress. The results prove the potential of Tibetan wild barley in developing barley cultivars with high tolerance to LP stress and understanding the mechanisms of LP stress tolerance in plants.3. Identification of the differentially accumulated proteins associated with low phosphorous stress tolerance in a Tibetan wild barley accessionA comparatively proteomic analysis was conducted using three barley genotypes differing in LP stress tolerance so as to reveal the mechanisms underlying XZ99in LP stress tolerance. Totally,31differentially accumulated proteins were identified and compared in the roots and leaves of the three genotypes, and these proteins are involved in the many biological processes, including metabolism, signal transduction. cell growth and division, and stress defense. In comparison with XZ100and ZD9, XZ99had more developed root system, which is mainly attributed to enhanced carbohydrate metabolizing proteins under LP condition. Moreover, among the indentified16proteins preferentially accumulated under LP condition in roots and leaves.9proteins were specifically accumulated in XZ99. In addition, the accumulated levels of the9LP responsive proteins were verified by qRT-PCR. This is the first report on the proteomic analysis of LP stress tolerance in wild barley and the results proved the existence of the differentially accumulated proteins associated with tolerance to LP condition in the Tibetan wild barley. 4. The regulation of root growth in response to phosphorus deficiency mediated by phytohormones and sugar signaling in a Tibetan wild barley accessionA pot experiment was conducted to investigate the mechanisms of altered root system architecture (RSA) in response to LP stress. In general. LP stress tolerant genotype XZ99differed from LP stress sensitive genotype XZ100and moderate LP stress tolerant genotype ZD9in terms of altered RSA. It was found that sucrose, phytohormone (mainly auxin and ethylene) and phosphate (Pi) co-ordinate the local and systemic signal pathway in response to the LP stress, as a result enhancing Pi acquisition by plants. Changes in RSA mediated by local signals and systemic signals enhanced translocation of sucrose to roots via phloem, resulting in promoted root growth. More importantly, in this pathway auxin-ethylene regulated gene (TIR1, ARF19and ACCO), sucrose transpoter2(SUC2) gene, and phosphate transporter (Phtl;1, Phtl;2) gene were up-regulated in response to LP stress in XZ99roots. The current results made the first insight into the roles of plant hormones and sucrose as signals in regulation of LP stress response and tolerant development.