Identification and Fine Mapping of the Recessive Gene BK-5, Which Affects Cell Wall Biosynthesis and Plant Brittleness in Maize.

The cellulose of the plant cell wall indirectly affects the cell shape and straw stiffness of the plant. Here, the novel brittleness mutant brittle stalk-5 (bk-5) of the maize inbred line RP125 was characterized. We found that the mutant displayed brittleness of the stalk and even the whole plant, and that the brittleness phenotype existed during the whole growth period from germination to senescence. The compressive strength was reduced, the cell wall was thinner, and the cellulose content was decreased compared to that of the wild type. Genetic analysis and map-based cloning indicated that bk-5 was controlled by a single recessive nuclear gene and that it was located in a 90.2-Kb region on chromosome 3 that covers three open reading frames (ORFs). Sequence analysis revealed a single non-synonymous missense mutation, T-to-A, in the last exon of Zm00001d043477 (B73: version 4, named BK-5) that caused the 951th amino acid to go from leucine to histidine. BK-5 encodes a cellulose synthase catalytic subunit (CesA), which is involved with cellulose synthesis. We found that BK-5 was constitutively expressed in all tissues of the germinating stage and silking stage, and highly expressed in the leaf, auricula, and root of the silking stage and the 2-cm root and bud of the germinating stage. We found that BK-5 mainly localized to the Golgi apparatus, suggesting that the protein might move to the plasma membrane with the aid of Golgi in maize. According to RNA-seq data, bk-5 had more downregulated genes than upregulated genes, and many of the downregulated genes were enzymes and transcription factors related to cellulose, hemicellulose, and lignin biosynthesis of the secondary cell wall. The other differentially expressed genes were related to metabolic and cellular processes, and were significantly enriched in hormone signal transduction, starch and sucrose metabolism, and the plant-pathogen interaction pathway. Taken together, we propose that the mutation of gene BK-5 causes the brittle stalk phenotype and provides important insights into the regulatory mechanism of cellulose biosynthesis and cell wall development in maize.

Population-tailored mock genome enables genomic studies in species without a reference genome.

Based on molecular markers, genomic prediction enables us to speed up breeding schemes and increase the response to selection. There are several high-throughput genotyping platforms able to deliver thousands of molecular markers for genomic study purposes. However, even though its widely applied in plant breeding, species without a reference genome cannot fully benefit from genomic tools and modern breeding schemes. We used a method to assemble a population-tailored mock genome to call single-nucleotide polymorphism (SNP) markers without an available reference genome, and for the first time, we compared the results with standard genotyping platforms (array and genotyping-by-sequencing (GBS) using a reference genome) for performance in genomic prediction models. Our results indicate that using a population-tailored mock genome to call SNP delivers reliable estimates for the genomic relationship between genotypes. Furthermore, genomic prediction estimates were comparable to standard approaches, especially when considering only additive effects. However, mock genomes were slightly worse than arrays at predicting traits influenced by dominance effects, but still performed as well as standard GBS methods that use a reference genome. Nevertheless, the array-based SNP markers methods achieved the best predictive ability and reliability to estimate variance components. Overall, the mock genomes can be a worthy alternative for genomic selection studies, especially for those species where the reference genome is not available.

Genome-Wide Identification and Characterization of Polygalacturonase Gene Family in Maize (Zea mays L.).

Polygalacturonase (PG, EC 3.2.1.15) is a crucial enzyme for pectin degradation and is involved in various developmental processes such as fruit ripening, pollen development, cell expansion, and organ abscission. However, information on the PG gene family in the maize (Zea mays L.) genome and the specific members involved in maize anther development are still lacking. In this study, we identified 55 PG family genes from the maize genome and further characterized their evolutionary relationship and expression patterns. Phylogenetic analysis revealed that ZmPGs are grouped into six Clades, and gene structures of the same Clade are highly conserved, suggesting their functional conservation. The ZmPGs are randomly distributed across maize chromosomes, and collinearity analysis showed that many ZmPGs might be derived from tandem duplications and segmental duplications, and these genes are under purifying selection. Furthermore, gene expression analysis provided insights into possible functional divergence among ZmPGs. Based on the RNA-seq data analysis, we found that many ZmPGs are expressed in various tissues while 18 ZmPGs are highly expressed in maize anther, and their detailed expression profiles in different anther developmental stages were further investigated by using RT-qPCR analysis. These results provide valuable information for further functional characterization and application of the ZmPGs in maize.

Chromosome-level genome assembly of a regenerable maize inbred line A188.

BACKGROUND: The maize inbred line A188 is an attractive model for elucidation of gene function and improvement due to its high embryogenic capacity and many contrasting traits to the first maize reference genome, B73, and other elite lines. The lack of a genome assembly of A188 limits its use as a model for functional studies. RESULTS: Here, we present a chromosome-level genome assembly of A188 using long reads and optical maps. Comparison of A188 with B73 using both whole-genome alignments and read depths from sequencing reads identify approximately 1.1 Gb of syntenic sequences as well as extensive structural variation, including a 1.8-Mb duplication containing the Gametophyte factor1 locus for unilateral cross-incompatibility, and six inversions of 0.7 Mb or greater. Increased copy number of carotenoid cleavage dioxygenase 1 (ccd1) in A188 is associated with elevated expression during seed development. High ccd1 expression in seeds together with low expression of yellow endosperm 1 (y1) reduces carotenoid accumulation, accounting for the white seed phenotype of A188. Furthermore, transcriptome and epigenome analyses reveal enhanced expression of defense pathways and altered DNA methylation patterns of the embryonic callus. CONCLUSIONS: The A188 genome assembly provides a high-resolution sequence for a complex genome species and a foundational resource for analyses of genome variation and gene function in maize. The genome, in comparison to B73, contains extensive intra-species structural variations and other genetic differences. Expression and network analyses identify discrete profiles for embryonic callus and other tissues.

Functional Characterization of an Anthocyanin Dimalonyltransferase in Maize.

Anthocyanins are pigments with appealing hues that are currently being used as sources of natural colorants. The interaction of acylation on the stability of anthocyanin molecules has long been known. Maize is an abundant source of malonylglucoside and dimalonylglucoside anthocyanins. The enzyme Aat1 is an anthocyanin acyltransferase known to synthesize the majority of acylated anthocyanins in maize. In this paper, we characterize the substrate specificity and reaction kinetics of Aat1. It was found that Aat1 has anthocyanin 3-O-glucoside dimalonyltransferase activity and is only the second enzyme of this type characterized to this date. Our results indicate that Aat1 can utilize malonyl-CoA; succinyl-CoA and every anthocyanin 3-O-glucoside tested. Results of this study provide insight into the structure-function relations of dimalonyltransferases and give a unique insight into the activity of monocot anthocyanin acyltransferases.

Genetic diversity and population structure of early and extra-early maturing maize germplasm adapted to sub-Saharan Africa.

BACKGROUND: Assessment and effective utilization of genetic diversity in breeding programs is crucial for sustainable genetic improvement and rapid adaptation to changing breeding objectives. During the past two decades, the commercialization of the early and extra-early maturing cultivars has contributed to rapid expansion of maize into different agro-ecologies of sub-Saharan Africa (SSA) where maize has become an important component of the agricultural economy and played a vital role in food and nutritional security. The present study aimed at understanding the population structure and genetic variability among 439 early and extra-early maize inbred lines developed from three narrow-based and twenty-seven broad-based populations by the International Iinstitute of Tropical Agriculture Maize Improvement Program (IITA-MIP). These inbreds were genotyped using 9642 DArTseq-based single nucleotide polymorphism (SNP) markers distributed uniformly throughout the maize genome. RESULTS: About 40.8% SNP markers were found highly informative and exhibited polymorphic information content (PIC) greater than 0.25. The minor allele frequency and PIC ranged from 0.015 to 0.500 and 0.029 to 0.375, respectively. The STRUCTURE, neighbour-joining phylogenetic tree and principal coordinate analysis (PCoA) grouped the inbred lines into four major classes generally consistent with the selection history, ancestry and kernel colour of the inbreds but indicated a complex pattern of the genetic structure. The pattern of grouping of the lines based on the STRUCTURE analysis was in concordance with the results of the PCoA and suggested greater number of sub-populations (K = 10). Generally, the classification of the inbred lines into heterotic groups based on SNP markers was reasonably reliable and in agreement with defined heterotic groups of previously identified testers based on combining ability studies. CONCLUSIONS: Complete understanding of potential heterotic groups would be difficult to portray by depending solely on molecular markers. Therefore, planned crosses involving representative testers from opposing heterotic groups would be required to refine the existing heterotic groups. It is anticipated that the present set of inbreds could contribute new beneficial alleles for population improvement, development of hybrids and lines with potential to strengthen future breeding programs. Results of this study would help breeders in formulating breeding strategies for genetic enhancement and sustainable maize production in SSA.

Molecular characterization, expression and functional analysis of acyl-CoA-binding protein gene family in maize (Zea mays).

BACKGROUND: Acyl-CoA-binding proteins (ACBPs) possess a conserved acyl-CoA-binding (ACB) domain that facilitates binding to acyl-CoA esters and trafficking in eukaryotic cells. Although the various functions of ACBP have been characterized in several plant species, their structure, molecular evolution, expression profile, and function have not been fully elucidated in Zea mays L. RESULTS: Genome-wide analysis identified nine ZmACBP genes in Z. mays, which could be divided into four distinct classes (class I, class II, class III, and class IV) via construction of a phylogenetic tree that included 48 ACBP genes from six different plant species. Transient expression of a ZmACBP-GFP fusion protein in tobacco (Nicotiana tabacum) epidermal cells revealed that ZmACBPs localized to multiple different locations. Analyses of expression profiles revealed that ZmACBPs exhibited temporal and spatial expression changes during abiotic and biotic stresses. Eight of the nine ZmACBP genes were also found to have significant association with agronomic traits in a panel of 500 maize inbred lines. The heterologous constitutive expression of ZmACBP1 and ZmACBP3 in Arabidopsis enhanced the resistance of these plants to salinity and drought stress, possibly through alterations in the level of lipid metabolic and stress-responsive genes. CONCLUSION: The ACBP gene family was highly conserved across different plant species. ZmACBP genes had clear tissue and organ expression specificity and were responsive to both biotic and abiotic stresses, suggesting their roles in plant growth and stress resistance.

A cost-effective maize ear phenotyping platform enables rapid categorization and quantification of kernels.

High-throughput phenotyping systems are powerful, dramatically changing our ability to document, measure, and detect biological phenomena. Here, we describe a cost-effective combination of a custom-built imaging platform and deep-learning-based computer vision pipeline. A minimal version of the maize (Zea mays) ear scanner was built with low-cost and readily available parts. The scanner rotates a maize ear while a digital camera captures a video of the surface of the ear, which is then digitally flattened into a two-dimensional projection. Segregating GFP and anthocyanin kernel phenotypes are clearly distinguishable in ear projections and can be manually annotated and analyzed using image analysis software. Increased throughput was attained by designing and implementing an automated kernel counting system using transfer learning and a deep learning object detection model. The computer vision model was able to rapidly assess over 390 000 kernels, identifying male-specific transmission defects across a wide range of GFP-marked mutant alleles. This includes a previously undescribed defect putatively associated with mutation of Zm00001d002824, a gene predicted to encode a vacuolar processing enzyme. Thus, by using this system, the quantification of transmission data and other ear and kernel phenotypes can be accelerated and scaled to generate large datasets for robust analyses.

Identification of the variety of maize seeds based on hyperspectral images coupled with convolutional neural networks and subregional voting.

BACKGROUND: Maize is one of the most important food crops in the world. Many different varieties of maize seeds are similar in size and appearance, so distinguishing the varieties of maize seed is a significant research topic. This study used hyperspectral image processing coupled with convolutional neural network (CNN) and a subregional voting method to recognize different varieties of maize seed. RESULTS: First, visible and near-infrared (NIR-visible) hyperspectral images were obtained. Savitzky-Golay (SG) smoothing and first derivative (FD) were used to pretreat the raw spectra and highlight the spectral differences of samples of different varieties. Second, the region of interest (ROI) of each sample was divided into several subregions according to the shape and the number of pixels. Then, a method was proposed for reshaping the images of pixel spectra for the CNN and the training model was established. Finally, using subregional voting, one prediction result was generated from the prediction results of several original subregions in one sample. The results showed that, for six varieties of normal maize seeds, the tests identified embryoid and non-embryoid forms with 93.33% and 95.56% accuracy, respectively. For six varieties of sweet maize seeds, the test accuracy in embryoid and non-embryoid forms was 97.78% and 98.15%, respectively. CONCLUSION: The maize seed was identified accurately. The present study demonstrated that the CNN model for spectral image coupled with subregional voting represents a new approach for the identification of varieties of maize seed. © 2021 Society of Chemical Industry.

Starch physicochemical properties of double recessive sweet-waxy maize.

Sweet-waxy is a new type of maize with waxy and sugary double recessive genes. This study aims to clarify starch structural and functional properties of this maize type. Grains with sweet-waxy and waxy phenotypes were separated from an ear using the two sweet-waxy maize hybrids of ATN and NKY as materials. Compared with waxy maize starch, the sweet-waxy maize starch mainly comprises small-sized round granules despite the typical waxy character of both starches. Mw, Mn, and relative crystallinity of sweet-waxy starch were higher than those of waxy starch in both hybrids. The average chain length of waxy starch was higher in ATN but lower in NKY compared with that of sweet-waxy starch. However, polydispersity (Mw/Mn) and F1 fraction were high in sweet-waxy and waxy starches in ATN and NKY, respectively. Breakdown viscosity, gelatinization enthalpy and temperatures of both hybrids were low in sweet-waxy starch. Peak viscosity was higher in waxy starch in NKY and similar between sweet-waxy and waxy starches in ATN. Retrogradation percentage was high and low for sweet-waxy starches in ATN and NKY, respectively.

Identification of the volatile profiles of 22 traditional and newly bred maize varieties and their porridges by PTR-QiTOF-MS and HS-SPME GC-MS.

BACKGROUND: Low adoption of maize varieties bred to address the nutritional needs of the growing African population limits their impact. Aroma is essential in consumer preference, but has hitherto hardly been studied. We analysed the volatile organic compounds of flours and porridges of 22 maize varieties belonging to four nutritionally distinct groups, namely provitamin A maize, quality protein maize, yellow and white maize. RESULTS: Proton-transfer-reaction quadrupole ion time-of-flight mass spectrometry (PTR-QiTOF-MS) analysis generated 524 mass peaks ranging from 16.007 to 448.089 m/z. Principal component analysis separated the varieties belonging to the four groups. With headspace solid-phase microextraction gas chromatography-mass spectrometry (HS-SPME GC-MS), 48 volatile compounds were identified in maize flour and 21 in maize porridge, including hexane, nonane, pentanoic acid, 1-octen-3-ol, 1-hexanol, hexanal, nonanal, 2-pentylfuran and 2-heptanone. Volatile compounds such as 1,2,4-trimethyl benzene, associated with thermal degradation of carotenoids, increased in the porridge of yellow and provitamin A maize. CONCLUSION: The results indicate that PTR-QiTOF-MS and HS-SPME GC-MS combined with multivariate analysis are instrumental to study the volatile aroma compounds of different maize varieties.

Genetic characterization of maize doubled haploid lines for Fusarium stalk rot caused by Fusarium verticillioides.

Fusarium stalk rot disease (FSR) of maize caused by Fusarium verticillioides (Sacc.) Nirenberg is becoming an important biotic production constraint in many of the major maize growing areas causing substantial yield losses. Inbreds are preferred as parents in hybrid development owing to homozygous nature and high heterotic ability. Double haploid (DH) technology has emerged as a significant milestone. A total of 339 DH lines were generated from two inbred lines, VL1043 (susceptible) and CM212 (resistant), through in vivo haploid induction method. The 339 DH lines along with parents were phenotyped for their response to the FSR at the College of Agriculture, V. C. Farm, Mandya, India during summer, kharif and rabi seasons of the 2019-2020. Best linear unbiased predictors (BLUPs) were estimated for the FSR disease scores over three seasons. Awide range of BLUP scores of three to nine indicated the presence of higher variation for response of DH lines to FSR disease. The higher estimates of standardized range (1.31) and phenotypic coefficient of variation (19.80) also displayed higher variability. Nine lines were moderately resistant and 188 exhibited moderately susceptible reaction. The distribution of DH lines was positively skewed (1.34) and platykurtic (2.31) which suggested complementary epistasis and involvement of large number of genes in the disease expression.

Genetic analysis of maydis leaf blight resistance in subtropical maize (Zea mays L.) germplasm.

Knowledge on the genetics of maydis leaf blight (MLB) is crucial to breed the resistant maize cultivars to combat disease epidemics as a sustainable and cost-effective approach. The present investigation was framed to understand the genetics of MLB resistance in subtropical maize. Two contrasting genotypes CM119 (susceptible) and SC-7-2-1-2-6-1 (resistant) were used to generate six genetic populations, namely P1, P2, F1, F2, BC1P1 and BC1P2, and evaluated in three target environments for MLB resistance under artificial epiphytotic condition. The CM119 and SC-7-2-1-2-6-1 showed susceptible and resistant reactions with mean disease reaction of 3.89-3.98 and 1.88-2.00, respectively. The derived generations, namely F1, F2, BC1P1 and BC1P2 showed mean disease reaction of 2.15-2.28, 2.44-2.51, 2.19-2.24 and 2.22-2.28, respectively in the test locations. The segregating generations (F2: 0.35-0.37; BC1P1: 0.24-0.29 and BC1P2: 0.17-0.20) showed variation for MLB disease resistance over the parental and first filial generations (P1: 0.11-0.17; P2: 0.08-0.13 and F1: 0.12-0.14). The genetic analysis of MLB resistance revealed the nonallelic interactions of duplicate epistasis type across the test locations. Among the gene interactions, dominance x dominance [l] effect was predominant over additive x additive [i] and additive x dominance [j] effects. The segregation analysis and the prediction of the number of major loci revealed at least two major genes associated with MLB tolerance in subtropical maize. Our investigation paved the foundation for the improvement of subtropical maize germplasm of MLB resistance.

Analysis of metabolites and metabolic pathways in three maize (Zea mays L.) varieties from the same origin using GC-MS.

Metabolites of the Jinyu 88, Huanong 18, and Demeiya 9 maize varieties from the same origin were isolated and identified by GC-MS, and the specific metabolites and metabolic mechanisms of these three varieties of maize were preliminarily analysed and discussed. The metabolites were extracted with 80% methanol and derived with N,O-bis(trimethylsilyl)trifluoroacetamide. A total of 59 metabolites were identified. The specific metabolites of these three varieties of maize were identified. Four possible unknown-structure metabolites were hypothesized. The results showed that the specific metabolites of JY88 were only involved in fatty acid metabolism. The specific metabolites of HN18 were determined to be involved in fatty acid metabolism, glucose metabolism, and phytosterol metabolism. The specific metabolites of DM9 were observed to participate in glucose metabolism and fatty acid metabolism. The disease resistance of HN18 was higher than that of DM9, and its grain bulk density was higher than that of DM9. JY88 was determined to be significantly different from the other two varieties, and its appearance and disease resistance were worse than those of the other two varieties. The variety with the highest nutritional value was determined to be HN18, and the variety with the lowest nutritional value was JY88. This finding indicated that different maize varieties from the same origin had different metabolites and different metabolic mechanisms, which caused the three maize varieties to exhibit different characteristics and qualities.

Genomic Analysis of Selected Maize Landraces from Sahel and Coastal West Africa Reveals Their Variability and Potential for Genetic Enhancement.

Genetic adaptation of maize to the increasingly unpredictable climatic conditions is an essential prerequisite for achievement of food security and sustainable development goals in sub-Saharan Africa. The landraces of maize; which have not served as sources of improved germplasm; are invaluable sources of novel genetic variability crucial for achieving this objective. The overall goal of this study was to assess the genetic diversity and population structure of a maize panel of 208 accessions; comprising landrace gene pools from Burkina Faso (58), Ghana (43), and Togo (89), together with reference populations (18) from the maize improvement program of the International Institute of Tropical Agriculture (IITA). Genotyping the maize panel with 5974 DArTseq-SNP markers revealed immense genetic diversity indicated by average expected heterozygosity (0.36), observed heterozygosity (0.5), and polymorphic information content (0.29). Model-based population structure; neighbor-joining tree; discriminant analysis of principal component; and principal coordinate analyses all separated the maize panel into three major sub-populations; each capable of providing a wide range of allelic variation. Analysis of molecular variance (AMOVA) showed that 86% of the variation was within individuals; while 14% was attributable to differences among gene pools. The Burkinabe gene pool was strongly differentiated from all the others (genetic differentiation values >0.20), with no gene flow (Nm) to the reference populations (Nm = 0.98). Thus; this gene pool could be a target for novel genetic variation for maize improvement. The results of the present study confirmed the potential of this maize panel as an invaluable genetic resource for future design of association mapping studies to speed-up the introgression of this novel variation into the existing breeding pipelines.

Apoplastic maize fructan exohydrolase Zm-6-FEH displays substrate specificity for levan and is induced by exposure to levan-producing bacteria.

Fructan exohydrolases (FEHs) are structurally related to cell wall invertases. While the latter are ubiquitous in higher plants, the role of FEHs in non-fructan species has remained enigmatic. To explore possible roles of FEHs in maize, a full length putative Zm-6-FEH-encoding cDNA was cloned displaying high sequence similarity with cell wall invertases. For functional characterization, Zm-6-FEH protein was expressed in Picha pastoris and in Nicotiana benthamiana leaves. Enzyme activity of recombinant Zm-6-FEH protein showed a strong preference for levan as substrate. Expression profiling in maize seedlings revealed higher transcript amounts in the more mature leaf parts as compared to the growth zone at the base of the leaf, in good correlation with FEH enzyme activities. Subcellular localization analysis indicated Zm-6-FEH location in the apoplast. Noteworthy, incubation of leaf discs with levan and co-incubation with high levan-producing bacteria selectively up-regulated transcript levels of Zm-6-FEH, accompanied by an increase of 6-FEH enzyme activity. In summary, the results indicate that Zm-6-FEH, a novel fructan exohydrolase of a non-fructan species, may have a role in plant defense against levan-producing bacteria.

The genetic architecture of the maize progenitor, teosinte, and how it was altered during maize domestication.

The genetics of domestication has been extensively studied ever since the rediscovery of Mendel's law of inheritance and much has been learned about the genetic control of trait differences between crops and their ancestors. Here, we ask how domestication has altered genetic architecture by comparing the genetic architecture of 18 domestication traits in maize and its ancestor teosinte using matched populations. We observed a strongly reduced number of QTL for domestication traits in maize relative to teosinte, which is consistent with the previously reported depletion of additive variance by selection during domestication. We also observed more dominance in maize than teosinte, likely a consequence of selective removal of additive variants. We observed that large effect QTL have low minor allele frequency (MAF) in both maize and teosinte. Regions of the genome that are strongly differentiated between teosinte and maize (high FST) explain less quantitative variation in maize than teosinte, suggesting that, in these regions, allelic variants were brought to (or near) fixation during domestication. We also observed that genomic regions of high recombination explain a disproportionately large proportion of heritable variance both before and after domestication. Finally, we observed that about 75% of the additive variance in both teosinte and maize is "missing" in the sense that it cannot be ascribed to detectable QTL and only 25% of variance maps to specific QTL. This latter result suggests that morphological evolution during domestication is largely attributable to very large numbers of QTL of very small effect.

Regulation of Sulfur Homeostasis in Mycorrhizal Maize Plants Grown in a Fe-Limited Environment.

Sulfur is an essential macronutrient for growth of higher plants. The entry of the sulfate anion into the plant, its importation into the plastids for assimilation, its long-distance transport through the vasculature, and its storage in the vacuoles require specific sulfate transporter proteins. In this study, mycorrhizal and non-mycorrhizal maize plants were grown for 60 days in an S-deprived substrate, whilst iron was provided to the plants in the sparingly soluble form of FePO4. On day 60, sulfate was provided to the plants. The gene expression patterns of a number of sulfate transporters as well as sulfate assimilation enzymes were studied in leaves and roots of maize plants, both before as well as after sulfate supply. Prolonged sulfur deprivation resulted in a more or less uniform response of the genes' expressions in the roots of non-mycorrhizal and mycorrhizal plants. This was not the case neither in the roots and leaves after the supply of sulfur, nor in the leaves of the plants during the S-deprived period of time. It is concluded that mycorrhizal symbiosis modified plant demands for reduced sulfur, regulating accordingly the uptake, distribution, and assimilation of the sulfate anion.

Denser Markers and Advanced Statistical Method Identified More Genetic Loci Associated with Husk Traits in Maize.

The husk-the leaf-like outer covering of maize ear-has multiple functions, including protecting the ear from diseases infection and dehydration. In previous studies, we genotyped an association panel of 508 inbred lines genotyped with a total of ~550,000 SNPs (Illumina 50 K SNP Chip and RNA-seq). Genome-Wide Association Studies (GWAS) were conducted on four husk traits: husk length (HL), husk layer number (HN), husk thickness (HT), and husk width (HW). Minimal associations were identified and none of them passed the P-value threshold after a Bonferroni multiple-test correction using a single locus test in framework of mixed linear model. In this study, we doubled the number of SNPs (~1,250,000 in total) by adding GBS and 600 K SNP Chip. GWAS, performed with the recently developed multiple loci model (BLINK), revealed six genetic loci associated with HN and HT above the Bonferroni multiple-test threshold. Five candidate genes were identified based on the linkage disequilibrium with these loci, including GRMZM2G381691 and GRMZM2G012416. These two genes were up-regulation and down-regulation in all husk related tissues, respectively. GRMZM2G381691 associated with HT encoded a CCT domain protein, which expressed higher in tropical than temperate maize. GRMZM2G012416 associated with HN encoded an Armadillo (ARM) repeat protein, which regulated GA signal pathway. These associated SNPs and candidate genes paved a path to understand the genetic architecture of husk in maize.

Anthocyanin Profiling of Maize Grains Using DIESI-MSQD Reveals That Cyanidin-Based Derivatives Predominate in Purple Corn, whereas Pelargonidin-Based Molecules Occur in Red-Pink Varieties from Mexico.

Corn seeds contain natural pigments and antioxidants, such as the molecular variants of flavonoids and carotenoids. The aleurone and pericarp tissues from pigmented genotypes were extracted for metabolic fingerprinting and evaluated using UV-vis and mass spectrometry (MS). MS ionomic fingerprints classified samples according to genetic background and kernel color. The MS/MS fragmentation pattern (Daughter and Neutral Loss methods) allowed the tentative identification of 18 anthocyanins with glycosyl, malonyl, and succinyl moieties, including 535 m/z for cyanidin-3-O-(6″-malonyl-glucoside) and 621 m/z for cyanidin-3-O-(3″,6″-dimalonyl-glucoside). We also detected 663 m/z for pelargonidin-3-O-(disuccinyl-glucoside) and 633 m/z for peonidin-3-O-(disuccinyl-glucoside). Cyanidin-based anthocyanins were the most abundant in dark purple colored kernels, while pelargonidins predominated in the red-pink kernels of the "Elote occidental" landrace. Grains of "Conico negro" had a simultaneous pigmentation of aleurone and pericarp, while Vitamaize had purple pigmentation only in the aleurone layer. Most landraces had a white endosperm, while Vitamaize had a yellow endosperm and a dark seed coat. We conclude that Vitamaize grains contain both carotenes and anthocyanins, and therefore it is proposed as a nontransgenic agronomically improved variety of tropical purple maize, a good source for organic superfoods.