Identification and validation of genomic regions influencing kernel zinc and iron in maize.

KEY MESSAGE: Genome-wide association study (GWAS) on 923 maize lines and validation in bi-parental populations identified significant genomic regions for kernel-Zinc and-Iron in maize. Bio-fortification of maize with elevated Zinc (Zn) and Iron (Fe) holds considerable promise for alleviating under-nutrition among the world's poor. Bio-fortification through molecular breeding could be an economical strategy for developing nutritious maize, and hence in this study, we adopted GWAS to identify markers associated with high kernel-Zn and Fe in maize and subsequently validated marker-trait associations in independent bi-parental populations. For GWAS, we evaluated a diverse maize association mapping panel of 923 inbred lines across three environments and detected trait associations using high-density Single nucleotide polymorphism (SNPs) obtained through genotyping-by-sequencing. Phenotyping trials of the GWAS panel showed high heritability and moderate correlation between kernel-Zn and Fe concentrations. GWAS revealed a total of 46 SNPs (Zn-20 and Fe-26) significantly associated (P ≤ 5.03 × 10-05) with kernel-Zn and Fe concentrations with some of these associated SNPs located within previously reported QTL intervals for these traits. Three double-haploid (DH) populations were developed using lines identified from the panel that were contrasting for these micronutrients. The DH populations were phenotyped at two environments and were used for validating significant SNPs (P ≤ 1 × 10-03) based on single marker QTL analysis. Based on this analysis, 11 (Zn) and 11 (Fe) SNPs were found to have significant effect on the trait variance (P ≤ 0.01, R2 ≥ 0.05) in at least one bi-parental population. These findings are being pursued in the kernel-Zn and Fe breeding program, and could hold great value in functional analysis and possible cloning of high-value genes for these traits in maize.

Genome-wide association analysis reveals new targets for carotenoid biofortification in maize.

KEY MESSAGE: Genome-wide association analysis in CIMMYT's association panel revealed new favorable native genomic variations in/nearby important genes such as hydroxylases and CCD1 that have potential for carotenoid biofortification in maize. Genome-wide association studies (GWAS) have been used extensively to identify allelic variation for genes controlling important agronomic and nutritional traits in plants. Provitamin A (proVA) enhancing alleles of lycopene epsilon cyclase (LCYE) and ß-carotene hydroxylase 1 (CRTRB1), previously identified through candidate-gene based GWAS, are currently used in CIMMYT's maize breeding program. The objective of this study was to identify genes or genomic regions controlling variation for carotenoid concentrations in grain for CIMMYT's carotenoid association mapping panel of 380 inbred maize lines, using high-density genome-wide platforms with ~476,000 SNP markers. Population structure effects were minimized by adjustments using principal components and kinship matrix with mixed models. Genome-wide linkage disequilibrium (LD) analysis indicated faster LD decay (3.9 kb; r (2) = 0.1) than commonly reported for temperate germplasm, and therefore the possibility of achieving higher mapping resolution with our mostly tropical diversity panel. GWAS for various carotenoids identified CRTRB1, LCYE and other key genes or genomic regions that govern rate-critical steps in the upstream pathway, such as DXS1, GGPS1, and GGPS2 that are known to play important roles in the accumulation of precursor isoprenoids as well as downstream genes HYD5, CCD1, and ZEP1, which are involved in hydroxylation and carotenoid degradation. SNPs at or near all of these regions were identified and may be useful target regions for carotenoid biofortification breeding efforts in maize; for example a genomic region on chromosome 2 explained ~16% of the phenotypic variance for ß-carotene independently of CRTRB1, and a variant of CCD1 that resulted in reduced ß-cryptoxanthin degradation was found in lines that have previously been observed to have low proVA degradation rates.

Provitamin A Carotenoids in Grain Reduce Aflatoxin Contamination of Maize While Combating Vitamin A Deficiency.

Aflatoxin contamination of maize grain and products causes serious health problems for consumers worldwide, and especially in low- and middle-income countries where monitoring and safety standards are inconsistently implemented. Vitamin A deficiency (VAD) also compromises the health of millions of maize consumers in several regions of the world including large parts of sub-Saharan Africa. We investigated whether provitamin A (proVA) enriched maize can simultaneously contribute to alleviate both of these health concerns. We studied aflatoxin accumulation in grain of 120 maize hybrids formed by crossing 3 Aspergillus flavus resistant and three susceptible lines with 20 orange maize lines with low to high carotenoids concentrations. The hybrids were grown in replicated, artificially-inoculated field trials at five environments. Grain of hybrids with larger concentrations of beta-carotene (BC), beta-cryptoxanthin (BCX) and total proVA had significantly less aflatoxin contamination than hybrids with lower carotenoids concentrations. Aflatoxin contamination had negative genetic correlation with BCX (-0.28, p < 0.01), BC (-0.18, p < 0.05), and proVA (-0.23, p < 0.05). The relative ease of breeding for increased proVA carotenoid concentrations as compared to breeding for aflatoxin resistance in maize suggests using the former as a component of strategies to combat aflatoxin contamination problems for maize. Our findings indicate that proVA enriched maize can be particularly beneficial where the health burdens of exposure to aflatoxin and prevalence of VAD converge with high rates of maize consumption.