The industrial uses of cottonseed oil are limited by its fatty acid composition. Genetic modification of cotton lipid profiles using seed-specific promoters could allow cotton growers to produce valuable new oils in the seed without adverse effects on fiber quality and yield, therefore making this crop more commercially profitable. Transgenic cotton callus harboring a diverged fatty acid desaturase gene (FADX) from Momordica charantia was characterized for production of alpha-eleostearic acid (conjugated double bonds: 18:3 D9 cis, 11 trans, 13 trans), not normally found in cotton. Gas chromatography (GC) in conjunction with mass spectrometry (MS) confirmed production of alpha-eleostearic acid in the transgenic cotton tissues. A second series of transformation experiments introduced the cotton fatty acid thioesterase B (FATB) cDNA, fused to the seed-specific oleosin promoter into cotton to promote the over-expression of FATB, to generate cotton with increased palmitate in the cottonseed. PCR amplification, as well as fatty acid analysis by gas chromatography, confirmed introduction of the FATB cDNA in transgenic tissues. Collectively, these results demonstrate the feasibility of manipulating the fatty acid composition in cotton via transgenic approaches and form the basis for continued efforts to create novel oils in cottonseed.

Master of Science

M.S.

Thesis (MScAgric)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: This research study consists of three investigations with regard to ostrich (Struthio camelus var. domesticus) production, meat quality and the processing of ostrich meat into a value added meat product. The first study was conducted in order to establish whether the gradual replacement of soybean oilcake meal with cottonseed oilcake meal (CSOCM) as a protein source in the diet of slaughter ostriches would affect ostrich growth performance and meat quality. A total of 105 ostriches were divided into five feeding groups according to the CSOCM inclusion level: Control (0% CSOCM), 3%, 6%, 9% and 12% CSOCM, and fed with experimental diets from 6 to 13 months of age. As a result of feeding CSOCM, the final live weight and the average daily gain significantly increased in the 12% CSOCM group compared to the other treatment diets. The proximate composition, cholesterol content, mineral and fatty acid profile of the meat remained unaffected. Considering all the results, CSOCM may be used as an alternative protein source to soybean oilcake meal in ostrich nutrition, resulting in decreased feed costs. Secondly, a descriptive sensory analysis, together with chemical and physical measurements, was performed to determine whether the manipulation of the fatty acid composition in the fan fillet (Iliofibularis muscle) as a result of feeding CSOCM would be detected on a sensory level. Two levels of CSOCM were investigated; 0% as a control and 9% CSOCM. No significant differences were found for the physical measurements (cooking loss (%) and shear force) as well as for the pH and proximate composition of the raw fan fillet. The Control group presented a higher (P<0.05) mono-unsaturated fatty acid (MUFA) content in the cooked fan fillet whereas the 9% CSOCM group showed a favourable increased (P<0.05) poly-unsaturated fatty acid (PUFA) content when compared to the cooked Control samples. As a result, the poly-unsaturated:saturated fatty acid (PUFA:SFA) ratio in the 9% CSOCM group was also higher (P<0.05). No differences (P>0.05) were found between the treatments for the n-6:n-3 (omega 6 to omega 3) ratio. The 9% CSOCM group had a more intense beef aroma, had a higher level of initial and sustained juiciness as well as increased tenderness (P<0.05). Inclusion of 9% CSOCM resulted in a favourable cooked ostrich fan fillet. Finally, the effect of feeding CSOCM on a processed ostrich meat product was investigated. Fan fillet (Iliofibularis muscle) from 13 month old birds receiving no cottonseed oilcake meal (Control) or 9% cottonseed oilcake meal (9% CSOCM) was used. Olive oil was used as a replacement for pork fat, and warthog (Phacochoerus africanus) meat was used to replace commercial pork meat in the production of a semi dry sausage, cabanossi. Olive oil was included at three levels (0%, 1% and 2%). Six treatments were investigated: Control 0% olive oil, Control 1% olive oil, Control 2% olive oil, 9% CSOCM 1% olive oil, 9% CSOCM 2% olive oil en 9% CSOCM 2% olive oil. The Control and 9% CSOCM ostrich meat did not differ significantly in chemical composition nor fatty acid profile. After smoking and drying the fat content in the cabanossi containing 0%, 1% and 2% olive oil averaged 7.2%, 7.45% and 8.65% respectively. Processed meat products containing less than 10% fat are classified as a low-fat meat product. Olive oil is a mono-unsaturated vegetable oil containing mainly Oleic acid (C18:1n9c), and low quantities of saturated fatty acids and polyunsaturated fatty acids. Total mono unsaturated fatty acids in the cabanossi increased from 47.0% to 73.0% of total fat, whilst total saturated fatty acids and total polyunsaturated fatty acids decreased from 40.6% to 19.9% and 11.6% to 6.6% respectively as olive oil increased from 0% to 2%. The inclusion of olive oil at 2% resulted in cabanossi with increased (P<0.05) tenderness, juiciness and cured red meat colour, all factors that appeal greatly to the consumer. Overall flavour was not adversely affected by the inclusion of olive oil. This investigation indicated that the use of CSOCM had no negative effect on the production performance of ostriches whilst a 9% CSOCM inclusion level resulted in meat that was found to be favourable by a trained sensory panel. Furthermore, the use of CSOCM as a feed component also had no negative effect on a processed product (cabanossi) derived from the meat obtained from the birds fed this feed component. The CSOCM used in this investigation had low levels of gossypol (10 to 20ppm) and more research is required on the effect of the use of CSOCM with higher levels of gossypol on the production performance of ostriches. / AFRIKAANSE OPSOMMING: Die studie het bestaan uit drie ondersoeke met betrekking tot volstruis (Struthio camelus var. domesticus) -produksie, -vleiskwaliteit en die vervaardiging van waarde-toegevoegde geprosesseerde volstruis-vleisprodukte. Die doel van die eerste studie was om vas te stel of die geleidelike vervanging van sojaboonoliekoekmeel met katoensaad-oliekoekmeel (CSOCM) as ‘n proteïenbron in die voeding van volstruise, die groeipersentasie en vleiskwaliteit van die Iliofibiularis spier (fan fillet) sal affekteer. ‘n Totaal van 105 volstruise is verdeel in vyf voedingsgroepe volgens die katoensaad oliekoekmeel insluitingsvlak: Kontrole (0% CSOCM), 3%, 6%, 9% en 12% CSOCM. Die onderskeie voedingsgroepe was van ses tot 13 maande ouderdom op die eksperimentele voere geplaas. Die resultate het aangedui dat die voëls in die 12% CSOCM behandelingsgroep ‘n betekenisvolle (P<0.05) toename in finale lewende massa asook gemiddelde daaglikse toename gehad het. Die proksimale samestelling, cholesterol-inhoud, mineraal- en vetsuursamestelling van die vleis was nie geaffekteer deur die insluiting van CSOCM nie. Die CSOCM kan dus wel as ‘n alternatiewe proteïenbron in die voeding van volstruise gebruik word. Laasgenoemde bevinding kan ook lei tot verlaagde voerkostes, aangesien CSOCM heelwat goedkoper is as sojaboon-oliekoekmeel. Die tweede deel van die studie was van ‘n chemiese asook sensoriese aard. ‘n Beskrywende sensoriese analiese is uitgevoer om vas te stel of die manipulering van die vetsuursamestelling in die volstruis fan fillet as gevolg van die CSOCM sensories waargeneem kan word. Die chemiese en fisiese eienskappe van die vleis is ook ondersoek. Twee vlakke van CSOCM inhoud is ondersoek; 0% (as kontrole) en 9% CSOCM. Geen betekenisvolle verskille is gevind vir die fisiese vleiskwaliteit (kookverliespersentasie en taaiheid), asook vir die proksimale samestelling en pH van die fan fillet nie. Die gekookte fan fillet van die Kontrole behandeling het ‘n betekenisvolle (P<0.05) toename in mono-onversadigde vetsure (MUFA) getoon en die 9% CSOCM het ‘n voordelige toename in poli-onversadigde vetsuur-inhoud (PUFA) gehad. Die poli-onversadigde tot versadigde vetsuurverhouding (PUFA:SFA) was as ‘n gevolg ook betekenisvol hoër. Geen verskille (P>0.05) is opgemerk in die omega-6 tot omega-3 poli-onversadigde vetsuurverhouding (n-6:n-3) nie. Met betrekking tot die sensoriese eienskappe het die 9% CSOCM ‘n meer opvallende beesvleis aroma, hoër vlakke van aanvanklike sappigheid en ook sagter vleis in vergelyking met die kontrole behandeling gehad (P<0.05). Insluiting van 9% CSOCM het gelei tot ‘n gekookte volstruis fan fillet van voornemende kwaliteit. Laastens is daar ondersoek ingestel op die vervanging van varkvet met olyfolie in die vervaardiging van ‘n volstruis cabanossi. Chemiese asook sensoriese analises is uitgevoer op die gedroogde en gerookte volstruis cabanossi. Vir die vervaardiging van laasgenoemde produkte is die fan fillet van 13 maande oue voëls van die Kontrole (0% CSOCM) en 9% CSOCM behandelings gebruik. Addisioneel tot die volstruisvleis is daar ook vlakvarkvleis (Phacochoerus africanus) gebruik om die kommersiële varkvleis te vervang. Olyfolie was ingesluit teen drie vlakke (0%, 1% en 2% van die totale mengsel). Ses behandelings was ondersoek: Kontrole 0% olyfolie, Kontrole 1% olyfolie, Kontrole 2% olyfolie, 9% CSOCM 1% olyfolie, 9% CSOCM 2% olyfolie en 9% CSOCM 2% olyfolie. Daar was geen verskille (P>0.05) in die chemiese en vetsuursamestellings van die Kontrole en 9% CSOCM volstruisvleis nie. Na die droging en rooksiklus was die gemiddelde vet-inhoud van die 0%, 1% en 2% olyfolie cabanossi monsters onderskeidelik 7.2%, 7.45% en 8.65%. Geprosesseerde vleisprodukte met ‘n vet-inhoud van minder as 10% word in die kommersiële vleisindustrie na lae vet vleisprodukte verwys. Olyfolie is baie ryk aan MUFA, veral Oleïensuur (C18:1n9c) en dit bevat ook lae hoeveelhede SFA en PUFA. Die totale MUFA inhoud in die cabanossi het toegeneem van 47.0% tot 73.0% terwyl die totale SFA en PUFA onderskeidelik afgeneem het van 40.6% tot 19.9% en 11.6% tot 6.6%, met ‘n olyfolie toename van 0% tot 2%. Die insluiting van olyfolie teen 2% het gelei tot ‘n sagter cabanossi wat meer sappig was met ‘n meer opvallende rooi gekuurde vleiskleur, wat almal eienskappe is wat dié produk meer aantreklik maak vir die verbruiker. Hierdie studie het aangedui dat CSOCM geen negatiewe effek gehad het op die produksie van volstruise nie. Volstruisvleis van die behandelingsgroep wat CSOCM teen 9% van die dieet ontvang het, het wel vleis geproduseer wat as aanvaarbaar aanskou was deur ‘n opgeleide sensoriese paneel. Die gebruik van CSOCM as ‘n voerbestandeel het ook geen negatiewe effek gehad op ‘n geprosesseerde produk (cabanossi) gemaak van die volstruisvleis nie. Die CSOCM wat in die huidige studie gebruik is, het baie lae vlakke van gossypol (10 – 20dpm) gehad en verdere ondersoek is noodsaaklik om die effek van CSOCM met hoër vlakke van gossypol op die produksie van volstruise te bevestig.

This item was digitized as part of the Million Books Project led by Carnegie Mellon University and supported by grants from the National Science Foundation (NSF). Cornell University coordinated the participation of land-grant and agricultural libraries in providing historical agricultural information for the digitization project; the University of Arizona Libraries, the College of Agriculture and Life Sciences, and the Office of Arid Lands Studies collaborated in the selection and provision of material for the digitization project.

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The relatively high level of palmitic acid (22 mol%) in cottonseeds may be due in part to the activity of a palmitoyl-acyl carrier protein (ACP) thioesterase (PATE). In embryo extracts, PATE activity was highest at the maximum rate of reserve accumulation (oil and protein). The cotton FatB mRNA transcript abundance also peaked during this developmental stage, paralleling the profiles of PATE enzyme activity and seed oil accumulation. A cotton FatB cDNA clone was isolated by screening a cDNA library with a heterologous Arabidopsis FatB probe (Pirtle et al., 1999, Plant and Cell Physiology 40: 155-163). The predicted amino acid sequence of the cotton PATE preprotein had 63% identity to the Arabidopsis FatB thioesterase sequence, suggesting that the cotton cDNA clone probably encoded a FatB-type thioesterase. When acyl-CoA synthetase-minus E. coli mutants expressed the cotton cDNA, an increase in 16:0 free fatty acid content was measured in the culture medium. In addition, acyl-ACP thioesterase activity assays in E. coli lysates revealed that there was a preference for palmitoyl-ACP over oleoyl-ACP in vitro, indicating that the cotton putative FatB cDNA encoded a functional thioesterase with a preference for saturated acyl-ACPs over unsaturated acyl-ACPs (FatA). Overexpression of the FatB cDNA in transgenic cotton resulted in elevated levels of palmitic acid in transgenic somatic embryos compared to control embryos. Expression of the anti-sense FatB cDNA in transgenic cotton plants produced some plants with a dwarf phenotype. These plants had significantly smaller mature leaves, all with smaller cells, suggesting that these plants may have less palmitic acid available for incorporation into extraplastidial membrane lipids during cell expansion. Thus manipulation of FatB expression in cotton directly influenced palmitic acid levels. Collectively, data presented in this dissertation support the hypothesis that there indeed is a palmitoyl-ACP thioesterase in cotton, encoded by the isolated FatB cDNA, which plays a major role in regulating palmitic acid content of extraplastidial complex glycerolipids. This work forms the basis for future studies of the influence of palmitic acid content on plant membrane function and provides a key target for the metabolic engineering of palmitic acid levels in storage oils of developing cottonseeds.

The aim of this study was to investigate the effect of feeding cotton co-products with different gossypol concentrations on reproductive development of lambs close to puberty and also on carcass traits and fatty acid profile of meat. Twenty four 5-months old ram lambs (20.6 ± 1.9 kg BW) were used. These were housed in individual pens and received four diets: 20% of dry matter intake (DMI) of whole cottonseed (WCS), 20%DMI of cottonseed meal (CSM), 20%DMI of high oil cottonseed meal (CSC) and a control group without cottonseed co-products (CTL). Free gossypol intake was 0, 16.32, 6.98 and 5.47 mg/kgBW for CTL, WCS, CSC and CSM, respectively. At each 15 days, the animals were weighted, and blood and semen samples were collected. Sperm motility, vigor, mass movement, concentration and pathologies were evaluated. The free testosterone and cortisol concentrations in serum were determined. After 95 experimental days, the lambs were slaughtered and carcass traits were measured. Meat samples of Longissumus dorsi muscle were taken for fatty acid profile analysis. And testis samples were collected to analysis in light and transmission electron microscopes. The treatments did not differ in average daily weight gain, sperm volume, motility, vigor and concentration. The CTL group had higher testosterone concentration than CSC at the end of trial and had lower total sperm defects and higher mass movement than others. The number of mitochondrial sheath aplasia increased with increasing gossypol level in diet. There was no relation between the other variables evaluated and gossypol level in diets; however the groups that received cottonseed co-products showed worse reproductive parameters than CTL. Therefore, these co-products had negative impact on reproductive system of puberal lambs. In relation to fatty acid profile, meat from CSM and CSC groups had higher levels of conjugated linolenic acid (CLA) than others and yet CSC group showed higher vaccenic acid than others. Meat from animals that received whole cottonseed had less unsaturated fatty acids, CLA and vaccenic acid. Therefore, between cotton co-products, the processed (CSM and CSC) must be preferred for use in ruminant feed rather than whole cottonseed. The meat from animals that did not receive cotton co-products had higher n-3 fatty acid, and also better n-6 to n-3 ratio compared to others. This can impair the use of these co-products due to current great importance given to these fatty acids in human nutrition / Com este estudo, objetivou-se avaliar o efeito da alimentação com co-produtos do algodão tendo diferentes concentrações de gossipol no desenvolvimento reprodutivo de cordeiros no período próximo a puberdade e também nas características de carcaça e no perfil de ácidos graxos da carne. Vinte e quatro cordeiros com 5 meses de idade (20.6 ± 1.9 kg PV) foram utilizados. Estes foram alojados em baias individuais e receberam quatro dietas: 20% da ingestão de matéria seca (IMS) de caroço de algodão (CAROÇO), 20%IMS de farelo de algodão (FARELO), 20%IMS de torta de algodão (TORTA) e um grupo controle (CONTROLE) sem o uso de co-produtos do algodão. A ingestão de gossipol livre foi de 0, 16,32, 6,98 e 5,47 mg/kgPV para CONTROLE, CAROÇO, TORTA e FARELO, respectivamente. A cada 15 dias, os animais foram pesados, e amostras de sangue e sêmen foram coletadas. O sêmen foi analisado quanto a motilidade, vigor, turbilhão, concentração e patologias. Foram determinadas a concentração de testosterona livre e cortisol no soro sanguíneo. Após 95 dias de experimento, os cordeiros foram abatidos e as características de rendimento de carcaça e composição da 12ª costela foram mensuradas. Amostras do músculo Longissumus dorsi foram coletadas para a análise do perfil de ácidos graxos. Amostras dos testículos foram coletadas para análise em microscopia de luz e microscopia eletrônica de transmissão. Os tratamentos não diferiram em ganho em peso diário, volume ejaculado, motilidade, vigor e concentração espermática. Os animais do grupo CONTROLE apresentaram concentração de testosterona maior que os do grupo TORTA no final do período experimental. E ainda os animais do grupo CONTROLE tiveram menor número de defeitos totais e maior turbilhão que os outros. O número de lesões de aplasia da bainha mitocondrial aumentou com o aumento do teor de gossipol livre na dieta. Para as outras variáveis avaliadas, não houve relação com o nível de gossipol livre na dieta, no entanto os animais que receberam co-produtos do algodão apresentaram piores parâmetros reprodutivos do que os animais do grupo CONTROLE. Por isso, pode-se concluir que estes co-produtos tiveram um impacto negativo no sistema reprodutivo dos cordeiros durante a puberdade. Em relação ao perfil de ácidos graxos, a carne dos animais dos grupos TORTA e FARELO apresentaram maiores valores de ácido linolênico conjugado (CLA) que os outros dois tratamentos e ainda os animais do grupo TORTA apresentaram maior teor de ácido vacênico que os outros. A carne dos animais que receberam caroço de algodão mostrou menor quantidade de ácidos graxos insaturados, CLA e ácido vacênico. Portanto, dentre os co-produtos do algodão, os processados (FARELO e TORTA) devem ser preferidos para uso na nutrição de ruminantes em detrimento ao caroço. A carne dos animais do grupo CONTROLE apresentou maior teor de n-3, e ainda melhor proporção entre n-6 e n-3 comparado com as demais dietas, o que pode dificultar o uso desses co-produtos devido a grande importância dada atualmente a este grupo de ácidos graxos na nutrição humana