Amaranth Species Research Articles

The Amaranth Genome: Genome, Transcriptome, and Physical Map Assembly.

Amaranth ( L.) is an emerging pseudocereal native to the New World that has garnered increased attention in recent years because of its nutritional quality, in particular its seed protein and more specifically its high levels of the essential amino acid lysine. It belongs to the Amaranthaceae family, is an ancient paleopolyploid that shows disomic inheritance (2 = 32), and has an estimated genome size of 466 Mb. Here we present a high-quality draft genome sequence of the grain amaranth. The genome assembly consisted of 377 Mb in 3518 scaffolds with an N of 371 kb. Repetitive element analysis predicted that 48% of the genome is comprised of repeat sequences, of which -like elements were the most commonly classified retrotransposon. A de novo transcriptome consisting of 66,370 contigs was assembled from eight different amaranth tissue and abiotic stress libraries. Annotation of the genome identified 23,059 protein-coding genes. Seven grain amaranths (, , and ) and their putative progenitor () were resequenced. A single nucleotide polymorphism (SNP) phylogeny supported the classification of as the progenitor species of the grain amaranths. Lastly, we generated a de novo physical map for using the BioNano Genomics' Genome Mapping platform. The physical map spanned 340 Mb and a hybrid assembly using the BioNano physical maps nearly doubled the N of the assembly to 697 kb. Moreover, we analyzed synteny between amaranth and sugar beet ( L.) and estimated, using analysis, the age of the most recent polyploidization event in amaranth.

PIGMENT CONTENT AND COMPOSITION IN AUTOTROPHIC AND HETEROTROPHIC LEAF TISSUES OF AMARANTH SPECIES A. TRICOLOR L.

At present there is numerous evidence of the antioxidant positive role in the defensive reaction that is capable to protect not only plants, but also humans against oxidative stress. Plant pigments such as natural dyes from leaves, flowers and fruits are known to have high antioxidant activity. Amaranth species A. tricolor L. cultivar ‘Early Splendor’ is a convenient model for the comparative studying of the formation processes of differently colored pigment composition in leaf tissues that differs in the ability to photosynthesize. Leaves of amaranth cultivar ‘Valentina’ were as a standard. The aim of the experiment was a comparative studying of the pigments content: amaranthine, chlorophyll a and b, carotenoids in the cauline leaves of amaranth cultivars ‘Valentina’ and ‘Early Splendor’, as well as in the red and green areas of the leaves. Analysis of the aqueous extract of red Early Splendor amaranth apical leaves showed the presence of betacyanin pigment - amaranthine, in the absorption spectrum in which peak was seen in the green region at 540 nm. In addition to the antioxidant amaranthine there are also antioxidants which might be phenolic glycosides, and ascorbic acid in the extract, the total content of which is almost twice as small as in the leaves of amaranth cauline of this cultivar. Yellow fraction was found in the ethanolic extract of red leaves. Its absorption spectrum had peaks in the blue region at 445 nm and 472 nm and a shoulder at 422 nm that indicated the presence of betaxanthin, betalamic acid or carotenoids. Water-soluble antioxidants - amaranthine and ascorbic acid were found in auline leaves of studied species. Their content in the leaves of Valentina cultivar was higher than in the leaves of cultivar ‘Early Splendor’, and the maximum level of photosynthetic pigments was found in ‘Early Splendor’ leaves. The obtained results showed that the amaranth is a promising source of pigments with the antioxidant activity that can be used for production of food dyes.

Identification of novel saponins in vegetable amaranth and characterization of their hemolytic activity

Amaranth is a plant genus of global importance comprising more than 60 species that are dually used for human consumption. While the grains are used as pseudo-cereals mainly in America and Asia, leaves are also consumed as leafy vegetable in African countries. Besides further secondary plant metabolites, saponins are described as major bioactive constituents in amaranth species. These triterpenoid saponins belonging to the oleanane-type are assumed to be part of the plant defense system and are often also associated with potential health risks for the consumer, mainly due to hemolytic properties. However, data concerning amaranth saponins are limited to the grains of single cultivars of only a few species investigated.The aim of the present work was to determine the saponin profile in leaves of various amaranth cultivars grown under identical conditions. Out of 15 cultivars, six did not show any indications for the presence of saponins in HPLC–MS analysis. Two saponin-rich cultivars (one of Amaraanthus hybridus and one of Amaranthus hypochondriacus) as well as commercially available amaranth grains were selected for an in-depth analysis using a combined approach of high resolution and multi stage mass spectrometry. Three previously undescribed monodesmosidic and four bidesmosidic saponins could be assigned according to the MS data. Four novel saponins were also found in commercial grain amaranth analyzed for comparison. The investigation of the hemolytic effects revealed that only one saponin exerts significant activity whilst the further saponins did not lyse erythrocytes in vitro.The results show that the saponin profile of amaranth cultivars is more diverse than reported so far. However, the biological activity seems to be different for the single structures. Thus, a more comprehensive case-by-case investigation of amaranth saponins is required to evaluate the impact of these secondary metabolites on humans and plants.

Selenium Supplementation of Amaranth Sprouts Influences Betacyanin Content and Improves Anti-Inflammatory Properties via NFκB in Murine RAW 264.7 Macrophages.

Sprouts contain potent compounds which while influencing crucial transduction pathways in cell reveal anti-inflammatory and anticancer activities. In this study, we report the biological activity for seeds and colourful sprouts of four types of edible amaranth, as amaranth has recently attracted interest due to its appreciable nutritional value. MTT assay conducted for the amaranth seeds and sprouts did not show any adverse effect on the viability of murine RAW 264.7 cells. As amaranth accumulates selenium, the sprouts were supplemented with this trace element (10 mg/L; 15 mg/L Se as sodium selenite) while growing. Selenium concentration in sprouts was observed to be significantly correlated with betacyanins content of the tested species. The amounts of Se and betacyanins in sprouts varied for various Amaranth species. In the present study, Amaranthus cruentus sprouts with the highest betacyanins (19.30 ± 0.57–28.85 ± 2.23 mg of amaranthin/100 g of fresh weight) and high total selenium (22.51 ± 1.57–1044.75 ± 73.08 μg/L in methanol extracts) content prevented NFκB translocation to the cell nucleus and subsequently exerted an anti-inflammatory effect by significant decreasing inflammatory interleukin 6 production (587.3 ± 34.2–710.0 ± 88.1 pg/mL) in the cell culture of activated RAW 264.7 macrophages (vs LPS control 1520 ± 114 pg/mL).Electronic supplementary materialThe online version of this article (doi:10.1007/s12011-015-0429-x) contains supplementary material, which is available to authorized users.

Structures, physicochemical properties, and applications of amaranth starch

ABSTRACTAmaranth as a rediscovered “new” crop is becoming a research focus in the recent two decades. The major carbohydrate of some amaranth species is starch, which accounts up to around 60% of the dry grains. This review summarizes the present knowledge of the isolation, composition, structures, physiochemical properties, modifications, and applications of amaranth starches, and provides suggestions for research to further improve the utilization.

Amaranthus parganensis(Amaranthaceae), a New Species from West Bengal, India

Abstract. A new species of vegetable amaranth, Amaranthus parganensis Saubhik Das (Amaranthaceae), has been described from the lower Gangetic Plain of West Bengal, India. Its morphology and distributional range suggest that it is related to A. tricolor L., both adapted to tropical climate, having a sympatric distribution and similar ecology.

Fungi occurring on the plants of the genus Amaranthus L.

A study of fungi on Amaranthus spp. was performed in 2007-2009. The following forms of the genus were taken under consideration: cultivated amaranth (A. cruentus) and a wild form (A. retroflexus) growing as a weed on amaranth and sugar beet plantations and growing as a ruderal weed. The aim of the work was to determine which fungi communities occur in the phyllosphere, roots, rhizoplane, and rhizosphere of Amaranthus spp. To investigate the phyllosphere fungi communities, 5 plants were taken in the seed formation phase. From each plant, 3 healthy, symptomless leaves were taken. In addition, the isolation of fungi communities from the roots, rhizoplane, and rhizosphere was performed in the seed formation phase. Ten plants from each location were taken along the diagonal of the plot. In total, 38 species of fungi were isolated from the phyllosphere of Amaranthus spp., and of that number, 30 were collected from A. cruentus and 29 from A. retroflexus. In total, 29 fungi species were isolated from the roots of all the observed forms of amaranth. From the rhizosphere of all amaranth species tested, 44 fungi species were isolated. The most frequently recorded taxa within the associations of fungi isolated from the phyllosphere were Cladosporium spp. (C. cladosporioides and C. herbarum), Alternaria alternata, and Epicoccum nigrum. Fungi species of the genera Penicillium, Fusarium, Cladosporium, and Phoma, as well as species of A. alternata, were isolated in great abundance from the roots and the rhizosphere of the amaranth plants.

Amaranthus cruentus L. is suitable for cultivation in Central Italy: field evaluation and response to plant densities

The aim of this study was to determine the possibility of amaranth cultivation in Central Italy and to determine the optimum plant density. Field trials were carried out in 2011 and 2012 under non-irrigated conditions in Tuscany (43° 18’ N, 11° 47’ E). Twelve accessions of two amaranth species (Amaranthus cruentus L. and A. hypochondriacus L.) were utilised. Genotypes were evaluated over a two-year period using a RCB design with three replicates. The effects of plant density were investigated in 2012. A with a split-plot design was used, where the A. cruentus accessions (AMES 5148, PI 511719 and PI 643045) constituted the main plots. Plant densities (7.5, 15, 30 and 60 plants m–2) constituted the subplots. Plants were transplanted at the 3-4 true leaf stage. Morphological traits were determined using 5 plants selected from the two central rows of the sampling area. Plots were hand-harvested and cleaned with a mechanical grid with appropriate sieve diameters. A. cruentus was shown to be more suitable to the Central Italy agro-ecological conditions than A. hypochondriacus. The accessions derived from Mexico (PI 477913, PI 576481, PI 643045, PI 643053, and PI 6495079), Guatemala (PI 511719) and Puerto Rico (AMES 5148), had both higher grain yields and a greater stability over the two-year period, with a mean grain production ranging from 2.8 to 3.2 t ha–1. The severe climatic stress in 2012 (high temperatures and aridity), resulted in a 43-60% reduction in seed production compared to that of the previous year. Under these conditions, PI 511719, AMES 26015, AMES 5386, AMES 5148, PI 477913 yielded on average 1.9 t ha–1. Yields of A. hypochondriacus were negligible in both years, probably attributable to greater photoperiod sensitivity, resulting in reduced flowering and delayed maturity. By increasing density up to 60 and 30 plants m–2 for PI 511719 and AMES 5148, respectively, grain production was increased by 55%. As the plant population increased, plant height, panicle length and diameter, basal stem diameter and branches per plant were reduced. AMES 5148 plants were composed of one single stem with apical panicles at a density of 30 plants m–2, whilst the other two accessions were similar at the highest plant density. Branch number per plant was negatively correlated with grain production (r=–0.536**). Hence, A. cruentus may be grown in Central Italy even under arid conditions, using a density of either 30 or 60 plants m–2 according to the genotype used.

Analysis of the sequences of internal transcribed spacers ITS1, ITS2 and the 5.8S ribosomal gene of species of the Amaranthus genus

Analysis of the sequence ITS1-5.8S-ITS2 in 11 samples of the amaranth species (Amaranthus caudatus, A. cruentus, A. hybridus, A. tricolor, A. paniculatus, A. hypohondriacus) was performed. It has been shown that the variability of the sequences of the intergenic spacers ITS1, ITS2 and 5.8S rRNA gene of the amaranth species analyzed is extremely low. A possible secondary structure of the 5.8S rRNA molecule was determined for the first time; three conservative motifs were identified. A single nucleotide substitution found in A. hybridus did not change the loop topology. In the sample of Celosia cristata taken as an external group, a four-nucleotide insertion in the 5'-end of the gene and a one-nucleotide deletion in the fourth hairpin not affecting the general topology of the 5.8S rRNA molecule were found.

Antioxidant enzyme activity, proline accumulation, leaf area and cell membrane stability in water stressed Amaranthus leaves

Traditional crops are extremely important for food production in low income, food-deficit countries (LIFDCs) where they continue to be maintained by socio-cultural preferences and traditional uses. Significant potential exists to improve these crops, one of which is to select for improved productivity during moisture stress conditions. Germplasm of Amaranthus tricolor, Amaranthus hypochondriacus and Amaranthus hybridus were subjected to various screening methods to measure metabolic and physiological changes during water stress. The activities of enzymes involved in the oxygen-scavenging system during abiotic stress conditions (superoxide dismutase (SOD), ascorbate peroxidase (APX) and glutathione reductase (GR)), free proline production, leaf area (LA), cell membrane stability (CMS), leaf water potential (LWP) and relative water content (RWC) were measured in these three amaranth species during induced water stress. This study showed significant differences in metabolic responses during water deficit of the three species tested. Moisture stress and a decrease in RWC and LWP were first experienced in A. hybridus and A. hypochondriacus, followed by A. tricolor. There was an indirect correlation between leaf water status (RWC and LWP), enzyme activity, proline production and leaf area. The combined effect of GR, APX and SOD could ensure higher levels of regulation of the toxic effect of H2O2 which could be associated with drought tolerance in Amaranthus. Distinct differences in onset of proline accumulation and the amount of accumulated in leaves upon induced water stress was noticed for the three amaranth species tested. Proline accumulation during water stress conditions in amaranth seems to be indirect and could possibly have a protective role apart from osmoregulation during stress conditions. This contention is supported by the decrease in leaf area and high cell membrane stability for two of the species tested. This study forms part of a project aimed at the development of improved traditional crops to contribute to food production and quality for subsistence farmers in areas with low precipitation or variable rainfall patterns.

Nutritional diversity of leafy amaranth species grown in Kenya

Objectives: Despite the availability of many species of amaranth in Kenya, there is inadequate information on their nutritional diversity and how they can be best used in mitigation of malnutrition. Hence, this study was aimed at investigating the nutritional diversity of five leafy amaranth species grown in Kenya. Methodology and results: Amaranth vegetables were harvested 5-6 weeks after planting. They were analyzed for their nutritional and phytochemical content using standard methods. Moisture content was lowest in A.cruentus (79.29±1.26%). A.cruentus had the highest protein (4.37 ±0.02%).The fat content of A.hybridus (2.53±0.06%) and A.hypochodriacus (2.63±0.11%) was significantly higher than the other species. The crude fibre content (3.01±0.29%) was similar in all the species. A.dubius had the highest calcium (336.47±0.9mg/100g) and iron (18.64±0.23mg/100g) while A.cruentus was highest in zinc content (1.67±0.03mg/100g).The total phenols content was highest in A.cruentus (3.59±0.01GAE/100mg).Catechins were highest in A.cruentus (7.15±0.63CE/100mg) while quercetin was highest in A.cruentus (14.28±0.86QE/100mg). Conclusion and application of results: More than 50% of the Kenyan population live below the poverty line and lack access to adequate food. The food available food is of poor nutritional value, this causes malnutrition and other nutritional disorders. There is therefore a need to identify nutrient-rich foods that can be produced inexpensively to meet the nutrient requirements of everyone and especially the vulnerable groups, such as pregnant women, children and the elderly. Amaranth is drought tolerant, grows fast and is easily cultivated. This study indicates that the Amaranth species found in Kenya are a good source of key nutrients, which can be used in mitigation of malnutrition. A.dubius is a superior source of calcium and iron and can help curb the micronutrient deficiencies in Kenya, while A.cruentus is a superior source of protein and phytochemicals which are important in reducing the risk of chronic diseases which are so rampant in Kenya today.Keywords: A. dubius; A. hybridus and A. cruentus; A. albus and A. hypochondriacus; Nutrients

Nutritional diversity of leafy amaranth species grown in Kenya

Objectives : Despite the availability of many species of amaranth in Kenya, there is inadequate information on their nutritional diversity and how they can be best used in mitigation of malnutrition. Hence, this study was aimed at investigating the nutritional diversity of five leafy amaranth species grown in Kenya. Methodology and results : Amaranth vegetables were harvested 5-6 weeks after planting. They were analyzed for their nutritional and phytochemical content using standard methods. Moisture content was lowest in A.cruentus (79.29±1.26%). A.cruentus had the highest protein (4.37 ±0.02%).The fat content of A.hybridus (2.53±0.06%) and A.hypochodriacus (2.63±0.11%) was significantly higher than the other species. The crude fibre content (3.01±0.29%) was similar in all the species. A.dubius had the highest calcium (336.47±0.9mg/100g) and iron (18.64±0.23mg/100g) while A.cruentus was highest in zinc content (1.67±0.03mg/100g).The total phenols content was highest in A.cruentus (3.59±0.01GAE/100mg).Catechins were highest in A.cruentus (7.15±0.63CE/100mg) while quercetin was highest in A.cruentus (14.28±0.86QE/100mg). Conclusion and application of results : More than 50% of the Kenyan population live below the poverty line and lack access to adequate food. The food available food is of poor nutritional value, this causes malnutrition and other nutritional disorders. There is therefore a need to identify nutrient-rich foods that can be produced inexpensively to meet the nutrient requirements of everyone and especially the vulnerable groups, such as pregnant women, children and the elderly. Amaranth is drought tolerant, grows fast and is easily cultivated. This study indicates that the Amaranth species found in Kenya are a good source of key nutrients, which can be used in mitigation of malnutrition. A.dubius is a superior source of calcium and iron and can help curb the micronutrient deficiencies in Kenya, while A.cruentus is a superior source of protein and phytochemicals which are important in reducing the risk of chronic diseases which are so rampant in Kenya today. Keywords : A. dubius; A. hybridus and A. cruentus; A. albus and A. hypochondriacus; Nutrients

Molecular characterization and genetic diversity of the starch branching enzyme (SBE) gene from Amaranthus: the evolutionary origin of grain amaranths

Full-length genomic DNA clones encoding starch branching enzymes (SBEs) from three cultivated grain amaranths were isolated and characterized. A sequence analysis showed that the SBE genes from Amaranthus caudatus, A. cruentus, and A. hypochondriacus, viz. SBE-ca, SBE-cr, and SBE-hy, respectively, contained the same exon and intron structure and consisted of 961 amino acids with molecular weights of approximately 108.4 kDa. The lengths of the genes were 7,446 (SBE-ca), 7,453 (SBE-cr), and 7,443 bp (SBE-hy), and they are highly homologous. We also tested, for the first time, the potential use of SBE from diverse genetic sources to answer long-standing questions on the origin of grain amaranths. The maximum parsimony tree based on the SBE sequence data suggests that A. hybridus is the progenitor wild species of grain amaranths. The results could be summarized as follows: (1) the A. hybridus species comprise all grain amaranths, which can then be divided into two subgroups; (2) A. caudatus and A. hypochondriacus and/or A. caudatus and A. quitensis appear to be closely related to each other; (3) A. powellii was the most distantly related to the other five species in the A. hybridus complex. These results indicate that A. hybridus is the ancestor species in at least two separate domestication events, lending support to the theory of three separate, rather than one, domestication events involving A. hybridus as the common ancestor. A new approach to investigating the genetic diversity in Amaranthus using the nuclear gene SBE provided useful information on the evolutionary origin of three cultivated species.

Adaptation of Cultivated Amaranth (Amaranthus spp.) and Their Wild Relatives in Mexico

In a global climate change scenario, climate-resilient varieties are needed. This research aimed to identify possible shifts in suitable zones of adaptation of two predominant cultivated Mexican species of amaranth (Amaranthus cruentus and A. hypochondriacus) and their two possible wild relatives (A. hybridus and A. powellii) using DIVA-GIS with two different climatic databases. The first model used a current climatic database named WorldClim. The second model accounted for global climate change to obtain the predicted future suitable regions of adaptation using a CCM3 model. This research was performed to elucidate the best possible suitable areas to establish plantations and in vivo germplasm banks, and to predict migration pattern of these species. Our research suggests that the cultivated species of amaranth and their wild relatives had differences among their current and predicted suitable regions of adaptation. Temperature would be the main climatic variable affected. In predicted model, all species examined were more frequent in regions with higher temperature when compared to the current model. However, only A. powellii had a change in annual precipitation frequency in both models. A. hybridus could be a potential new genetic source for amaranth adaptation to potential climatic regions because it showed wider adaptation than the cultivated species.

Burkholderia ambifaria and B. caribensis promote growth and increase yield in grain amaranth (Amaranthus cruentus and A. hypochondriacus) by improving plant nitrogen uptake.

Grain amaranth is an emerging crop that produces seeds having high quality protein with balanced amino-acid content. However, production is restricted by agronomic limitations that result in yields that are lower than those normally produced by cereals. In this work, the use of five different rhizobacteria were explored as a strategy to promote growth and yields in Amaranthus hypochondriacus cv. Nutrisol and A. cruentus cv. Candil, two commercially important grain amaranth cultivars. The plants were grown in a rich substrate, high in organic matter, nitrogen (N), and phosphorus (P) and under greenhouse conditions. Burkholderia ambifaria Mex-5 and B. caribensis XV proved to be the most efficient strains and significantly promoted growth in both grain amaranth species tested. Increased grain yield and harvest index occurred in combination with chemical fertilization when tested in A. cruentus. Growth-promotion and improved yields correlated with increased N content in all tissues examined. Positive effects on growth also occurred in A. cruentus plants grown in a poor soil, even after N and P fertilization. No correlation between non-structural carbohydrate levels in roots of inoculated plants and growth promotion was observed. Conversely, gene expression assays performed at 3-, 5- and 7-weeks after seed inoculation in plants inoculated with B. caribensis XV identified a tissue-specific induction of several genes involved in photosynthesis, sugar- and N- metabolism and transport. It is concluded that strains of Burkholderia effectively promote growth and increase seed yields in grain amaranth. Growth promotion was particularly noticeable in plants grown in an infertile soil but also occurred in a well fertilized rich substrate. The positive effects observed may be attributed to a bio-fertilization effect that led to increased N levels in roots and shoots. The latter effect correlated with the differential induction of several genes involved in carbon and N metabolism and transport.

Nutritional Yield of African Indigenous Vegetables in Water-Deficient and Water-Sufficient Conditions

Water scarcity in certain regions of sub-Saharan Africa impacts agricultural production while prolonging dry seasons, and contributing to food insecurity and malnutrition. The objectives of the study were to evaluate the nutritional yield (edible yield × nutritional content) of indigenous leafy vegetables commonly consumed in Africa under water-stressed conditions. Two species of amaranth, two species of African nightshade, and two varieties of Ethiopian kale were examined under environment-controlled water-sufficient and water-deficient conditions. Fresh and dry stem and leaf weights significantly decreased in the three crop groups under the water-deficient treatment. Nutritional values for carotenoids, tocopherols, calcium and zinc showed significant differences at the content level among treatments. Among the 6 crops evaluated, the amaranth species named Amaranthus hypochondriacus and the African nightshade species named Solanum scabrum had the lowest nutrient losses in drought conditions.

PROPERTIES OF STAINING EXTRACT FROM IN FLORESCENCES AND LEAVES OF AMARANTH AND ITS APPLICATION POTENTIAL

The peculiarities of accumulation and sustainability of the betacyanins's pigment of antioxidant Amaranthin in the inflorescences of different species of amaranth have been discussed. The method of extraction of the betacyanins's staining pigment from inflorescences of amaranth (cv. Valentina) has been proposed. The optimal storage conditions of the betacyanins's staining pigment have been found.

Relationships between the Weedy Amaranthus hybridus (Amaranthaceae) and the Grain Amaranths

ABSTRACTPrevious phylogenetic work investigating the origins of the grain amaranths is plagued by two methodological faults. Primarily, no study to date has adequately sampled all of the grain species as well as the two putative weedy progenitors across their entire native species’ ranges. Second, no study to date adequately investigates the genetic diversity component of both putative weedy species (Amaranthus hybridus L. and Amaranthus quitensis Kunth). Two hundred fifty‐eight individuals from 56 taxa representing the three grain amaranth species and two putative weedy progenitors were sampled from their native species’ ranges and subjected to microsatellite testing. Data were analyzed using principal component analysis, clustering algorithms, and genetic distance neighbor‐joining tree dendrograms. Standard genetic diversity measures indicate strong inbreeding coefficients in all studied taxa. Principal component analysis, clustering algorithms, private alleles, and tree dendrograms all resolve a distinct A. quitensis group. The grain amaranths and A. hybridus were found to group together. Results from 11 simple sequence repeat loci indicate the following: (i) A. hybridus is the progenitor of all three grain amaranths in at least two separate domestication events, (ii) A. quitensis is its own species and not a variant of A. hybridus, and (iii) Amaranthus caudatus L. and Amaranthus hypochondriacus L. appear to be closely related.

Genetic analysis of three Amaranth species using ISSR markers

Genetic analysis of three Amaranth species using ISSR markers

Molecular characterization and expression analysis of a gene encoding an isoamylase-type starch debranching enzyme 3 (ISA3) in grain amaranths

A cDNA clone from amaranth perisperm that encodes an isoamylase (ISA)-type starch debranching enzyme 3 was isolated and analyzed for the first time. The cDNA consisted of 2,715 bp with a single open reading frame of 2,346 bp, encoding a protein of 781 amino acid residues. The deduced amino acid sequence of CrISA3 shared 63–71 % identity with those of other plant ISA3s. We also investigated the genetic diversity of ISA3 in three species of grain amaranth. A comparison of their ISA3 coding sequences revealed an extremely high level of conservation and only 11 single nucleotide polymorphisms were detected. The expression of the CrISA3 gene in amaranth developmental seeds and several tissues was investigated by qRT-PCR analysis. The results showed that CrISA3 was rapidly expressed at the early stage during seed maturation. It was also expressed in non-storage tissues (leaf, petiole, stem, and root) as well as in storage tissue. This observation demonstrates that CrISA3 may play an important role in perisperm starch accumulation at the early developmental stages. In addition, our results indicate that CrISA3 plays important roles in the synthesis of storage and transitory starches. The characterization of the CrISA3 gene will contribute to further studies on starch biosynthesis in Amaranthus.