| 1. |
- Aslan, Selcuk, et al.
(författare)
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Increased production of wax esters in transgenic tobacco plants by expression of a fatty acid reductase:wax synthase gene fusion
- 2015
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Ingår i: Transgenic Research. - : Springer Science and Business Media LLC. - 0962-8819 .- 1573-9368. ; 24, s. 945-953
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Tidskriftsartikel (refereegranskat)abstract
- Wax esters are hydrophobic lipids consisting of a fatty acid moiety linked to a fatty alcohol with an ester bond. Plant-derived wax esters are today of particular concern for their potential as cost-effective and sustainable sources of lubricants. However, this aspect is hampered by the fact that the level of wax esters in plants generally is too low to allow commercial exploitation. To investigate whether wax ester biosynthesis can be increased in plants using transgenic approaches, we have here exploited a fusion between two bacterial genes together encoding a single wax ester-forming enzyme, and targeted the resulting protein to chloroplasts in stably transformed tobacco (Nicotiana benthamiana) plants. Compared to wild-type controls, transgenic plants showed both in leaves and stems a significant increase in the total level of wax esters, being eight-fold at the whole plant level. The profiles of fatty acid methyl ester and fatty alcohol in wax esters were related, and C16 and C18 molecules constituted predominant forms. Strong transformants displayed certain developmental aberrations, such as stunted growth and chlorotic leaves and stems. These negative effects were associated with an accumulation of fatty alcohols, suggesting that an adequate balance between formation and esterification of fatty alcohols is crucial for a high wax ester production. The results show that wax ester engineering in transgenic plants is feasible, and suggest that higher yields may become achieved in the near future.
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| 2. |
- Aslan, Selcuk
(författare)
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Metabolic engineering for production of complex lipids in tobacco (Nicotiana benthamiana) leaves and rice (Oryza sativa) endosperm
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- World chemical and energy supplies for industry are today highly dependent on unsustainable fossil reserves. Vegetable lipids with similar chemical structures to fossil oil could be the ultimate renewable solution to replace fossil oil and provide environmentally friendly feedstocks and energy resources. Wax esters and triacylglycerols (TAG) are two groups of lipid with applications in industry as lubricants, surfactants and biodiesel. However, global production of wax esters and TAG is limited to a few crop species such as jojoba, carnauba, oil palm, soybean and rapeseed. Further, there are restrictions on arable land for oilseed crop growth, and the demand for vegetable lipids is predicted to increase in coming decades. Therefore, there is an urgent need to establish sustainable technologies and develop new alternative oil crop species to meet an expected future vegetable lipid demand. This thesis examined the potential for production of wax esters and TAG with different qualities in tobacco (Nicotiana benthamiana) leaves, and in rice (Oryza sativa) endosperm by metabolic engineering. In tobacco leaves, a new metabolic pathway was introduced into chloroplasts by combining bacterial- and plant-derived genes (AtFAR6, AtPES2, MaFAR, MhWS, tpMaFAR::MhWS, AtWRI1) for wax ester biosynthesis. Combinations of the gene functions resulted in production of wax esters with differing composition to a level of 0.9% of leaf dry weight (DW). The newly introduced pathway was further engineered by RNAi inhibition of the KASII gene. Additional co-expression of KASIIRNAi constructs resulted in increased palmitic/stearic acid ratio, which significantly improved wax ester production in AtFAR6-containing combinations. The novel gene fusion tpMaFAR::MhWS was further investigated by stable transformation of tobacco plants. Wax ester content in transformants was increased eight-fold compared with wild-type (WT). In rice, overexpression of both full-length and truncated Arabidopsis transcription factor AtWRI1 in endosperm tissues resulted respectively in sevenfold and four-fold accumulation of TAG content compared with WT. Taken together, these results showed that valuable oleochemicals could be increased considerably in plants.
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| 3. |
- Aslan, Selcuk, et al.
(författare)
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Molecular Genotyping of HistoricalBarley Landraces Reveals Novel CandidateRegions for Local Adaption
- 2015
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Ingår i: Crop science. - : Crop Science Society of America. - 0011-183X .- 1435-0653. ; 55:6, s. 2766-2776
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Tidskriftsartikel (refereegranskat)abstract
- Barley landraces from Northern Europe formgenetically distinct latitudinal groups, suggestingthat adaption plays an important role inthe geographical distribution of genetic diversity.Here, we investigate how Northern Europeanbarley landraces relate to landraces fromother parts of Europe and whether candidategenes for climate adaption can be identified.For this purpose, 27 barley landraces, availableas century-old seed specimens, were genotypedwith a 384 single nucleotide polymorphism(SNP) assay. Landraces from the Nordiccountries formed a genetically distinct grouprelative to landraces from Central and SouthernEurope. Polymorphic positions in the floweringtime genes HvCO1, HvFT1, Ppd-H1, and VRN1-H1 were genotyped. The previously known alleledistribution of Ppd-H1 with the responsive allelepresent in the South and the nonresponsiveallele in the North was confirmed. The otherthree genes were more variable in Central andSouthern Europe compared to the North andneither of the flowering time genes showedany geographically correlated variation withinthe Nordic countries. Allelic frequencies fromthe 384 SNP set were correlated with climaticvariables. This allowed us to identify five SNPsputatively associated with length of growth season,and two SNPs putatively associated withprecipitation. The results show how historicalcrop specimens can be used to study howgenetic variation has been geographically distributedand the genetics of adaption.
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| 4. |
- Aslan, Selcuk, et al.
(författare)
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Transient silencing of the KASII genes is feasible in Nicotiana benthamiana for metabolic engineering of wax ester composition
- 2015
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 5
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Tidskriftsartikel (refereegranskat)abstract
- The beta-ketoacyl-ACP synthase II (KASII) is an enzyme in fatty acid biosynthesis, catalyzing the elongation of 16:0-acyl carrier protein (ACP) to 18:0-ACP in plastids. Mutations in KASII genes in higher plants can lead to lethality, which makes it difficult to utilize the gene for lipid metabolic engineering. We demonstrated previously that transient expression of plastid-directed fatty acyl reductases and wax ester synthases could result in different compositions of wax esters. We hypothesized that changing the ratio between C16 (palmitoyl-compounds) and C18 (stearoyl-compounds) in the plastidic acyl-ACP pool by inhibition of KASII expression would change the yield and composition of wax esters via substrate preference of the introduced enzymes. Here, we report that transient inhibition of KASII expression by three different RNAi constructs in leaves of N. benthamiana results in almost complete inhibition of KASII expression. The transient RNAi approach led to a shift of carbon flux from a pool of C18 fatty acids to C16, which significantly increased wax ester production in AtFAR6-containing combinations. The results demonstrate that transient inhibition of KASII in vegetative tissues of higher plants enables metabolic studies towards industrial production of lipids such as wax esters with specific quality and composition.
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| 5. |
- Aslan, Selcuk, et al.
(författare)
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Wax esters of different compositions produced via engineering of leaf chloroplast metabolism in Nicotiana benthamiana
- 2014
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Ingår i: Metabolic Engineering. - : Elsevier BV. - 1096-7176 .- 1096-7184. ; 25, s. 103-112
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Tidskriftsartikel (refereegranskat)abstract
- In a future bin based economy, renewable sources for lipid compounds at attractive cost are needed for applications where today petrochemical derivatives are dominating. Wax esters and fatty alcohols provide diverse industrial uses, such as in lubricant and surfactant production. In this study, chloroplast metabolism was engineered to divert intermediates from de nova fatty acid biosynthesis to wax ester synthesis. To accomplish this, chloroplast targeted fatty acyl recluctases (EAR) and wax ester synthases (WS) were transiently expressed in Nic"onana benthamiuna loaves. Wax esters of different qualities and quantities were produced providing insights to the properties and interaction of the individual enzymes used. In particular, a phytyl ester synthase was found to be a premium candidate for medium chain wax ester synthesis. Catalytic activities of FAR and WS were also expressed as a fusion protein and determined functionally equivalent to the expression of individual enzymes for wax ester synthesis in chloroplasts. (C) 2014 The Authors. Published by Elsevier Inc. On behalf of International Metabolic Engineering Society.
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| 6. |
- Jin, Yunkai, et al.
(författare)
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Improved bioenergy value of residual rice straw by increased lipid levels from upregulation of fatty acid biosynthesis
- 2023
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Ingår i: Biotechnology for Biofuels and Bioproducts. - 2731-3654. ; 16
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundRice (Oryza sativa) straw is a common waste product that represents a considerable amount of bound energy. This energy can be used for biogas production, but the rate and level of methane produced from rice straw is still low. To investigate the potential for an increased biogas production from rice straw, we have here utilized WRINKLED1 (WRI1), a plant AP2/ERF transcription factor, to increase triacylglycerol (TAG) biosynthesis in rice plants. Two forms of Arabidopsis thaliana WRI1 were evaluated by transient expression and stable transformation of rice plants, and transgenic plants were analyzed both for TAG levels and biogas production from straw.ResultsBoth full-length AtWRI1, and a truncated form lacking the initial 141 amino acids (including the N-terminal AP2 domain), increased fatty acid and TAG levels in vegetative and reproductive tissues of Indica rice. The stimulatory effect of the truncated AtWRI1 was significantly lower than that of the full-length protein, suggesting a role for the deleted AP2 domain in WRI1 activity. Full-length AtWRI1 increased TAG levels also in Japonica rice, indicating a conserved effect of WRI1 in rice lipid biosynthesis. The bio-methane production from rice straw was 20% higher in transformants than in the wild type. Moreover, a higher producing rate and final yield of methane was obtained for rice straw compared with rice husks, suggesting positive links between methane production and a high amount of fatty acids.ConclusionsOur results suggest that heterologous WRI1 expression in transgenic plants can be used to improve the metabolic potential for bioenergy purposes, in particular methane production.
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