Since the dawn of agriculture man has been genetically modifying crop plants to increase yield, quality and utility. In addition to selective breeding and hybridization we can utilize mutant populations and biotechnology to have greater control over crop plant modification than ever before. Increasing the production of plant oils such as soybean oil as a renewable resource for food and fuel is valuable. Successful breeding for higher oil levels in soybean, however, usually results in reduced protein, a second valuable seed component. We show that by manipulating a highly active acyl-CoA: diacylglycerol acyltransferase (DGAT) the hydrocarbon flux to oil in oilseeds can be increased without reducing the protein component. Compared to other plant DGATs, a DGAT from Vernonia galamensis (VgDGAT1A) produces much higher oil synthesis and accumulation activity in yeast, insect cells and soybean. Soybean lines expressing VgDGAT1A show a 4% increase in oil content without reductions in seed protein contents or yield per unit land area. Furthermore, we have screened a soybean fast neutrino population derived from M92-220 variety and found three high oil mutants that do not have reduced levels of protein. From the F2 plant populations we quantitatively pooled the high oil and low oil plants and performed comparative genomics hybridization (CGH). From the data it appears that two families have a 0.3 kb aberration in chromosome 14. We are performing further analysis to study this aberration and develop markers for molecular breeding. Mutagenic techniques are also useful for developing other traits such as early flowering varieties and adapting new high oil crops to a new region. Chia (Salvia hispanica) is an ancient crop that has experienced an agricultural resurgence in recent decades due to the high omega 3 fatty acid (ω-3) content of the seeds and good production potential. The area of cultivation has been expanded to Kentucky using mutagenized populations and the composition traits are similar to that of the original regions of cultivation in Central and South America.

New soybean lines have been developed with significantly higher oil, protein + oil and higher meal protein. These soybeans contain a VgD1 gene (highly active acyl-CoA:diacylglycerol acyltransferase, DGAT from Vernonia galamensis (VgDGAT1A) produces much higher oil synthesis and accumulation activity in soybean. Soybean with active DGAT from Vernonia galamensis (VgDGAT1A) has active TAG biosynthesis relative to other DGATs including from soybeans and Arabidopsis. DGATs catalyze the final step of TAG synthesis: DAG (diacylglycerol) + acyl-CoA → TAG + CoASH (Coenzyme A is notable for its role in the synthesis and oxidation of fatty acids, and the oxidation of pyruvate in the citric acid cycle). A thorough analysis of the major components in VgD1 lines, especially those of nutritional or anti-nutritional value including what else changed (decreased); and what remained at normal levels was conducted. A field study was conducted in Spindletop and Princeton KY, reviled no reduction in yield nor protein, and about 4 % (DW) more oil was obtained in Princeton and 2% (DW) in Spindeltop. No consistent reduction in the other seed composition.VgDGAT1A soybean lines indicated noticeably early maturation compared to the parental line. This is associated with higher expression of the flowering genes FT2 (FLOWERING LOCUS T2) and FT5 (FLOWERING LOCUS T5), for the high oil lines. A single recessive mutation in soybean (MIPS) myo-inositol 1-phosphate synthase, confers a seed phenotype of increase inorganic phosphate (Pi) crossed with high oil lines expressing a DGAT from Vernonia galamensis (VgDGAT1A) (VgD). The oil and protein were maintained compart to VgD. VgD X MIPS (VM), had 21.2, and 22 % oil in 2015, and 23.3 and 24.0 oil in 2016, and protein 46, 49 in 2015, and 37 and 39 % in 2016. Phosphate results suggesting the cross MV is still segregating for MIPS and more selection and planting are needed. Measurement of seed phosphate levels is an established technique for screening for low phytate mutants but to date, it has not been performed non-destructively from single soybean seeds. A protocol was developed greatly reducing the sample size thereby reducing the cost and time and saving a generation in the selection of low phytate mutant seeds based on the high Pi phenotype. Genotyping single seeds are useful in breeding and genetics while maintaining high germination rates. Nondestructive single-seed genomic DNA extraction protocols using 12 mg cotyledon tissue with a modified cetyl trimethyl ammonium bromide (CTAB) technique and a commercial seed DNA extraction kit using 1 mg cotyledon tissue were developed for dry soybean seeds and cross-verified with leaf DNA analysis.

Fructose consumption has increased exponentially during the past four decades. The physiological effects of a high fructose diet include obesity and insulin resistance. In animal models, the effects of a high fructose diet on fat distribution are inconclusive in that some studies find increases in body mass and lipids while others find no effect. Recent findings indicate that a high fructose diet causes hippocampal insulin resistance in hamsters, raising the possibility that the diet causes impairments in cognition. The following experiments tested the hypotheses that a high fructose diet alters fat distribution rather than total body mass and impairs hippocampal-dependent memory. Results indicated that the high fructose diet did not affect fat distribution, but did increase plasma triacylglycerides. Interestingly, the diet also impaired spatial reference memory in the Morris water maze, and this effect was correlated with plasma triacylglycerides. These results indicate that a high fructose diet impairs brain function.

Concrete in its unaltered form allows the mass transfer of fluids into and out of its microstructure. These fluids can contain detrimental solutes which change the chemistry of the cement paste and/or the corrosion properties of the reinforcement bars, most noticeably hydrogen carbonates (HCO3-), oxygen (O2) and chloride ions (Cl-). Water and its solutions containing salts, mostly sodium chloride (NaCl), can also cause physical damage due to phase changes (freezing and thawing).  External application of hydrophobic agents onto the cement paste surface is a well-known method to alter the mass transfer at this interface. Bulk application of hydrophobic agents in ready mixed concrete is also a possible route but alters the entire cement paste. This thesis presents relevant aspects concerning the use of bulk hydrophobic agents in concrete within a spectrum water to cement ratio   (w/c) = 0.40-0.50. The main focus was on triacylglycerides (TAG) and alkyl alkoxysilanes (“silanes”) with application rates 1-3% based on cement weight.  Alterations to the compressive strengths have been observed and documented over a three-year period. The relative drop in mechanical strength is inversely proportional to w/c. The higher the addition rate, the lower the compressive strength. Chemical differences within the hydrophobic groupings (TAG or “silanes”) resulted in different outcomes. This was most noticeable in the water absorption, compressive strengths and chloride diffusion.  Freeze thaw testing did show noticeable differences, the use of “silanes” was detrimental in these tests even in deionsed water. The exact mechanism is unknown, but thin section analysis shows a lack of air entrainer (even when added on the fresh concrete mix) and extensive cracking in the entire cement paste. The scaling in concrete with TAGs was smaller but needs further improving.  The main properties intended with these agents were the ability to alter the mass transfer of water or solutions into the cement paste. Capillary suction and diffusion were examined. Increasing the w/c reduces the effectiveness of the hydrophobic agents to resist water uptake. This was seen in capillary suction and uni-directional chloride diffusion testing. Processed TAGs were more effective in reducing chloride diffusion than the unprocessed chemical whereas, in some cases, the “silanes” actually increased the amount of chloride ions transferred into the cement paste. Only a slight positive effect can be seen at the lower inclusion rate (1%). Increasing the w/c reduces the resistance to chloride ion diffusion with the same dosage rate.  A field test station close to vehicular traffic was also established in 2018 and 2019, but the specimens have not been tested at this point in time. It is hoped that these and other future studies will lead to a complete PhD project. / Betong i sin oförändrade form tillåter masstransport av vätskor i och ur dess mikrostruktur. Dessa vätskor kan innehålla skadliga lösta ämnen som förändrar cementpastans kemi och/eller korrosionsegenskaperna hos armeringsjärnen, framför allt vätekarbonater (HCO3-), syre (O2) och kloridjoner (Cl-). Vatten och dess lösningar som innehåller salter, mestadels natriumklorider (NaCl), kan till och med orsaka fysisk skada på grund av fasförändringar (frysning och upptining). Extern applicering av hydrofoba medel på cementpastans yta är en välkänd metod för att ändra masstransport genom denna gränsyta. Bulkapplicering av hydrofoba medel i färdigblandad betong är också en möjlig väg, och resulterar i förändringar i hela cementpastan. Denna licentiatavhandling presenterar relevanta aspekter rörande användningen av bulk-hydrofoba medel i betong inom intervall av vattencementtal = 0,40-0,50 (vct). Huvudfokus låg på triacylglyceroler (TAG) och alkyl-alkoxisilaner (”silaner”) med inblandning 1-3 % baserat på cementvikt. Förändringar av tryckhållfasthet har observerats och dokumenterats under en treårsperiod. Den relativa reduktionen i mekanisk hållfasthet är omvänt proportionell mot vct. Ju högre tillsatsmängd i cementpastan desto lägre tryckhållfasthet. Kemiska skillnader inom de hydrofoba grupperna (TAG eller "silaner") resulterade i olika resultat. Detta var mest märkbart i vattenabsorption, tryckhållfasthet och kloriddiffusion. Frysprovning visade märkbara skillnader, användningen av "silaner" var skadlig i dessa tester även i avjoniserat vatten. Den exakta mekanismen är okänd, men tunnslipsanalys visar på brist på luftporbildare (även om den tillsätts i stora mängder i den färska betongblandningen) och omfattande sprickbildning i hela cementpastan. Avskalningen i betong med TAG var mindre men behöver ytterligare förbättras. De huvudsakliga egenskaperna avsedda med dessa medel var förmågan att förändra masstransport av vatten eller lösningar till cementpastan. Kapillärsugning och diffusion undersöktes. Att öka vct minskar effektiviteten hos de hydrofoba medlen för att motstå vattenupptagning. Detta sågs vid kapillärsugning och enkelriktad kloriddiffusionsprovning. Raffinerade TAG:er var effektivare att minska kloriddiffusion än den oraffinerade, medan "silanerna" i visa fall faktiskt ökade mängden kloridjoner som överfördes inne i cementpastan. Något positiva effekt kan ses vid den lägre inkluderingstillsatsen (1 %). Att öka vct minskar motståndet mot kloridjondiffusion. En fältteststation intill fordonstrafik etablerades också 2018 och 2019 men provkropparna har inte testats vid denna tidpunkt. Förhoppningen är att dessa och andra framtida studier ska leda till ett komplett doktorandprojekt. / <p>QC 230330</p>

Made available in DSpace on 2016-06-02T19:32:06Z (GMT). No. of bitstreams: 1 4940.pdf: 1724181 bytes, checksum: edce5d0a5db2ddb6103377e2a01b20fd (MD5) Previous issue date: 2011-08-26 / Universidade Federal de Sao Carlos / The chemical compounds called triacylglycerides (TAGs) have been the main source for biodiesel. The isolation of high lipid yield microalgae is important for the success of algae biomass production for biodiesel and, therefore, a search not only for organisms that fulfill such profile, but also for techniques effective in rapid lipid detection is fundamental. A number of methodologies on the characterization of microalgae lipid production, such as FTIR and RMN spectroscopy and spectrofluorimetry using the Nile Red dye are presented in this study. The Fourier Transform Infrared Spectroscopy (FTIR) technique have been largely used in biological system studies due to its rapid analysis, no pretreatment data gathering and easy biochemical characterization of intact cells. This study proposes a technical optimization of the FTIR technique for microalgae usage purposes as a practical approach for concomitantly prospecting large amounts of microalgae species with focus on biodiesel production. The Red Nile fluorescence has been successfully applied in microalgae lipid determination. In this research, the lipid (neutral) amount was determined by fluorescence in 430-570nm and a linear correlation established with the data gathered through gravimetry. After prospecting a number of species from the laboratory s stock cultures, the freshwater microalgae Scenedesmus sp (Clone MJ13) has been selected for study under traditional growth practices and under conditions of different nitrogen and phosphorus concentrations. The results showed a fast metabolic answer of Scenedesmus sp to the nutritional availability and the efficiency of the FTIR spectroscopy as a fast and reliable method for the determination of lipid production in a prospecting process. / A principal fonte para o biodiesel tem sido os compostos químicos denominados triacilglicerídeos (TAGs). O Isolamento de microalgas com alto rendimento lipídico é a chave para o sucesso da produção de biodiesel a partir de biomassa algal. Desta forma fazse necessário uma busca não somente por organismos que atendam tal exigência, mas também por técnicas eficazes que nos auxiliem na investigação destes. No presente estudo foram apresentadas algumas metodologias úteis na caracterização da produção de lipídeos por microalgas, tais como as espectroscopias de FTIR e RMN e a espectrofluorimetria com o corante Vermelho do Nilo. A técnica de espectroscopia de Infravermelho com Transformada de Fourier (FTIR) vem sendo muito utilizada nos estudos de sistemas biológicos, devido a sua rapidez de análise, obtenção de dados sem tratamento prévio e facilidade na caracterização bioquímica de células intactas. Foi proposto neste trabalho, uma otimização da técnica de FTIR para utilização em microalgas, como um método prático de prospectar concomitantemente uma grande quantidade de espécies de microalgas com finalidade de produção de biodiesel. A fluorescência por vermelho do Nilo tem sido aplicada com sucesso na determinação de lipídeos em microalgas. Desta forma a quantidade de lipídeos (neutros) foi determinada pela fluorescência em 430-570nm, e feita uma correlação linear com os dados obtidos por gravimetria. Após prospecção de várias espécies do banco de culturas do laboratório, foi selecionada a microalga de água doce Scenedesmus sp (Clone MJ13) para um estudo de caso em condições normais de cultivo e sob condições de diferentes concentrações de nitrogênio e fósforo. Os resultados demonstraram uma rápida resposta metabólica da Scenedesmus sp às mudanças na disponibilidade de nutrientes e indicam a eficiência da espectroscopia de FTIR como um método confiável e rápido para a determinação de produção lipídica em um processo de prospecção.