Storage Root Starch Research Articles

Molecular Cloning, Expression Analysis, and Functional Analysis of Nine IbSWEETs in Ipomoea batatas (L.) Lam

Sugar Will Eventually be Exported Transporter (SWEET) genes play an important regulatory role in plants’ growth and development, stress response, and sugar metabolism, but there are few reports on the role of SWEET proteins in sweet potato. In this study, nine IbSWEET genes were obtained via PCR amplification from the cDNA of sweet potato. Phylogenetic analysis showed that nine IbSWEETs separately belong to four clades (Clade I~IV) and contain two MtN3/saliva domains or PQ-loop superfamily and six~seven transmembrane domains. Protein interaction prediction showed that seven SWEETs interact with other proteins, and SWEETs interact with each other (SWEET1 and SWEET12; SWEET2 and SWEET17) to form heterodimers. qRT-PCR analysis showed that IbSWEETs were tissue-specific, and IbSWEET1b was highly expressed during root growth and development. In addition to high expression in leaves, IbSWEET15 was also highly expressed during root expansion, and IbSWEET7, 10a, 10b, and 12 showed higher expression in the leaves. The expression of SWEETs showed a significant positive/negative correlation with the content of soluble sugar and starch in storage roots. Under abiotic stress treatment, IbSWEET7 showed a strong response to PEG treatment, while IbSWEET10a, 10b, and 12 responded significantly to 4 °C treatment and, also, at 1 h after ABA, to NaCl treatment. A yeast mutant complementation assay showed that IbSWEET7 had fructose, mannose, and glucose transport activity; IbSWEET15 had glucose transport activity and weaker sucrose transport activity; and all nine IbSWEETs could transport 2-deoxyglucose. These results provide a basis for further elucidating the functions of SWEET genes and promoting molecular breeding in sweet potato.

Polyploid GWAS reveals the basis of molecular marker development for complex breeding traits including starch content in the storage roots of sweet potato.

Given the importance of prioritizing genome-based breeding of sweet potato to enable the promotion of food and nutritional security for future human societies, here, we aimed to dissect the genetic basis of storage root starch content (SC) when associated with a complex set of breeding traits including dry matter (DM) rate, storage root fresh weight (SRFW), and anthocyanin (AN) content in a mapping population containing purple-fleshed sweet potato. A polyploid genome-wide association study (GWAS) was extensively exploited using 90,222 single-nucleotide polymorphisms (SNPs) obtained from a bi-parental 204 F1 population between 'Konaishin' (having high SC but no AN) and 'Akemurasaki' (having high AN content but moderate SC). Through the comparison of polyploid GWAS on the whole set of the 204 F1, 93 high-AN-containing F1, and 111 low-AN-containing F1 populations, a total of two (consists of six SNPs), two (14 SNPs), four (eight SNPs), and nine (214 SNPs) significantly associated signals were identified for the variations of SC, DM, SRFW, and the relative AN content, respectively. Of them, a novel signal associated with SC, which was most consistent in 2019 and 2020 in both the 204 F1 and 111 low-AN-containing F1 populations, was identified in homologous group 15. The five SNP markers associated with homologous group 15 could affect SC improvement with a degree of positive effect (~4.33) and screen high-starch-containing lines with higher efficiency (~68%). In a database search of 62 genes involved in starch metabolism, five genes including enzyme genes granule-bound starch synthase I (IbGBSSI), α-amylase 1D, α-amylase 1E, and α-amylase 3, and one transporter gene ATP/ADP-transporter were located on homologous group 15. In an extensive qRT-PCR of these genes using the storage roots harvested at 2, 3, and 4 months after field transplantation in 2022, IbGBSSI, which encodes the starch synthase isozyme that catalyzes the biosynthesis of amylose molecule, was most consistently elevated during starch accumulation in sweet potato. These results would enhance our understanding of the underlying genetic basis of a complex set of breeding traits in the starchy roots of sweet potato, and the molecular information, particularly for SC, would be a potential platform for molecular marker development for this trait.

MeGATAs, functional generalists in interactions between cassava growth and development, and abiotic stresses.

The proteins with DNA-binding preference to the consensus DNA sequence (A/T) GATA (A/G) belong to a GATA transcription factor family, with a wide array of biological processes in plants. Cassava (Manihot esculenta) is an important food crop with high production of starch in storage roots. Little was however known about cassava GATA domain-containing genes (MeGATAs). Thirty-six MeGATAs, MeGATA1 to MeGATA36, were found in this study. Some MeGATAs showed a collinear relationship with orthologous genes of Arabidopsis, poplar and potato, rice, maize and sorghum. Eight MeGATA-encoded proteins (MeGATAs) analysed were all localized in the nucleus. Some MeGATAs had potentials of binding ligands and/or enzyme activity. One pair of tandem-duplicated MeGATA17-MeGATA18 and 30 pairs of whole genome-duplicated MeGATAs were found. Fourteen MeGATAs showed low or no expression in the tissues. Nine analysed MeGATAs showed expression responses to abiotic stresses and exogenous phytohormones. Three groups of MeGATA protein interactions were found. Fifty-three miRNAs which can target 18 MeGATAs were identified. Eight MeGATAs were found to target other 292 cassava genes, which were directed to radial pattern formation and phyllome development by gene ontology enrichment, and autophagy by Kyoto Encyclopaedia of Genes and Genomes enrichment. These data suggest that MeGATAs are functional generalists in interactions between cassava growth and development, abiotic stresses and starch metabolism.

Cassava shrunken-2 homolog MeAPL3 determines storage root starch and dry matter content and modulates storage root postharvest physiological deterioration

Key messageAmong the five cassava isoforms (MeAPL1–MeAPL5), MeAPL3 is responsible for determining storage root starch content. Degree of storage root postharvest physiological deterioration (PPD) is directly correlated with starch content.AGPase is heterotetramer composed of two small and two large subunits each coded by small gene families in higher plants. Studies in cassava (Manihot esculenta) identified and characterized five isoforms of Manihot esculenta ADP-glucose pyrophosphorylase large subunit (MeAPL1–MeAPL5) and employed virus induced gene silencing (VIGS) to show that MeAPL3 is the key isoform responsible for starch and dry matter accumulation in cassava storage roots. Silencing of MeAPL3 in cassava through stable transgenic lines resulted in plants displaying significant reduction in storage root starch and dry matter content (DMC) and induced a distinct phenotype associated with increased petiole/stem angle, resulting in a droopy leaf phenotype. Plants with reduced starch and DMC also displayed significantly reduced or no postharvest physiological deterioration (PPD) compared to controls and lines with high DMC and starch content. This provides strong evidence for direct relationships between starch/dry matter content and its role in PPD and canopy architecture traits in cassava.

A novel effect of glycine on the growth and starch biosynthesis of storage root in sweetpotato (Ipomoea batatas Lam.)

Sweetpotato (Ipomoea batatas Lam.) plays an indispensable role in feed, starch-based industries and ethanol biofuel production. Few studies have investigated on how external amino acids affect the growth and production of sweetpotato. In the study, we evaluated morphological, physiological and molecular effects of external glycine (Gly) on the root growth and starch metabolism of sweetpotato, Xushu16. At morphological level, the Xushu16 with Gly stimuli had larger plant biomass than that under control condition. At physiological level, the photosynthesis strength of the Xushu16 with Gly treatments showed significant differences relative to those under control condition. The relative content of plant hormone and starch in storage roots was higher under Gly conditions than that under control condition. At molecular level, a total of 4836 differentially expression genes were identified in the storage roots with different Gly treatments by RNA-Seq. Among them, as many as 1830 genes were involved in carbohydrate metabolism, which held maximum proportion among all the DEGs. Further, a few genes involved in starch biosynthesis were proved to be Gly-induced significantly by RT-qPCR. All the results indicated extrinsic Gly promotes the growth of storage roots by strengthening photosynthesis and increasing plant hormone, and enhances starch biosynthesis of storage roots by accelerating carbohydrate metabolism and regulating the expression of starch-related genes.

Production of very-high-amylose cassava by post-transcriptional silencing of branching enzyme genes.

High amylose starch can be produced by plants deficient in the function of branching enzymes (BEs). Here we report the production of transgenic cassava (Manihot esculenta Crantz) with starches containing up to 50% amylose due to the constitutive expression of hair-pin dsRNAs targeting the BE1 or BE2 genes. All BE1-RNAi plant lines (BE1i) and BE2-RNAi plant lines (BE2i) were grown up in the field, but with reduced total biomass production. Considerably high amylose content in the storage roots of BE2i plant lines was achieved. Storage starch granules of BE1i and BE2i plants had similar morphology as wild type (WT), however, the size of BE1i starch granules were bigger than that of WT. Comparisons of amylograms and thermograms of all three sources of storage starches revealed dramatic changes to the pasting properties and a higher melting temperature for BE2i starches. Glucan chain length distribution analysis showed a slight increase in chains of DP>36 in BE1i lines and a dramatic increase in glucan chains between DP 10-20 and DP>40 in BE2i lines. Furthermore, BE2i starches displayed a B-type X-ray diffraction pattern instead of the A-type pattern found in BE1i and WT starches. Therefore, cassava BE1 and BE2 function differently in storage root starch biosynthesis.

Modification of Cassava Root Starch Phosphorylation Enhances Starch Functional Properties.

Cassava (Manihot esculenta Crantz) is a root crop used as a foodstuff and as a starch source in industry. Starch functional properties are influenced by many structural features including the relative amounts of the two glucan polymers amylopectin and amylose, the branched structure of amylopectin, starch granule size and the presence of covalent modifications. Starch phosphorylation, where phosphates are linked either to the C3 or C6 carbon atoms of amylopectin glucosyl residues, is a naturally occurring modification known to be important for starch remobilization. The degree of phosphorylation has been altered in several crops using biotechnological approaches to change expression of the starch-phosphorylating enzyme GLUCAN WATER DIKINASE (GWD). Interestingly, this frequently alters other structural features of starch beside its phosphate content. Here, we aimed to alter starch phosphorylation in cassava storage roots either by manipulating the expression of the starch phosphorylating or dephosphorylating enzymes. Therefore, we generated transgenic plants in which either the wild-type potato GWD (StGWD) or a redox-insensitive version of it were overexpressed. Further plants were created in which we used RNAi to silence each of the endogenous phosphoglucan phosphatase genes STARCH EXCESS 4 (MeSEX4) and LIKE SEX4 2 (MeLSF), previously discovered by analyzing leaf starch metabolism in the model species Arabidopsis thaliana. Overexpressing the potato GWD gene (StGWD), which specifically phosphorylates the C6 position, increased the total starch-bound phosphate content at both the C6 and the C3 positions. Silencing endogenous LSF2 gene (MeLSF2), which specifically dephosphorylates the C3 position, increased the ratio of C3:C6 phosphorylation, showing that its function is conserved in storage tissues. In both cases, other structural features of starch (amylopectin structure, amylose content and starch granule size) were unaltered. This allowed us to directly relate the physicochemical properties of the starch to its phosphate content or phosphorylation pattern. Starch swelling power and paste clarity were specifically influenced by total phosphate content. However, phosphate position did not significantly influence starch functional properties. In conclusion, biotechnological manipulation of starch phosphorylation can specifically alter certain cassava storage root starch properties, potentially increasing its value in food and non-food industries.

Genetic Variability for Yield and Nutritional Quality in Yam Bean (Pachyrhizus sp.)

The amount of genotypic and phenotypic variability that exists in a species is important for selection and initiating breeding programs. Yam bean is grown locally in tropical countries of the Americas and Asia for their tasty storage roots, which usually have low dry matter content. The crop was recently introduced in Uganda and other East and Central African countries to supplement iron (Fe) and protein content in diets. This study aimed to estimate genetic variability for root yield and quality traits among 26 yam bean accessions in Uganda. A randomized complete block design was used with two replications across two ecogeographical locations and two seasons during 2012 and 2013. Near-infrared reflectance spectroscopy (NIRS) was used to determine quality of storage root samples. Significant differences among genotypes were observed for all traits except root protein, zinc (Zn), and phosphorus contents. Genotypic variance components () were significant for storage root fresh yield (SRFY), storage root dry matter (SRDM), storage root dry yield (SRDY), vine yield (VNY), fresh biomass yield (FBY), and storage root starch (STA) and Fe contents. For traits with significant the broad sense heritability estimates ranged from 58.4% for SRDY to 83.6% for FBY; and phenotypic coefficients of variation were high for SRFY (66%), SRDY (53.3%), VNY (60.5%), and FBY (59%), but low to medium for SRDM (22.6%), STA (15.1%), and Fe (21.3%). Similarly, the genotypic coefficients of variation were high for SRFY (56.7%), SRDY (53.3%), VNY (55%), and FBY (53.9%); and low for SRDM (20%), STA (12.4%), and Fe (17.8%). There were strong positive correlations between SRFY and both SRDY (r = 0.926) and FBY (r = 0.962), but low-to-moderate correlations among quality traits. It should be possible to breed for high dry matter yam beans by using low dry matter accessions due to the observed genetic variation ( = 9.3%2), which is important if the high dry matter Pachyrhizus tuberosus accessions (known as chuin) from Peru cannot be accessed. This study indicated substantial genetic variation for yield and quality traits in yam bean, demonstrating potential for adaptability to growing conditions and consumer needs in East and Central Africa and for genetic improvement through selection.

Differential Expression of Genes Involved in Sucrose Synthesis in Source and Sink Organs of Cassava Plants Undergoing Seasonal Drought Stress

Sucrose functions as a regulator of gene expression allowing plants to respond to cellular energy demands during both normal and stress conditions. Previous studies indicated that drought followed with onset of rain could negatively affect yield of starch in storage roots of cassava. In this study, molecular approaches were conducted to investigate expression profiling of sucrose phosphate synthase (SPS), sucrose phosphatase (SPP) and 14-3-3, which are importance as regulatory points to modulate availability of sucrose in cassava. In source organs, expression of cassava (Manihot esculenta Crantz.) sucrose phosphate synthase (MeSPS) and cassava sucrose phosphatase (MeSPP) varied depending on developmental stage and day/night cycle. In sink organ, the levels of MeSPS, MeSPP and Me14-3-3 transcripts were affected to a much greater degree by different planting season than by root developmental stage. In response to fluctuating level of rainfall, a correlation between MeSPS and Me14-3-3 expression patterns was observed. This finding suggests the role of these enzymes and the importance of sucrose metabolism in seasonal drought stress response in cassava. Given the identified new role for cassava as an important biofuel crop as well as its current usage as a food crop, understanding on the most appropriate time to plant and harvest cassava in order to maximize starch quantity and quality is warranted.