Date of Award
Master of Science (MS)
Elizabeth A. Weretilnyk
In global soils phosphorus is both the most abundant yet the least available essential macronutrient for plants. Phosphorus is a component of nucleic acids, some signalling molecules, membrane lipids and, as ATP, for metabolic control and energy transfer. Consequently, a phosphorus deficiency impacts many aspects of growth and development leading to plants that are smaller in stature, develop slowly and yield less than plants with sufficient phosphate.
Diversity among native plants with respect to phosphorus use can allow researchers to identify traits meriting consideration for crop improvement. Thus the objective of this thesis research was to study the response of a native crucifer found in the Yukon Territory, Thellungiella salsuginea, to various concentrations of external phosphate. Seedlings were grown on nutrient medium containing O.OS , 0.1 , 0.2 or 0.5 mM phosphate. No significant differences were found with respect to root elongation rates for concentrations below 0.2 mM phosphate but root elongation was 1.4-fold faster for seedlings grown on 0.5 mM phosphate. Thellungiella seedlings grown on low (0.05 mM) phosphate medium showed a 2.5-fold increased biomass allocation to roots when compared to plants on high (0.5 mM) phosphate medium consistent with an increased allocation to root mass under low phosphate seen with plants optimizing foraging for limiting nutrients. Even at the lowest external Pi content no seedlings showed the classic phosphate starvation response typified by short primary roots with many lateral branches.
The lack of a starvation phenotype raised questions of how Thellungiella allocates phosphate to major sinks, one being phospholipids. Therefore, a 31p_NMR profiling strategy was developed for leaf phospholipids. This approach required more tissue than was available from seedlings so rosette leaves were harvested from five-week-old plants grown in PromixBX and watered weekly with a low (0.05 mM) or high (1.0 mM) phosphate solution. Shoot fresh weight was not significantly different between treatments and visual inspection revealed no overt phenotypic differences. Shoot tissue was solventextracted to yield polar and non-polar phases. The polar phase of plants grown under low phosphate conditions contained significantly less (1.3-fold) inorganic phosphate (Pi) than plants watered with 1.0 mM phosphate (P < 0.05). Non-polar phases dissolved in 5% (w/v) sodium cholate produced high resolution spectra by 31 p-NMR. Spectra from low and high phosphate-treated plant samples usually showed five peaks tentatively identified as free Pi, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol and phosphatidylcholine. Statistical analysis indicated that only the putative phosphatidylinositol peak showed treatment-related differences, with low phosphate plants showing about 50% of the lipid species relative to high phosphate plants.
Use of the Electronic Reference To Access In Vivo Concentrations (ERETIC) method was tested on a limited number of extracts and results show this to be a promising approach for quantifying phospholipid species profiled by 31 p_NMR. Moreover, this profiling protocol was successfully extended to Arabidopsis tissue producing spectra with a signal-to-noise comparable to those from Thellungiella samples despite using 2-fold less tissue. In the case of Arabidopsis, only four distinct peaks were identified with the one attributed to phosphatidylinositol apparently absent or below the detection limits. The difference in spectra found between Thellungiella and Arabidopsis plants warrants further investigation.
DiBattista, Alicia, "Growth and Phospholipid Analysis of Thellungiella salsuginea Exposed to Varying Environmental Phosphate" (2010). Open Access Dissertations and Theses. Paper 4098.
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