Triacylglycerol (TAG) is a class of lipid molecules composed of three fatty acyl chains esterified to a glycerol backbone. In plants, TAG is synthesized in various tissues and serves as a carbon and energy source. Oil biosynthesis is well understood in oilseeds however how plants store oil in non-seed tissue is yet to be determined. In Avocado (Persea americana), a basal angiosperm, TAG is exclusively accumulated in mesocarp tissue and therefore is emerging as a model system to uncover underlying mechanisms of TAG biosynthesis in tissues other than seed. The mesocarp of Avocado fruit contains ~60-70% of oil by dry weight. Recent transcriptome studies revealed that the TAG biosynthesis is transcriptionally regulated in non-seed tissues. In seed tissues, TAG biosynthesis is regulated by many seed maturation factors directly or indirectly through downstream transcription factor WRINKLED1 (WRI1). Transcriptome studies revealed that in addition to ortholog of WRI1, orthologs for WRI2 and WRI3 were also highly expressed in avocado mesocarp during the period of oil accumulation. Based on the transcriptome data, I hypothesize that putative WRI genes (WRI1, 2, 3) of avocado enhance oil content in nonseed tissues. Currently, cloning of Putative PaWRI 1, 2 and 3 genes into a binary vector, followed by agrobacterium-mediated transformation to generate transient and stable transient lines, is underway. Full-length cDNA for PaWRI genes (1 & 2) were amplified and cloned into pK34 entry vector followed by sequence confirmation. PaWRI genes (1 & 2) were subcloned into pB110 destination vector and will be transformed into agrobacterium for their integration into the plants. Cloning of WRI3 is still ongoing. Transient expression of putative PaWRI 1, 2 and 3 genes, will be validated using tobacco leaf assay, are expected to enhance oil accumulation in leaf tissues. Agrobacterium bearing PaWRI genes and a viral silencing protein (p19) will be co-infiltrated on to the underside of Nicotiana benthamiana leaves. Infiltrated plants will be placed in growth room with 16:8 light/dark cycle. Four days post infiltration, infected leaf areas will be harvested and TAG content and composition will be determined by gas chromatography coupled with flame ionization detector. Functional validation of these orthologs is expected to reveal the preferred WRI isoform that likely participates in regulation of oil biosynthesis in avocado mesocarp. Additionally, this work may also elucidate the differences between regulation of TAG accumulation in seed and non-seed tissues and identify new targets to enhance TAG biosynthesis in plants.

The avocado mesocarp contains up to 60-70% oil by dry weight where triacylglycerol (TAG) is the major constituent. This neutral lipid, TAG is utilized by plants for the carbon and energy source when stores in seed tissue. There is significant human nutritional demand for vegetable oil, but its use in production of renewable biomaterials and fuels has intensified the need to increase oil production. In plants, the final and committed step in TAG biosynthesis is catalyzed by diacylglycerol acyltransferases (DGAT) and/or a phospholipid: diacylglycerol acyltransferases (PDAT). Both DGAT and PDAT contribute to seed TAG biosynthesis in an independent or overlapping manner, depending on the species. However, the regulation of TAG biosynthesis is not well-studied in nonseed tissues such as mesocarp of avocado. Based on the transcriptome data of Persea americana it is hypothesized that both DGAT and PDAT are likely to catalyze the conversion of diacylglycerol to TAG. In this study, putative DGAT1 and DGAT2 were identified and comprehensive in silico analyses were conducted to determine the respective start codons, full-length coding sequences, transmembrane domains, predicted protein structures and phylogenetic relationships with other known DGATs. These data reveal that the putative DGATs of a basal angiosperm species retain features that are conserved not only among angiosperms but also other eukaryotes. For further functional analysis, the avocado DGATs were expressed in H1246, a TAG-deficient yeast strain and lipotoxicity rescue assays were conducted. The complementation of this yeast strain confirmed enzyme activity and supported the possible role of both avocado DGATs in TAG biosynthesis. Future studies will be focused on determining the substrate specificity of DGAT and its role, relative to PDATs in TAG biosynthesis in avocado mesocarp.

Triacylglycerol (TAG) is a class of lipid molecules composed of three fatty acyl chains esterified to a glycerol backbone. In plants, TAG is synthesized in various tissues and serves as a carbon and energy source. Oil biosynthesis is well understood in oilseeds however how plants store oil in non-seed tissue is yet to be determined. In Avocado (Persea americana), a basal angiosperm, TAG is exclusively accumulated in mesocarp tissue and therefore is emerging as a model system to uncover underlying mechanisms of TAG biosynthesis in tissues other than seed. The mesocarp of Avocado fruit contains ~60-70% of oil by dry weight. Recent transcriptome studies revealed that the TAG biosynthesis is transcriptionally regulated in non-seed tissues. In seed tissues, TAG biosynthesis is regulated by many seed maturation factors directly or indirectly through downstream transcription factor WRINKLED1 (WRI1). Transcriptome studies revealed that in addition to ortholog of WRI1, orthologs for WRI2 and WRI3 were also highly expressed in avocado mesocarp during the period of oil accumulation. Currently, cloning of Putative PaWRI 1, 2 and 3 genes into a binary vector, followed by agrobacterium-mediated transformation to generate transient and stable transient lines, is underway. Transient expression of putative PaWRI 1, 2 and 3 genes, using tobacco leaf assay, are expected to enhance oil accumulation in leaf tissues. Stable expression of PaWRI 1, 2, and 3 in Atwri-/- is expected to restore oil accumulation in seeds. TAG content and composition will be determined by gas chromatography coupled with flame ionization detector. Functional validation of these orthologs is expected to reveal the preferred WRI isoform that likely participates in regulation of oil biosynthesis in avocado mesocarp. Additionally, this work may also elucidate the differences between regulation of TAG accumulation in seed and non-seed tissues and identify new targets to enhance TAG biosynthesis in plants.

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A variety of plants are natural resources for oil and are capable of synthesizing and storing up to 90% oil (dry weight) in the form of triacylglycerols (TAGs). TAGs are commonly used as vegetable oils of which, >35% is derived from fleshy part of the fruits, such as mesocarp of oil palm, avocado, and olive. Studies on TAG synthesis in seed tissues mostly implicated an acyl CoA-dependent enzyme, diacylglycerol (DAG) acyltransferase (DGAT) to catalyze the conversion of DAG to TAG. However, recent studies on Arabidopsis and oil palm suggested participation of a phospholipid:diacylglycerol acyltransferase (PDAT), which is an acyl-CoA-independent enzyme. In avocados, which store up to 70% oil in mesocarp, I hypothesize that both DGAT and PDAT are likely involved in TAG synthesis. To test the hypothesis, I determined TAG content and composition by gas chromatography (GC) and expression levels of DGAT and PDAT genes by real-time PCR, in developing mesocarp. These data will be compared to that of seed tissues of avocado to associate gene expression levels with changes in oil accumulation. Future studies on cloning and characterization of these potential acyltransferase genes involved in TAG synthesis will allow us to develop genetic tools that may increase oil yield; a step towards meeting the consumption demand for oil that is expected to almost double by 2030.

El texto completo de este trabajo no está disponible en el Repositorio Académico UPC por restricciones de la casa editorial donde ha sido publicado. / Avocado exports have increased considerably between 2016 and 2019 in Peru. For this reason, it is important that farmers develop strategies in their supply chain that will generate dwell value for their final product. In response to the above, Supply Chain Management Leagile (SCM Leagile) surface. This supply chain strategy Reduce processes that do not generate value to the product and have to rapid response to the constant change in demand for avocado to the international market. The objective of this paper is present a model of SCM Leagile which can be used by the farmers in various agrifood supply chains.

The study is aimed to determine the role of avocado DGAT1 and PDAT1 in seed oil synthesis. Triacylglycerol (TAG) has a nutritional and industrial value and is essential for plant growth. DGAT1 and PDAT1 catalyze the final step of TAG Assembly. We hypothesized that both PaPDAT1 and PaDGAT1, although predominantly expressed in non-seed tissues, could contribute to oil accumulation in seeds. Agrobacterium transformants with PaPDAT1 and PaDGAT1 cloned in pCAMBIA were generated to test this. Subsequently, the Agrobacterium-mediated transformation of Arabidopsis mutant lines and camelina was carried out by floral dipping. The T1 camelina seeds expressing the genes of interest were selected using fluorescence screening. Homozygous T3 lines were generated. The transgenic camelina seeds were evaluated for TAG content and fatty acid composition relative to wild-type seeds. Line D1 3-3-2 expressing PaDGAT1 and line P1 7-8 expressing PaPDAT1 showed a significant increase in C18:1 compared to the wild type.

Premature and excessive leaf abscission during flowering time in the late avocado (Persea americana Mill.) cultivar ‘Ryan’ is a considerable problem for avocado growers. They are especially concerned that premature and excessive leaf abscission will have a negative effect on yield. No previous investigations have been performed where premature and excessive leaf abscission in avocado has been studied in detail. This study therefore aimed to investigate the pattern of premature and excessive leaf abscission in ‘Ryan’, and compare it with two other important commercial cultivars, ‘Fuerte’ and ‘Hass’, which do not display this phenomenon. Time course studies of leaf abscission in the orchard were performed during 2006 and 2007 to determine the pattern of leaf abscission on ‘Ryan’, Fuerte’ and ‘Hass’. This also included anatomical studies to determine the time of leaf abscission zone formation. Possible stress factors, which accelerate leaf abscission were also investigated, namely unfavourable climatic conditions (temperature, solar radiation, rainfall, relative humidity and evapotranspiration), nutrient imbalances, excessive flowering and leaf area. The possible impact leaf abscission may have on yield was then assessed by determining levels of reserve carbohydrates in the bark of the tree. In addition, practical solutions, i.e. the application of fertilizers, plant growth regulators (PGRs) and kaolin, were investigated in order to reduce or eliminate premature and excessive leaf abscission. This study was carried out over the period 2005 until 2007, with experiments being modified on an annual basis as information was gathered on the phenomenon. Experiments began in 2005 with a study on the pattern of leaf abscission in ‘Ryan’, which revealed an increase in leaf abscission just prior to flowering. However, this increase was not significant. During 2006, the leaf abscission pattern for ‘Ryan’ was compared with the leaf abscission patterns of ‘Fuerte’ and ‘Hass’. Leaf abscission for ‘Ryan’ was significantly higher than for ‘Fuerte’ and ‘Hass’ during 2006. During 2006 ‘Ryan’ displayed two periods of high leaf abscission, namely the spring flush between bud dormancy and bud swell, and a drastic increase in spring and summer flush leaf abscission between inflorescence development and full bloom. These periods of increased leaf abscission were absent during the 2007 season. In addition, ‘Fuerte’ and ‘Hass’ did not display these peaks of high leaf abscission, with leaf abscission occurring in these cultivars at higher rates from full bloom onwards. Premature and excessive leaf abscission is therefore not an annual event in ‘Ryan’ and is in all likelihood influenced by external factors. Anatomical studies did not reveal any results in terms of initiation of leaf abscission, with only the protective layer of the abscission zone being visible after leaf yellowing occurred. During 2006, two peaks of extremely low temperatures (<4°C) occurred just prior to the acceleration of leaf abscission. During the second period of low temperatures, the solar radiation:temperature-ratio was also considerably higher. These periods of low temperatures were absent during 2007, indicating that cold and light stress could be contributing to premature and excessive leaf abscission in ‘Ryan’ in 2006. In addition, ‘Ryan’ flowered excessively during 2006, which could have been triggered by low temperature stress just prior to flower initiation. A significant correlation was found between excessive flowering and excessive leaf abscission in ‘Ryan’ during 2006. The occurrence of reduced flowering in ‘Fuerte’ and ‘Hass’ may possibly be due to these two cultivars being more tolerant to stress, and it is possible that ‘Ryan’ is genetically more prone to excessive flowering than ‘Fuerte’ and ‘Hass’. Excessive flowering could accelerate leaf abscission by causing an unusually high demand for water, nutrients and carbohydrates, resulting in the acceleration of leaf abscission. No significant relationship between nutrient levels and excessive leaf abscission was found for either 2006 or 2007. In addition, no significant correlation could be found between leaf abscission on a branch and the total leaf area of that branch during the 2007 season. During 2007, leaf abscission was low and it is possible that a significant correlation could be found in a season with excessive leaf abscission. During 2005, chemical applications to reduce leaf abscission did not yield any significant improvement in leaf retention. In fact, the 50 g/tree Solubor® and 50 g/tree Solubor® in combination with 2 kg/tree dolomitic lime had a significant negative effect on fruit set, possibly because too high concentrations were applied too close to fruit set. Chemical applications during 2006 were therefore made at bud dormancy and bud swell, as it was found that leaf abscission occurred before flowering time. However, no effect was observed on leaf retention or fruit set. During 2007, emphasis was placed on treatments that might reduce stress, as it became evident that stress could be responsible for premature and excessive leaf abscission in ‘Ryan’. Most treatments showed a slight positive effect on leaf retention, but no significant results were obtained possibly because that particular season was a season of low leaf abscission. Further research on application of stress-reducing treatments is therefore recommended. Best farm management practices including optimal fertilization and irrigation is therefore vital to prevent stress, accelerating leaf abscission. Copyright / Dissertation (MSc)--University of Pretoria, 2009. / Plant Science / unrestricted