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Molecular Cloning and Functional Expression of Glyphosates Target Enzyme (EPSPS) in C3 turfgrass species

Final Activity Report Summary - EPSPS (Molecular Cloning and Functional Expression of Glyphosates Target Enzyme (EPSPS) in C3 turfgrass species)

The herbicide glyphosate competitively inhibits 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) enzyme (EC.2.5.1.19) which is a nuclear-encoded plastid localized enzyme involved in biosynthesis of aromatic amino acids. Upon inhibition of EPSPS by glyphosate, shikimic acid, a metabolic precursor of shikimate-3-phosphate (S3P), has been reported to accumulate rapidly in sensitive plants. Glyphosate is also known to effect production of chlorophyll, cytochromes and micronutrient balance of plants. In order to investigate the physiological response of turfgrass species to glyphosate, chlorophyll content, dry matter production, shikimate accumulation and micro and macronutrient elements uptake were determined for control, 5 % (1.58mM) 10 % (3.16mM) 20 % (6.32mM) and 100 % (31.55mM; recommended rate) glyphosate application rates.

F. arundinacea 'Falcon' and B. dactyloides 'Bowie' were selected as the most resistant and sensitive species to applied sublethal rates of glyphosate as determined with visual injury ratings, respectively. The chlorophyll content and dry matter production in leaves of turfgrass species decreased with increasing rates of glyphosate application. The control (untreated) plants of all species had very small amounts of shikimate (less than 0,03 umol/g fresh weight). In contrast, shikimic acid accumulated in leaves of selected turfgrass species as early as days after treatment (DAT1) for all glyphosate rates applied. Even the lowest rate of glyphosate (5 %) caused a significant increase in the concentration of shikimate in the treated plants. The concentration of shikimic acid increased markedly in the glyphosate treated plants with increasing rate of glyphosate.

There were species-specific differences in accumulation rate of shikimate and F. arundinacea 'Falcon' species which was determined as the most resistant one to sublethal rates by visual injury ratings, was detected to accumulate considerable amount of shikimate. Evaluation of the levels of shikimate accumulated in the glyphosate treated turfgrass plants could be helpful in understanding the extent of inhibition of EPSPS enzyme by glyphosate, however, the visual injury ratings and chlorophyll content determination also provide valuable information.

The specific objective of this research was also to characterise the variation in response of selected turfgrass populations to glyphosate at molecular level. EPSPS gene was isolated and characterised only from buffalograss using degenerate primers. However, expression level of this gene was very low based on RT-PCR and QRT-PCR study.

The main achievements of the project included one postdoctoral fellow and two graduate students were trained and two articles were published due to this funding and two more publications were in preparation.