Publications by authors named "Mariette Andersson"

19 Publications

  • Page 1 of 1

Amylose starch with no detectable branching developed through DNA-free CRISPR-Cas9 mediated mutagenesis of two starch branching enzymes in potato.

Sci Rep 2021 Feb 22;11(1):4311. Epub 2021 Feb 22.

Department of Plant Breeding, Swedish University of Agricultural Sciences, P.O. Box 101, 23053, Alnarp, Sweden.

DNA-free genome editing was used to induce mutations in one or two branching enzyme genes (Sbe) in tetraploid potato to develop starch with an increased amylose ratio and elongated amylopectin chains. By using ribonucleoprotein (RNP) transfection of potato protoplasts, a mutation frequency up to 72% was achieved. The large variation of mutations was grouped as follows: Group 1 lines with all alleles of Sbe1 mutated, Group 2 lines with all alleles of Sbe1 as well as two to three alleles of Sbe2 mutated and Group 3 lines having all alleles of both genes mutated. Starch from lines in Group 3 was found to be essentially free of amylopectin with no detectable branching and a chain length (CL) distribution where not only the major amylopectin fraction but also the shortest amylose chains were lost. Surprisingly, the starch still formed granules in a low-ordered crystalline structure. Starch from lines of Group 2 had an increased CL with a higher proportion of intermediate-sized chains, an altered granule phenotype but a crystalline structure in the granules similar to wild-type starch. Minor changes in CL could also be detected for the Group 1 starches when studied at a higher resolution.
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http://dx.doi.org/10.1038/s41598-021-83462-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7900246PMC
February 2021

A simplified method of determining the internal structure of amylopectin from barley starch without amylopectin isolation.

Carbohydr Polym 2021 Mar 13;255:117503. Epub 2020 Dec 13.

Department of Molecular Sciences, Swedish University of Agricultural Sciences, Box 7015, SE-750 07, Uppsala, Sweden. Electronic address:

To determine the internal structure of barley starch without amylopectin isolation, whole starch was hydrolyzed using β-amylase to remove the linear amylose and obtain β-limit dextrins (β-LDs). The β-LDs were treated with extensive α-amylase to prepare α-limit dextrins (α-LDs), and the α-LDs were further hydrolyzed with β-amylase into building blocks. The chain-length distribution of β-LD and building block composition were analyzed by size-exclusion chromatography and anion-exchange chromatography. The internal structure of the barley whole starches had similar pattern to barley amylopectins analyzed by conventional methods. The starch of barley amo1-mutated varieties contained more short internal B-chains and less long internal B-chains than that of other varieties. The starch from amo1-mutated varieties had more large building blocks than that from waxy varieties. The simplified method presented in this study can effectively characterize starch internal structure that relates to physicochemical properties of starch, although some details of amylopectin structure are not assessable.
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http://dx.doi.org/10.1016/j.carbpol.2020.117503DOI Listing
March 2021

Protoplast-Based Method for Genome Editing in Tetraploid Potato.

Methods Mol Biol 2021 ;2264:177-186

Department of Plant Breeding, Swedish University of Agricultural Sciences, Alnarp, Sweden.

The cultivated potato is tetraploid with four probably equivalent loci for each gene. A potato variety is furthermore commonly genetically heterogeneous and selected based on a beneficial genetic context which is maintained by clonal propagation. When introducing genetic changes by genome editing it is then desirable to achieve edits in all four loci for a certain gene target. This is in order to avoid crosses to achieve homozygosity for edited gene loci and at the same time reduce risk of inbreeding depression. In such a context transient transfection of protoplasts for the introduction of mutations, avoiding stable insertion of foreign DNA, would be very attractive. The protocol of this chapter has been shown to be applicable for the introduction of mutations by DNA vectors containing expression cassettes of TALEN, Cas9, and Cas9 deaminase fusions together with sgRNA expression cassettes on either single or separate vectors. Furthermore, the protoplast-based system has been shown to work very efficiently for mutations introduced by in vitro-produced and transfected RNP (ribonucleoprotein) complexes.
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http://dx.doi.org/10.1007/978-1-0716-1201-9_12DOI Listing
March 2021

Reduced Enzymatic Browning in Potato Tubers by Specific Editing of a Polyphenol Oxidase Gene Ribonucleoprotein Complexes Delivery of the CRISPR/Cas9 System.

Front Plant Sci 2019 9;10:1649. Epub 2020 Jan 9.

Laboratorio de Agrobiotecnología, INTA - EEA Balcarce, Balcarce, Argentina.

Polyphenol Oxidases (PPOs) catalyze the conversion of phenolic substrates to quinones, leading to the formation of dark-colored precipitates in fruits and vegetables. This process, known as enzymatic browning, is the cause of undesirable changes in organoleptic properties and the loss of nutritional quality in plant-derived products. In potato ( L.), PPOs are encoded by a multi-gene family with different expression patterns. Here, we have studied the application of the CRISPR/Cas9 system to induce mutations in the gene in the tetraploid cultivar Desiree. We hypothesized that the specific editing of this target gene would result in a lower PPO activity in the tuber with the consequent reduction of the enzymatic browning. Ribonucleoprotein complexes (RNPs), formed by two sgRNAs and Cas9 nuclease, were transfected to potato protoplasts. Up to 68% of regenerated plants contained mutations in at least one allele of the target gene, while 24% of edited lines carried mutations in all four alleles. No off-target mutations were identified in other analyzed genes. Mutations induced in the four alleles of gene, led to lines with a reduction of up to 69% in tuber PPO activity and a reduction of 73% in enzymatic browning, compared to the control. Our results demonstrate that the CRISPR/Cas9 system can be applied to develop potato varieties with reduced enzymatic browning in tubers, by the specific editing of a single member of the gene family.
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http://dx.doi.org/10.3389/fpls.2019.01649DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6962139PMC
January 2020

Altered Tuber Yield in Genetically Modified High-Amylose and Oil Potato Lines Is Associated With Changed Whole-Plant Nitrogen Economy.

Front Plant Sci 2018 15;9:342. Epub 2018 Mar 15.

Department of Crop Production Ecology, Swedish University of Agricultural Sciences, Uppsala, Sweden.

Breeding for improved crop quality traits can affect non-target traits related to growth and resource use, and these effects may vary in different cultivation conditions (e. g., greenhouse vs. field). The objectives of this study are to investigate the growth and whole-plant nitrogen (N) economy of two genetically modified (GM) potato lines compared to their non-GM parental varieties and when grown in different cultivation conditions. A high-amylose GM potato line and its parent were grown under field and greenhouse conditions for one growing season in Sweden; and a GM oil potato line and its parent were grown in greenhouse conditions only. Tuber yield, above ground biomass, N uptake efficiency and other plant N economy traits were assessed. In both cultivation conditions, the GM lines produced between 1.5 and two times more tubers as compared with their parents. In the greenhouse, fresh tuber yield and N uptake efficiency were unaffected by the genetic modifications, but the GM-lines produced less tuber biomass per plant-internal N compared to their parents. In the field, the fresh tuber yield was 40% greater in the high-amylose line as compared with its parent; the greater fresh tuber yield in the high-amylose GM line was accomplished by higher water allocation to the harvested tubers, and associated with increased N recovery from soil (+20%), N uptake efficiency (+53%), tuber N content (+20%), and N accumulation (+120%) compared with the non-GM parent. The cultivation conditions influenced the yield and N economy. For example, the final fresh above-ground plant biomass and N pool were considerably higher in the greenhouse conditions, whilst the tuber yield was higher in the field conditions. In conclusion, the genetic modification inducing high accumulation of amylose in potato tubers affected several non-target traits related to plant N economy, and increased the plant N uptake and accumulation efficiency of the field-grown plants. Due to strongly increased plant N accumulation compared to the parental variety, the cultivation of the high-amylose line is expected to require higher N fertilization rates. However, starch productivity per unit land area or soil N still is expected to be higher in the high-amylose line.
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http://dx.doi.org/10.3389/fpls.2018.00342DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5862821PMC
March 2018

Genome editing in potato via CRISPR-Cas9 ribonucleoprotein delivery.

Physiol Plant 2018 Dec 27;164(4):378-384. Epub 2018 Apr 27.

Department of Plant Breeding, Swedish University of Agricultural Sciences, P.O. Box 101, SE 23053 Alnarp, Sweden.

Clustered regularly interspaced short palindromic repeats and CRISPR-associated protein-9 (CRISPR-Cas9) can be used as an efficient tool for genome editing in potato (Solanum tuberosum). From both a scientific and a regulatory perspective, it is beneficial if integration of DNA in the potato genome is avoided. We have implemented a DNA-free genome editing method, using delivery of CRISPR-Cas9 ribonucleoproteins (RNPs) to potato protoplasts, by targeting the gene encoding a granule bound starch synthase (GBSS, EC 2.4.1.242). The RNP method was directly implemented using previously developed protoplast isolation, transfection and regeneration protocols without further adjustments. Cas9 protein was preassembled with RNA produced either synthetically or by in vitro transcription. RNP with synthetically produced RNA (cr-RNP) induced mutations, i.e. indels, at a frequency of up to 9%, with all mutated lines being transgene-free. A mutagenesis frequency of 25% of all regenerated shoots was found when using RNP with in vitro transcriptionally produced RNA (IVT-RNP). However, more than 80% of the shoots with confirmed mutations had unintended inserts in the cut site, which was in the same range as when using DNA delivery. The inserts originated both from DNA template remnants from the in vitro transcription, and from chromosomal potato DNA. In 2-3% of the regenerated shoots from the RNP-experiments, mutations were induced in all four alleles resulting in a complete knockout of the GBSS enzyme function.
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http://dx.doi.org/10.1111/ppl.12731DOI Listing
December 2018

Inhibition of plastid PPase and NTT leads to major changes in starch and tuber formation in potato.

J Exp Bot 2018 04;69(8):1913-1924

Department of Plant Breeding, Swedish University of Agricultural Sciences, Alnarp, Sweden.

The importance of a plastidial soluble inorganic pyrophosphatase (psPPase) and an ATP/ADP translocator (NTT) for starch composition and tuber formation in potato (Solanum tuberosum) was evaluated by individual and simultaneous down-regulation of the corresponding endogenous genes. Starch and amylose content of the transgenic lines were considerably lower, and granule size substantially smaller, with down-regulation of StpsPPase generating the most pronounced effects. Single-gene down-regulation of either StpsPPase or StNTT resulted in increased tuber numbers per plant and higher fresh weight yield. In contrast, when both genes were inhibited simultaneously, some lines developed only a few, small and distorted tubers. Analysis of metabolites revealed altered amounts of sugar intermediates, and a substantial increase in ADP-glucose content of the StpsPPase lines. Increased amounts of intermediates of vitamin C biosynthesis were also observed. This study suggests that hydrolysis of pyrophosphate (PPi) by action of a psPPase is vital for functional starch accumulation in potato tubers and that no additional mechanism for consuming, hydrolysing, or exporting PPi exists in the studied tissue. Additionally, it demonstrates that functional PPi hydrolysis in combination with efficient ATP import is essential for tuber formation and development.
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http://dx.doi.org/10.1093/jxb/ery051DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6018912PMC
April 2018

Resistant starch and other dietary fiber components in tubers from a high-amylose potato.

Food Chem 2018 Jun 3;251:58-63. Epub 2018 Jan 3.

Department of Molecular Sciences, Swedish University of Agricultural Sciences, Box 7015, SE-750 07 Uppsala, Sweden. Electronic address:

Tubers from a genetically modified high-amylose line T-2012 and its parental potato cultivar Dinamo were analyzed for resistant starch (RS) and dietary fiber (DF) after cooking and cold storage. For uncooked potatoes, the high-amylose tubers (30% of dry matter, DM) had much lower RS than the parent tubers (56% of DM). However, after cooking, the high-amylose tubers gave more RS (13% of DM) than the parent (4% of DM), and the RS level increased further to about 20% of DM after 1 day of cold storage. The altered RS content was attributable to changes in amylose content, starch granule structure, and amylopectin structure induced by the genetic modification. The high-amylose tubers also contained more DF (10-14% of DM) than the parent (5-7% of DM). Furthermore, cell wall composition was indirectly affected by the genetic modification, giving more cellulose and less pectin in the high-amylose tubers than the parent.
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http://dx.doi.org/10.1016/j.foodchem.2018.01.028DOI Listing
June 2018

A Specialized Diacylglycerol Acyltransferase Contributes to the Extreme Medium-Chain Fatty Acid Content of Seed Oil.

Plant Physiol 2017 May 21;174(1):97-109. Epub 2017 Mar 21.

Center for Plant Science Innovation and Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, Nebraska 68588 (U.I., J.E.S., H.J.K., R.E.C., E.B.C.);

Seed oils of many sp. contain >90% of medium-chain fatty acids, such as decanoic acid (10:0). These seed oils, which are among the most compositionally variant in the plant kingdom, arise from specialized fatty acid biosynthetic enzymes and specialized acyltransferases. These include lysophosphatidic acid acyltransferases (LPAT) and diacylglycerol acyltransferases (DGAT) that are required for successive acylation of medium-chain fatty acids in the -2 and -3 positions of seed triacylglycerols (TAGs). Here we report the identification of a cDNA for a DGAT1-type enzyme, designated CpuDGAT1, from the transcriptome of var developing seeds. Microsomes of camelina () seeds engineered for CpuDGAT1 expression displayed DGAT activity with 10:0-CoA and the diacylglycerol didecanoyl, that was approximately 4-fold higher than that in camelina seed microsomes lacking CpuDGAT1. In addition, coexpression in camelina seeds of CpuDGAT1 with a FatB thioesterase (CvFatB1) that generates 10:0 resulted in TAGs with nearly 15 mol % of 10:0. More strikingly, expression of CpuDGAT1 and CvFatB1 with the previously described CvLPAT2, a 10:0-CoA-specific LPAT, increased 10:0 amounts to 25 mol % in camelina seed TAG. These TAGs contained up to 40 mol % 10:0 in the -2 position, nearly double the amounts obtained from coexpression of CvFatB1 and CvLPAT2 alone. Although enriched in diacylglycerol, 10:0 was not detected in phosphatidylcholine in these seeds. These findings are consistent with channeling of 10:0 into TAG through the combined activities of specialized LPAT and DGAT activities and demonstrate the biotechnological use of these enzymes to generate 10:0-rich seed oils.
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http://dx.doi.org/10.1104/pp.16.01894DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5411140PMC
May 2017

Efficient targeted multiallelic mutagenesis in tetraploid potato (Solanum tuberosum) by transient CRISPR-Cas9 expression in protoplasts.

Plant Cell Rep 2017 Jan 3;36(1):117-128. Epub 2016 Oct 3.

Department of Plant Breeding, Swedish University of Agricultural Sciences, P.O. Box 101, SE-23053, Alnarp, Sweden.

Key Message: Altered starch quality with full knockout of GBSS gene function in potato was achieved using CRISPR-Cas9 technology, through transient transfection and regeneration from isolated protoplasts. Site-directed mutagenesis (SDM) has shown great progress in introducing precisely targeted mutations. Engineered CRISPR-Cas9 has received increased focus compared to other SDM techniques, since the method is easily adapted to different targets. Here, we demonstrate that transient application of CRISPR-Cas9-mediated genome editing in protoplasts of tetraploid potato (Solanum tuberosum) yielded mutations in all four alleles in a single transfection, in up to 2 % of regenerated lines. Three different regions of the gene encoding granule-bound starch synthase (GBSS) were targeted under different experimental setups, resulting in mutations in at least one allele in 2-12 % of regenerated shoots, with multiple alleles mutated in up to 67 % of confirmed mutated lines. Most mutations resulted in small indels of 1-10 bp, but also vector DNA inserts of 34-236 bp were found in 10 % of analysed lines. No mutations were found in an allele diverging one bp from a used guide sequence, verifying similar results found in other plants that high homology between guide sequence and target region near the protospacer adjacent motif (PAM) site is essential. To meet the challenge of screening large numbers of lines, a PCR-based high-resolution fragment analysis method (HRFA) was used, enabling identification of multiple mutated alleles with a resolution limit of 1 bp. Full knockout of GBSS enzyme activity was confirmed in four-allele mutated lines by phenotypic studies of starch. One remaining wild-type (WT) allele was shown sufficient to maintain enough GBSS enzyme activity to produce significant amounts of amylose.
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http://dx.doi.org/10.1007/s00299-016-2062-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5206254PMC
January 2017

Potato tuber expression of Arabidopsis WRINKLED1 increase triacylglycerol and membrane lipids while affecting central carbohydrate metabolism.

Plant Biotechnol J 2016 09 17;14(9):1883-98. Epub 2016 Mar 17.

Department of Plant Breeding, Swedish University of Agricultural Sciences, Alnarp, Sweden.

Tuber and root crops virtually exclusively accumulate storage products in the form of carbohydrates. An exception is yellow nutsedge (Cyperus esculentus) in which tubers have the capacity to store starch and triacylglycerols (TAG) in roughly equal amounts. This suggests that a tuber crop can efficiently handle accumulation of energy dense oil. From a nutritional as well as economic aspect, it would be of interest to utilize the high yield capacity of tuber or root crops for oil accumulation similar to yellow nutsedge. The transcription factor WRINKLED1 from Arabidopsis thaliana, which in seed embryos induce fatty acid synthesis, has been shown to be a major factor for oil accumulation. WRINKLED1 was expressed in potato (Solanum tuberosum) tubers to explore whether this factor could impact tuber metabolism. This study shows that a WRINKLED1 transcription factor could induce triacylglycerol accumulation in tubers of transformed potato plants grown in field (up to 12 nmol TAG/mg dry weight, 1% of dry weight) together with a large increase in polar membrane lipids. The changes in metabolism further affected starch accumulation and composition concomitant with massive increases in sugar content.
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http://dx.doi.org/10.1111/pbi.12550DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5069604PMC
September 2016

Structurally divergent lysophosphatidic acid acyltransferases with high selectivity for saturated medium chain fatty acids from Cuphea seeds.

Plant J 2015 Dec;84(5):1021-33

Department of Biochemistry and Center for Plant Science Innovation, University of Nebraska-Lincoln, Lincoln, NE, 68588, USA.

Lysophosphatidic acid acyltransferase (LPAT) catalyzes acylation of the sn-2 position on lysophosphatidic acid by an acyl CoA substrate to produce the phosphatidic acid precursor of polar glycerolipids and triacylglycerols (TAGs). In the case of TAGs, this reaction is typically catalyzed by an LPAT2 from microsomal LPAT class A that has high specificity for C18 fatty acids containing Δ9 unsaturation. Because of this specificity, the occurrence of saturated fatty acids in the TAG sn-2 position is infrequent in seed oils. To identify LPATs with variant substrate specificities, deep transcriptomic mining was performed on seeds of two Cuphea species producing TAGs that are highly enriched in saturated C8 and C10 fatty acids. From these analyses, cDNAs for seven previously unreported LPATs were identified, including cDNAs from Cuphea viscosissima (CvLPAT2) and Cuphea avigera var. pulcherrima (CpuLPAT2a) encoding microsomal, seed-specific class A LPAT2s and a cDNA from C. avigera var. pulcherrima (CpuLPATB) encoding a microsomal, seed-specific LPAT from the bacterial-type class B. The activities of these enzymes were characterized in Camelina sativa by seed-specific co-expression with cDNAs for various Cuphea FatB acyl-acyl carrier protein thioesterases (FatB) that produce a variety of saturated medium-chain fatty acids. CvLPAT2 and CpuLPAT2a expression resulted in accumulation of 10:0 fatty acids in the Camelina sativa TAG sn-2 position, indicating a 10:0 CoA specificity that has not been previously described for plant LPATs. CpuLPATB expression generated TAGs with 14:0 at the sn-2 position, but not 10:0. Identification of these LPATs provides tools for understanding the structural basis of LPAT substrate specificity and for generating altered oil functionalities.
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http://dx.doi.org/10.1111/tpj.13063DOI Listing
December 2015

Improved material properties of solution-cast starch films: Effect of varying amylopectin structure and amylose content of starch from genetically modified potatoes.

Carbohydr Polym 2015 Oct 19;130:388-97. Epub 2015 May 19.

Department of Food Science, Swedish University of Agricultural Sciences, Box 7051, SE-750 07 Uppsala, Sweden.

High-amylose potato starches were produced through genetic modification resulting in changed granule morphology and composition, with higher amylose content and increased chain length of amylopectin. The increased amylose content and structural changes in amylopectin enhanced film-forming behavior and improved barrier and tensile properties in starch films. The molecular structure in these starches was related to film-forming properties. Solution-cast films of high-amylose starch revealed a homogeneous structure with increasing surface roughness at higher amylose content, possibly due to amylose aggregation. Films exhibited significantly higher stress and strain at break compared with films of wild-type starch, which could be attributable to the longer chains of amylopectin being involved in the interconnected network and more interaction between chains, as shown using transmission electron microscopy. The oxygen permeability of high-amylose starch films was significantly decreased compared with wild-type starch. The nature of the modified starches makes them an interesting candidate for replacement of non-renewable oxygen and grease barrier polymers used today.
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http://dx.doi.org/10.1016/j.carbpol.2015.05.024DOI Listing
October 2015

Targeted gene mutation in tetraploid potato through transient TALEN expression in protoplasts.

J Biotechnol 2015 Jun 4;204:17-24. Epub 2015 Apr 4.

Department of Plant Breeding, Swedish University of Agricultural Sciences, Alnarp, Sweden. Electronic address:

Potato is the third largest food crop in the world, however, the high degree of heterozygosity, the tetrasomic inheritance and severe inbreeding depression are major difficulties for conventional potato breeding. The rapid development of modern breeding methods offers new possibilities to enhance breeding efficiency and precise improvement of desirable traits. New site-directed mutagenesis techniques that can directly edit the target genes without any integration of recombinant DNA are especially favorable. Here we present a successful pipeline for site-directed mutagenesis in tetraploid potato through transient TALEN expression in protoplasts. The transfection efficiency of protoplasts was 38-39% and the site-directed mutation frequency was 7-8% with a few base deletions as the predominant type of mutation. Among the protoplast-derived calli, 11-13% showed mutations and a similar frequency (10%) was observed in the regenerated shoots. Our results indicate that the site-directed mutagenesis technology could be used as a new breeding method in potato as well as for functional analysis of important genes to promote sustainable potato production.
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http://dx.doi.org/10.1016/j.jbiotec.2015.03.021DOI Listing
June 2015

Nanostructural morphology of plasticized wheat gluten and modified potato starch composites: relationship to mechanical and barrier properties.

Biomacromolecules 2015 Mar 17;16(3):695-705. Epub 2015 Feb 17.

Department of Plant Breeding, Swedish University of Agricultural Sciences , Box 101, SE-230 53 Alnarp, Sweden.

In the present study, we were able to produce composites of wheat gluten (WG) protein and a novel genetically modified potato starch (MPS) with attractive mechanical and gas barrier properties using extrusion. Characterization of the MPS revealed an altered chain length distribution of the amylopectin fraction and slightly increased amylose content compared to wild type potato starch. WG and MPS of different ratios plasticized with either glycerol or glycerol and water were extruded at 110 and 130 °C. The nanomorphology of the composites showed the MPS having semicrystalline structure of a characteristic lamellar arrangement with an approximately 100 Å period observed by small-angle X-ray scattering and a B-type crystal structure observed by wide-angle X-ray scattering analysis. WG has a structure resembling the hexagonal macromolecular arrangement as reported previously in WG films. A larger amount of β-sheets was observed in the samples 70/30 and 30/70 WG-MPS processed at 130 °C with 45% glycerol. Highly polymerized WG protein was found in the samples processed at 130 °C versus 110 °C. Also, greater amounts of WG protein in the blend resulted in greater extensibility (110 °C) and a decrease in both E-modulus and maximum stress at 110 and 130 °C, respectively. Under ambient conditions the WG-MPS composite (70/30) with 45% glycerol showed excellent gas barrier properties to be further explored in multilayer film packaging applications.
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http://dx.doi.org/10.1021/bm5017496DOI Listing
March 2015

Starch biosynthetic genes and enzymes are expressed and active in the absence of starch accumulation in sugar beet tap-root.

BMC Plant Biol 2014 Apr 23;14:104. Epub 2014 Apr 23.

Department of Plant Breeding, Swedish University of Agricultural Sciences, P,O, Box 101, SE-23053 Alnarp, Sweden.

Background: Starch is the predominant storage compound in underground plant tissues like roots and tubers. An exception is sugar beet tap-root (Beta vulgaris ssp altissima) which exclusively stores sucrose. The underlying mechanism behind this divergent storage accumulation in sugar beet is currently not fully known. From the general presence of starch in roots and tubers it could be speculated that the lack in sugar beet tap-roots would originate from deficiency in pathways leading to starch. Therefore with emphasis on starch accumulation, we studied tap-roots of sugar beet using parsnip (Pastinaca sativa) as a comparator.

Results: Metabolic and structural analyses of sugar beet tap-root confirmed sucrose as the exclusive storage component. No starch granules could be detected in tap-roots of sugar beet or the wild ancestor sea beet (Beta vulgaris ssp. maritima). Analyses of parsnip showed that the main storage component was starch but tap-root tissue was also found to contain significant levels of sugars. Surprisingly, activities of four main starch biosynthetic enzymes, phosphoglucomutase, ADP-glucose pyrophosphorylase, starch synthase and starch branching enzyme, were similar in sugar beet and parsnip tap-roots. Transcriptional analysis confirmed expression of corresponding genes. Additionally, expression of genes involved in starch accumulation such as for plastidial hexose transportation and starch tuning functions could be determined in tap-roots of both plant species.

Conclusion: Considering underground storage organs, sugar beet tap-root upholds a unique property in exclusively storing sucrose. Lack of starch also in the ancestor sea beet indicates an evolved trait of biological importance.Our findings in this study show that gene expression and enzymatic activity of main starch biosynthetic functions are present in sugar beet tap-root during storage accumulation. In view of this, the complete lack of starch in sugar beet tap-roots is enigmatic.
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http://dx.doi.org/10.1186/1471-2229-14-104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4108048PMC
April 2014

Comparative transcriptome analysis of three oil palm fruit and seed tissues that differ in oil content and fatty acid composition.

Plant Physiol 2013 Jul 4;162(3):1337-58. Epub 2013 Jun 4.

Institut de Recherche pour le Développement, Unité Mixte de Recherche Diversité, Adaptation et Développement des Plantes, BP 64501, 34394 Montpellier, France.

Oil palm (Elaeis guineensis) produces two oils of major economic importance, commonly referred to as palm oil and palm kernel oil, extracted from the mesocarp and the endosperm, respectively. While lauric acid predominates in endosperm oil, the major fatty acids (FAs) of mesocarp oil are palmitic and oleic acids. The oil palm embryo also stores oil, which contains a significant proportion of linoleic acid. In addition, the three tissues display high variation for oil content at maturity. To gain insight into the mechanisms that govern such differences in oil content and FA composition, tissue transcriptome and lipid composition were compared during development. The contribution of the cytosolic and plastidial glycolytic routes differed markedly between the mesocarp and seed tissues, but transcriptional patterns of genes involved in the conversion of sucrose to pyruvate were not related to variations for oil content. Accumulation of lauric acid relied on the dramatic up-regulation of a specialized acyl-acyl carrier protein thioesterase paralog and the concerted recruitment of specific isoforms of triacylglycerol assembly enzymes. Three paralogs of the WRINKLED1 (WRI1) transcription factor were identified, of which EgWRI1-1 and EgWRI1-2 were massively transcribed during oil deposition in the mesocarp and the endosperm, respectively. None of the three WRI1 paralogs were detected in the embryo. The transcription level of FA synthesis genes correlated with the amount of WRI1 transcripts and oil content. Changes in triacylglycerol content and FA composition of Nicotiana benthamiana leaves infiltrated with various combinations of WRI1 and FatB paralogs from oil palm validated functions inferred from transcriptome analysis.
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http://dx.doi.org/10.1104/pp.113.220525DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3707537PMC
July 2013

Field performance and starch characteristics of high-amylose potatoes obtained by antisense gene targeting of two branching enzymes.

Plant Biotechnol J 2004 Jul;2(4):311-20

Plant Science Sweden AB, Herman Ehles Väg 2-4, 268 31 Svalöv, Sweden.

Potato is an important crop for starch production, but there are limitations regarding the genetic variation of starch quality. In maize, starches with various properties have been available for a long time by mutational breeding. Amylose starch from potatoes differs from cereal amyloses in several functionally important aspects, such as a higher degree of polymerization. Areas of application in which the degree of polymerization is of importance include film forming and the polymeric properties of bioplastics. High-amylose potato lines have been achieved by inhibiting the two known branching enzyme forms of potato. A single inserted gene construct for the inhibition of both forms resulted in structural changes of the starch to levels of branching that were below the commercially available amylose standards of potato. The high-amylose potato lines were tested in multiple year field trials of agronomic performance and were used for the pilot plant production of starch. The introduced trait was confirmed to be stable over multiple years. The consequences of the modification were found to be an increased tuber yield, reduced starch content, smaller granule size and an increase in reducing sugars.
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http://dx.doi.org/10.1111/j.1467-7652.2004.00073.xDOI Listing
July 2004

Targeted gene suppression by RNA interference: an efficient method for production of high-amylose potato lines.

J Biotechnol 2006 May 8;123(2):137-48. Epub 2006 Feb 8.

Plant Science Sweden AB, Herman Ehles Väg 2-4, 268 31 Svalöv, Sweden.

Production of high-amylose potato lines can be achieved by inhibition of two genes coding for starch branching enzymes. The use of antisense technology for gene inhibition have yielded a low frequency of high-amylose lines that mostly was correlated with high numbers of integrated T-DNA copies. To investigate whether the production of high-amylose lines could be improved, RNA interference was used for gene inhibition of the genes Sbe1 and Sbe2. Two constructs with 100 bp segments (pHAS2) or 200 bp segments (pHAS3) of both branching enzyme genes were cloned as inverted repeats controlled by a potato granule-bound starch synthase promoter. The construct pHAS3 was shown to be very efficient, yielding high-amylose quality in more than 50% of the transgenic lines. An antisense construct, included in the study as a comparator, resulted in only 3% of the transgenic lines being of high-amylose type. Noticeable was also that pHAS3 yielded low T-DNA copy inserts with an average of 83% of backbone-free transgenic lines being single copy events.
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http://dx.doi.org/10.1016/j.jbiotec.2005.11.001DOI Listing
May 2006
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