Genetic analysis and hybrid prediction in tropical maize (Zea mays L.) using phenotypic and single nucleotide polymorphic markers.

Abstract

Maize (Zea mays L., 2n = 2x = 20) is a commodity crop serving the food, feed, and processing industries globally. The productivity of maize in Africa remains low (< 2 t/ha) due to various yieldlimiting factors, including abiotic stresses (such as drought, heat stress, flooding, waterlogging, erosion and poor soil health), and biotic stresses (e.g. foliar diseases and insect pests). Limited adoption of new high yielding varieties, slow rate of varietal turnover , socio-economic constraints, and policy issues further hinder productivity. Seed Co Limited is a Pan-African seed company involved in the research, development, and commercialization of seeds of major food security grain crops, including maize. The Seed Co breeding program aims to enhance the yields of new generation maize cultivars via hybrid breeding by utilizing complementary and contrasting inbred lines. New lines and experimental hybrids are developed and phenotyped using economic agronomic traits and genotyped using high-resolution Single Nucleotide Polymorphism (SNP) markers to facilitate effective selection. Integrating phenotypic and genomic selection accelerates the development of inbred lines with desirable traits to create high-performing single crosses and three-way hybrids. The new hybrids should undergo rigorous field testing for yield gains and stability across various locations to guide cultivar release and commercialization. Therefore, to complement this breeding initiative, the objectives of the study were: to assess a maize germplasm panel's genetic diversity and population structure comprising 182 founder lines and 866 derived inbred lines using Single Nucleotide Polymorphism (SNP) markers to identify genetically unique lines for hybrid breeding, to conduct genome-wide prediction of yield and component traits using qualitative and quantitative phenotypic traits and SNP markers based on the additive-dominant genomic best linear unbiased predictions model to compute genomic estimated breeding values and genomic estimated genetic values to guide inbred line development and hybrid breeding, to assess the gains in yield and yield components among single cross maize hybrids selected through genomic prediction across representative locations to guide breeding and production and to determine the combining ability effects of newly selected inbred lines and quantify the magnitude of heterosis and genotype by environmental interaction (GEI) effects of single cross hybrids to select and recommend contrasting elite lines and experimental hybrids. In the first study, 182 founder and 866 derived maize inbred lines were characterized for genetic diversity and population structure analyses using SNP markers to identify genetically unique lines for hybrid breeding through beneficial allelic combinations. Genotyping was performed using the Affymetrix platform for the 182 founder lines (1201 SNP markers) and the Midseq platform for the 866 derived lines (1484 markers). Moderate genetic variation with genetic distance ranging from 0.004 to 0.44 (mean: 0.25) for founder lines and 0.004 to 0.34 (mean: 0.13) for derived lines was observed. Heterozygosity values ranged from 0.00 to 0.24 for both lines. About 82% of the 1201 markers and 84% of the 1484 markers exhibited polymorphism information content ranging from 0.25 to 0.50, detecting a high level of genetic diversity and that the SNPs were highly informative in distinguishing the tested lines. Analysis of molecular variance revealed significant genetic differences (P ≤ 0.001) among and within populations in the founder and derived lines. Notably, within-population variations accounted for 97% (founder lines) and 88.38% (derived lines) of the detected variations. Population structure analysis identified three subpopulations among founder lines and two among derived lines, which was supported by cluster analysis. Based on pairwise comparisons, genetically distant lines were selected, including G15NL337 and G15NL312 (Cluster 1), 15ARG152 and RGS-PL44 (Cluster 2), RGS-PL44 and 15ARG149 (Cluster 2), and RGS-PL33 and RGS-PL44 (Cluster 2). The selected lines are genetically distinct and recommended for marker-assisted hybrid maize breeding to leverage beneficial alleles. The second study genotyped 1,102 genetically diverse inbred lines from two heterotic groups (N3 and SC) using high-density SNP markers. The 1,102 lines and 4 testers were crossed in a line-by-tester design to generate 2,830 single cross hybrids (SCHs). Phenotypic data were collected from field trials with the following SCHs: 684 evaluated at five locations in 2018/19, 760 at four locations (2019/20), 646 at four locations (2020/21), and 740 at four locations (2021/22) summer seasons in Zimbabwe. The trials were laid out in a 6 x 7 alpha lattice design with two replications at each site. 20 highperforming and contrasting inbred lines with the highest genomic estimated breeding values (GEBVs) and genomic estimated genetic values (GEGVs), each from the two heterotic groups, were identified for genetic advancement, combining ability tests and commercial hybrid development. 20 highperforming candidate SCHs with high GEGVs were identified for three-way hybrid development, variety registration and commercialization. In the third study, 30 SCHs were developed from 11 inbred lines (6 from the N3 group and 5 from the SC group) with the highest predicted GEGVs for grain yield and associated traits using the genotypic best linear unbiased prediction (GBLUP) model. The lines were crossed using a factorial mating design with the six N3 lines used as female and five SC lines as male. The derived 30 SCHs and six commercial single cross check hybrids were field evaluated in seven locations, four in Zimbabwe and three in Zambia using a 6 x 6 alpha lattice design with two replications at each location. A combined analysis of variance revealed significant (P≤0.05) variation among the hybrids for the assessed 11 quantitative traits. Significant yield gains were realized over the mean of checks (at 13.09%), mean of the population (10.83%) and mean of best check (1.47%). Moderate to high broad-sense heritability (50 to 94%) and genetic advance were recorded for most of the assessed traits, indicating the success of selection assisted by genomic predictions. The study identified three best single cross hybrids (i.e., CTL03 x G16NL721, CTL03 x G17NL544 and GS-PL07 x G17NL544) with high and stable yields and recommended for commercialization. In the fourth study, 11 elite inbred lines (6 female parents from N3 and 5 male parents from SC group) were crossed using a factorial mating design, resulting in 30 SCHs. The lines were selected based on the highest GEGVs for yield and component traits through GS using the GBLUP model. The 30 SCHs and six commercial check hybrids were field evaluated at seven locations (four in Zimbabwe and three in Zambia) during the 2022/2023 summer season. The trials were arranged in a 6 x 6 alpha lattice design with two replications at each location. Data were recorded on yield and yield components, and general combining ability (GCA) and specific combining ability (SCA) effects were computed. Significant GCA effects for grain yield (GY) were noted for lines CTL03, G17NL544, G16NL721, and GS-PL07, while significant SCA effects were recorded for crosses 15AG163 x G16NL679, G15NL304 x G17NL642, and 15AG162 x G16NL679. The additive main effects and multiplicative interaction (AMMI) model explained 38.95%, 50.58% and 7.24% of the total variation in GY due to genotype (G), environment (E), and genotype x environment interaction (GEI) effects in that order. The test locations were clustered into two mega environments: Rattray Arnold Research Station (RARS), Agricultural Research Trust (ART), Mpongwe Research Station (MPRS), and Lusaka West Research Station (LWRS) (Environment 1), and Mkushi Research Station (MKRS), Stapleford Research Centre (STAP), and Kadoma Research Centre (KRC) (Environment 2). The genotype and genotype-by-environment interaction (GGE) biplot analysis identified hybrids G15NL304 x G17NL544 and 15AG162 x G17NL544 as high-yielding and stable, suitable for commercialization. The two mega-environments and the selected stable, high-yielding general and specific combiners are recommended for genotype evaluation and production in Zimbabwe, Zambia, and comparable agroecologies. Overall, the present study identified contrasting and genetically delineated inbred lines and enhanced the existing heterotic groups using high-throughput SNP markers. Best-performing lines (e.g. CTL03 and GS-PL07) were selected from the N3 heterotic group and G17NL544 and G16NL721 from the SC heterotic group. New single cross hybrids, such as CTL03 x G16NL721, CTL03 x G17NL544, and GS-PL07 x G17NL544, were selected with grain yields of 8.38 t/ha, 8.24 t/ha, and 8.23 t/ha, respectively. The new experimental hybrids are recommended for three-way hybrid development or release following multi-environment evaluation.