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University of the Western Cape (2016)

Comparative analysis of molecular and physiological responses of two canola genotypes to drought stress

Nkomo, Mbukeni Andrew

Titre : Comparative analysis of molecular and physiological responses of two canola genotypes to drought stress

Auteur : Nkomo, Mbukeni Andrew

Université de soutenance : University of the Western Cape

Grade : Magister Scientiae – MSc 2016

Résumé
Food security has always been one of the priority concerns in Africa, and it is mostly threatened by drought stress due to climate change. Drought-induced stress is one of the serious limiting factors of plant production, and it is known to impose oxidative stress as a consequence of excessive reactive oxygen species (ROS) accumulation that lead to lipid peroxidation, which is manifested as increased cell death. Hence, this study investigated the influence of drought stress on two contrasting canola genotypes (Agamax and Garnet), by monitoring their physiological and molecular changes. The results showed that the plant growth and biomass of both genotypes were significantly affected by drought stress as a consequence of excessive ROS accumulation (manifested as H₂O₂ and OH· content). However, under drought stress conditions, the reduction in biomass and shoot length was more pronounced in the Garnet genotype when compared to that of the Agamax genotype. This was further supported by the increase in lipid peroxidation and cell death, which were shown to be significantly higher in the Garnet genotype when compared to the Agamax genotype under drought stress. ROS scavenging ability which prevents oxidative stress and ultimately ROS-induced cellular damage. Hence, given the higher levels of antioxidant activity coupled with the reduction in ROS accumulation that was observed in the Agamax genotype, we suggest that the Agamax genotype might be slightly less susceptible to drought stress, when compared to the Garnet genotype. Furthermore, understanding the proteomic responses of these two contrasting genotypes that showed a marked difference in response to drought stress might help in unlocking complex biological networks of proteins underlying drought stress tolerance. Hence we use two-dimensional (2D) gel electrophoresis coupled with Matrix assisted laser desorption/ionisation-time of flight/time of flight tandem mass spectrometry (MALDI TOFTOF MS) analysis for this part of the study, in order to detect and analyze those differentially expressed proteins or proteins whose abundance levels were influenced as a consequence of drought stress. To gain additional insight into the leaf proteomes of the two canola genotypes, a protamine sulphate precipitation (PSP) method was used to remove RuBisCo and confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis. A total of 55 well resolved protein spots were selected for mass spectrometry analysis of which 31 (56%) were positively identified using the selective criteria analysis (SCA). All positively identified proteins were then classified into functional categories including protein folding (3%), photosynthetic (29%), detoxification and protection (20%), and energy related proteins whereas 16% could not be classified into any functional category. Apart from spot 32 (Fe superoxide dismutase) and spot 34 (chloroplast beta-carbonic anhydrase), no further significant difference in protein expression/abundance was observed for all the identified proteins for both genotypes in response to drought stress. Both proteins (spots 32 and 34) have been shown to contain antioxidant activity properties which suggest that they might play a crucial role in improving drought stress tolerance in canola plants.

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