Molecular marker-based genetic diversity assessment of Striga-resistant maize inbred lines

Striga-resistant maize inbred lines are of interest to maize breeding programs in the savannas of Africa where the parasitic weed is endemic and causes severe yield losses in tropical maize. Assessment of the genetic diversity of such inbred lines is useful for their systematic and efficient use in a breeding program. Diversity analysis of 41 Striga-resistant maize inbred lines was conducted using amplified fragment length polymorphism (AFLP) and simple sequence repeat (SSR) markers to examine the genetic relationships among these lines and to determine the level of genetic diversity that exists within and between their source populations. The two marker systems generated 262 and 101 polymorphic fragments, respectively. Genetic similarity (GS) values among all possible pairs of inbred lines varied from 0.45 to 0.95, with a mean of 0.61±0.002 for AFLPs, and from 0.21 to 0.92, with a mean of 0.48±0.003, for SSRs. The inbred lines from each source population exhibited a broad range of GS values with the two types of markers. Both AFLPs and SSRs revealed similar levels of within population genetic variation for all source populations. Cluster and principal component analysis of GS estimates with the two markers revealed clear differentiation of the Striga-resistant inbred lines into groups according to their source populations. There was clear separation between early- and late-maturing Striga-resistant inbred lines. Considering the paucity of germplasm with good levels of resistance to Striga in maize, the broad genetic diversity detected within and among source populations demonstrates the genetic potential that exists to improve maize for resistance to Striga.     

Molecular characterization and assessment of genetic diversity of inbred lines showing variability for drought tolerance in maize

A set of 24 genotypes bred at different centres in India as well as in CIMMYT showing variability for drought tolerance were selected for molecular and morpho-physiological characterization. A set of 35 SSR markers, having genome-wide coverage, was chosen for genotyping the inbreds. These markers generated a total of 111 polymorphic alleles with an average of 3.17 alleles per locus. The minimum and maximum PIC value was 0.27 and 0.77 with a mean of 0.5. A total of 13 unique alleles were found in the 24 inbred lines. The coefficient of genetic dissimilarity ranged from 0.192 to 0.803. NJ-based tree suggested the presence of three major clusters of which, two of them had subgroups. Phenotyping of inbreds by morpho-physiological traits revealed that there was a positive relationship among root length, chlorophyll content, relative water content while anthesis-silking interval was negative relationship with all these traits. Genotyping data complemented by morpho-physiological parameters were used to identify a number of pair-wise combinations for the development of mapping population segregating for drought tolerance and potential heterotic pairs for the development of drought tolerant hybrids.     

Genetic dissection of intermated recombinant inbred lines using a new genetic map of maize

A new genetic map of maize, ISU-IBM Map4, that integrates 2029 existing markers with 1329 new indel polymorphism (IDP) markers has been developed using intermated recombinant inbred lines (IRILs) from the intermated B73xMo17 (IBM) population. The website http://magi.plantgenomics.iastate.edu provides access to IDP primer sequences, sequences from which IDP primers were designed, optimized marker-specific PCR conditions, and polymorphism data for all IDP markers. This new gene-based genetic map will facilitate a wide variety of genetic and genomic research projects, including map-based genome sequencing and gene cloning. The mosaic structures of the genomes of 91 IRILs, an important resource for identifying and mapping QTL and eQTL, were defined. Analyses of segregation data associated with markers genotyped in three B73/Mo17-derived mapping populations (F2, Syn5, and IBM) demonstrate that allele frequencies were significantly altered during the development of the IBM IRILs. The observations that two segregation distortion regions overlap with maize flowering-time QTL suggest that the altered allele frequencies were a consequence of inadvertent selection. Detection of two-locus gamete disequilibrium provides another means to extract functional genomic data from well-characterized plant RILs.     

Evaluating genetic relationships between tropical maize inbred lines by means of AFLP profiling

Diversity among tropical maize inbred lines that compose breeding programs, is not well known. The lack of this information has made the arrangement of heterotic groups to be used for breeding purposes difficult. Methods of molecular analysis have been used as efficient alternatives for evaluating genetic diversity, aiming at heterotic group arrangement and acquisition of new hybrids. In this study, AFLP (amplified fragment length polymorphism) was used to investigate the genetic relationships among 96 tropical maize inbred lines from two different origins. The polymorphism level among the genotypes and the possibility of their allocation in heterotic groups were evaluated. Besides, correlations among genetic diversity and flowering time were analyzed. Nine primer combinations were used to obtain AFLP markers, producing 638 bands, 569 of which were polymorphic. Genetic similarities (GS), determined by Jaccard's similarity coefficient, varied from 0.345 to 0.891, with an average of 0.543. The dendrogram based on the GS and on the UPGMA cluster method did not separate the inbred lines in well-defined groups. Aiming at separating the lines into more accurate groups, Tocher's optimization procedure was carried out, 17 groups being identified. Association between flowering time and germplasm pools was detected. AFLP showed itself to be a robust assay, revealing a great power of detection of genetic variability in the tropical germplasm, and also demonstrated to be very useful for guiding breeding programs.     

Genetic diversity of maize inbred lines within and among heterotic groups revealed by RFLPs

Summary The objectives of this study were (1) to investigate genetic diversity for RFLPs in a set of important maize inbreds commonly used in Italian breeding programs, (2) to compare genetic similarities between unrelated lines from the same and different heterotic groups, and (3) to examine the potential of RFLPs for assigning maize inbreds to heterotic groups. Forty inbreds were analyzed for RFLPs with two restriction enzymes (EcoRI and HindIII) and 82 DNA clones uniformly distributed over the maize genome. Seventy clone-enzyme combinations gave single-banded RFLP patterns, and 79 gave multiple-banded RFLP patterns. The average number of RFLP patterns detected per clone-enzyme combination across all inbreds was 5.8. RFLP data revealed a wide range of genetic diversity within the two heterotic groups assayed, Iowa Stiff Stalk Synthetic (BSSS) and Lancaster Sure Crop (LSC). Genetic similarity (GS) between lines was estimated from binary RFLP data according to the method of Nei and Li (1979). The mean GS for line combinations of type BSSS × LSC (0.498) was substantially smaller than for unrelated line combinations or type BSSS × BSSS (0.584) but almost as great as for un-related line combinations of type LSC × LSC (0.506). Principal coordinate and cluster analyses based on GS values resulted in the separate groupings of lines, which is consistent with known pedigree information. A comparison between both methods for multivariate analyses of RFLP data is presented.     

Microsatellite marker-based genetic diversity assessment among exotic and native maize inbred lines of Bangladesh

Hybrid development is basically dependent on the variability among available genetic resources. Polymorphism among the maize inbreds is essentially needed for maize hybridization. This study aimed at the assessment of diversity among 22 maize inbreds by 18 microsatellite markers. The study identified 187 alleles at 18 SSR loci. The amplified allele frequency per microsatellite locus was 10.4 and the highest allele per locus was 17 in SSR primer pair phi026. SSR primer set p-umc1292, phi074 and phi090 showed the lowest 6 alleles per genotype per locus. The locus phi026 showed the highest degree of gene diversity (0.92), and the locus p-umc1292 had the lowest of gene diversity (0.77) with a mean value of 0.862 among the microsatellites. At each site, the most prevalent allele varied between 0.14 (bnlg371) and 0.36. (p-umc1292). At any given locus, an average of 0.22 out of the 22 selected maize inbred lines had a common major allele. The average value of the polymorphic information content (PIC) was 0.85, within the range of 0.74 at the lowest to 0.92 at the highest. The higher PIC values of phi026 and nc013 established them to be the best markers for maize inbred lines. The UPGMA clustering generated seven distinct groups having 12.5% of similarity coefficient. The results revealed that inbred lines E10, E27, E19, E34, E35, E4, E43, E28, E11, E21, E17, E38, E25, E34, E14, E16, E39 and E3 were more diversified. These lines are promising to be used as parent materials for hybrid maize development in the future.     

Patterns of genomic variation in Chinese maize inbred lines and implications for genetic improvement

Key message

Genetic relationships among Chinese maize germplasms reveal historical trends in heterotic patterns from Chinese breeding programs and identify line Dan340 as a potential genome donor for elite inbred line Zheng58.

Abstract

The characterization of the genetic relationships, heterotic patterns and breeding history of lines in maize breeding programs allows breeders to efficiently use maize germplasm for line improvement over time. In this study, 269 temperate inbred lines, most of which have been widely used in Chinese maize breeding programs since the 1970s, were genotyped using the Illumina MaizeSNP50 BeadChip, which contains 56,110 single-nucleotide polymorphisms. The STRUCTURE analysis, cluster analysis and principal coordinate analysis results consistently revealed seven groups, of which five were consistent with known heterotic groups within the Chinese maize germplasm—Domestic Reid, Lancaster, Zi330, Tang SPT and Tem-tropic I (also known as “P”). These genetic relationships also allowed us to determine the historical trends in heterotic patterns during the three decades from 1970 to 2000, represented by Mo17 from Lancaster, HuangZaoSi (HZS) from Tang SPT, Ye478 from Domestic Reid and P178 from Tem-tropic I heterotic groups. Mo17-related commercial hybrids were widely used in the 1970s and 1980s, followed by the release of HZS- and Ye478-related commercial hybrids in the 1980s and 1990s, and the introduction of Tem-tropic I group in the 1990s and 2000s. Additionally, we identified inbred line Dan340 as a potential genome donor for Zheng58, which is the female parent of the most widely grown commercial hybrid ZhengDan958 in China. We also reconstructed the recombination events of elite line HZS and its 14 derived lines. These findings provide useful information to direct future maize breeding efforts.

     

SSR analysis of genetic diversity among maize inbred lines adapted to cold regions of Japan

Information regarding diversity and relationships among breeding material is necessary for hybrid maize (Zea mays L.) breeding. Simple-sequence repeat (SSR) analysis of the 60 loci distributed uniformly throughout the maize genome was carried out for 65 inbred lines adapted to cold regions of Japan in order to assess genetic diversity among the inbred lines and to assign them to heterotic groups. The mean value (0.69) of the polymorphic-index content (PIC) for the SSR loci provided sufficient discrimination-ability for the assessment of genetic diversity among the inbred lines. The correlation between the genetic-similarity (GS) estimates and the coancestry coefficient was significant (r = 0.70). The average-linkage (UPGMA) cluster analysis and principal-coordinate analysis (PCOA) for a matrix of the GS estimates showed that the Northern flint inbred lines bred in Japan were similar to a Canadian Northern flint inbred line CO12 and a European flint inbred line F283, and that dent inbred lines bred in Japan were similar to BSSS inbred lines such as B73. These associations correspond to the known pedigree records of these inbred lines. The results indicate that SSR analysis is effective for the assessment of genetic diversity among maize inbred lines and for the assignment of inbred lines to heterotic groups.     

Classical genetic and quantitative trait loci analyses of heterosis in a maize hybrid between two elite inbred lines

The exploitation of heterosis is one of the most outstanding advancements in plant breeding, although its genetic basis is not well understood yet. This research was conducted on the materials arising from the maize single cross B73 x H99 to study heterosis by procedures of classical genetic and quantitative trait loci (QTL) analyses. Materials were the basic generations, the derived 142 recombinant inbred lines (RILs), and the three testcross populations obtained by crossing the 142 RILs to each parent and their F(1). For seedling weight (SW), number of kernels per plant (NK), and grain yield (GY), heterosis was >100% and the average degree of dominance was >1. Epistasis was significant for SW and NK but not for GY. Several QTL were identified and in most cases they were in the additive-dominance range for traits with low heterosis and mostly in the dominance-overdominance range for plant height (PH), SW, NK, and GY. Only a few QTL with digenic epistasis were identified. The importance of dominance effects was confirmed by highly significant correlations between heterozygosity level and phenotypic performance, especially for GY. Some chromosome regions presented overlaps of overdominant QTL for SW, PH, NK, and GY, suggesting pleiotropic effects on overall plant vigor.     

Uptake and distribution of cadmium in maize inbred lines

Genotypic variation in uptake and distribution of cadmium (Cd) was studied in 19 inbred lines of maize (Zea mays L.). The inbred lines were grown for 27 days on an in situ Cd-contaminated sandy soil or for 20 days on nutrient solution culture with 10 µg Cd L^-1. The Cd concentrations in the shoots showed large genotypic variation, ranging from 0.9 to 9.9 µg g^-1 dry wt. for the Cd-contaminated soil and from 2.5 to 56.9 µg g^-1 dry wt. for the nutrient solution culture. The inbred lines showed a similar ranking for the Cd concentrations in the shoots for both growth media (r²=0.89). Two main groups of inbreds were distinguished: a group with low shoot, but high root Cd concentrations (shoot: 7.4±5.3 µg g^-1 dry wt.; root: 206.0±71.2 µg g^-1 dry wt.; shoot Cd excluder) and a group with similar shoot and root Cd concentrations (shoot: 54.2±3.4 µg g^-1 dry wt.; root: 75.6±11.2 µg g^-1 dry wt.; non-shoot Cd excluder). The classification of the maize inbred lines and the near equal whole-plant Cd uptake between the two groups demonstrates that internal distribution rather than uptake is causing the genotypic differences in shoot Cd concentration of maize inbred lines. Zinc (Zn), a micronutrient chemically related to Cd, showed an almost similar distribution pattern for all maize inbred lines. The discrepancy in the internal distribution between Cd and Zn emphasizes the specificity of the Cd distribution in maize inbred lines.     

Quality control genotyping for assessment of genetic identity and purity in diverse tropical maize inbred lines

Quality control (QC) genotyping is an important component in breeding, but to our knowledge there are not well established protocols for its implementation in practical breeding programs. The objectives of our study were to (a) ascertain genetic identity among 2–4 seed sources of the same inbred line, (b) evaluate the extent of genetic homogeneity within inbred lines, and (c) identify a subset of highly informative single-nucleotide polymorphism (SNP) markers for routine and low-cost QC genotyping and suggest guidelines for data interpretation. We used a total of 28 maize inbred lines to study genetic identity among different seed sources by genotyping them with 532 and 1,065 SNPs using the KASPar and GoldenGate platforms, respectively. An additional set of 544 inbred lines was used for studying genetic homogeneity. The proportion of alleles that differed between seed sources of the same inbred line varied from 0.1 to 42.3?%. Seed sources exhibiting high levels of genetic distance are mis-labeled, while those with lower levels of difference are contaminated or still segregating. Genetic homogeneity varied from 68.7 to 100?% with 71.3?% of the inbred lines considered to be homogenous. Based on the data sets obtained for a wide range of sample sizes and diverse genetic backgrounds, we recommended a subset of 50–100 SNPs for routine and low-cost QC genotyping, verified them in a different set of double haploid and inbred lines, and outlined a protocol that could be used to minimize errors in genetic analyses and breeding.     

Analysis of genetic diversity and relationships among waxy maize inbred lines in Korea using SSR markers

Information regarding the genetic diversity and genetic relationships among elite inbred lines is necessary to improve new cultivars in maize breeding programs. In this study, genetic diversity and genetic relationships were investigated among 84 waxy maize inbred lines using 50 SSR markers. A total of 269 alleles were identified at all the loci with an average of 5.38 and a range between 2 and 13 alleles per locus. The gene diversity values varied from 0.383 to 0.923 with an average of 0.641. The cluster tree generated using the described SSR markers recognized two major groups at 32% genetic similarity. Group I included 33 inbred lines while group II included 51 inbred lines. The clustering patterns of most of the waxy maize inbred lines did not clearly agree with their source, pedigree or geographic location. The average GS among all inbred lines was 35.7 ± 10.8. Analysis of waxy maize inbred lines collected from Korea and China at 50 SSR loci revealed higher values of average number of alleles (4.9) and gene diversity (0.638) in Korean inbred lines as compared to Chinese inbred lines (3.5 and 0.563, respectively). The information obtained from the present studies would be very useful for maize breeding programs in Korea.     

Comparison of whole-genome prediction models for traits with contrasting genetic architecture in a diversity panel of maize inbred lines

ABSTRACT: BACKGROUND: There is increasing empirical evidence that whole-genome prediction (WGP) is a powerful tool for predicting line and hybrid performance in maize. However, there is a lack of knowledge about the sensitivity of WGP models towards the genetic architecture of the trait. Whereas previous studies exclusively focused on highly polygenic traits, important agronomic traits such as disease resistances, nutrifunctional or climate adaptational traits have a genetic architecture which is either much less complex or unknown. For such cases, information about model robustness and guidelines for model selection are lacking. Here, we compared five WGP models with different assumptions about the distribution of the underlying genetic effects. As contrasting model traits, we chose three highly polygenic agronomic traits and three metabolites each with a major QTL explaining 22 to 30 % of the genetic variance in a panel of 289 diverse maize inbred lines genotyped with 56,110 SNPs. RESULTS: We found the five WGP models to be remarkable robust towards trait architecture with the largest differences in prediction accuracies ranging between 0.05 and 0.14 for the same trait, most likely as the result of the high level of linkage disequilibrium prevailing in elite maize germplasm. Whereas RR-BLUP performed best for the agronomic traits, it was inferior to LASSO or elastic net for the three metabolites. We found the approach of genome partitioning of genetic variance, first applied in human genetics, as useful in guiding the breeder which model to choose, if prior knowledge of the trait architecture is lacking. CONCLUSIONS: Our results suggest that in diverse germplasm of elite maize inbred lines with a high level of LD, WGP models differ only slightly in their accuracies, irrespective of the number and effects of QTL found in previous linkage or association mapping studies. However, small gains in prediction accuracies can be achieved if the WGP model is selected according to the genetic architecture of the trait. If the trait architecture is unknown e.g. for novel traits which only recently received attention in breeding, we suggest to inspect the distribution of the genetic variance explained by each chromosome for guiding model selection in WGP.     

Agrobacterium-mediated genetic transformation of selected tropical inbred and hybrid maize (Zea mays L.) lines

The study was carried out to evaluate the amenability of tropical inbred and hybrid maize lines, using Agrobacterium mediated transformation technique. Agrobacterium tumefaciens strains EHA101 harbouring a pTF102 binary vector, EHA101, AGL1, and LBA4404 harbouring pBECK2000.4 plasmid, LBA4404, GV and EHA105 harbouring pCAMBIA2301 plasmid, and AGL1 harbouring the pSB223 plasmid were used. Delivery of transgenes into plant tissues was assessed using transient β-glucuronidase (gus) activity on the 3rd and 4th day of co-cultivation of the infected Immature Zygotic Embryos (IZEs) and embryogenic callus. Transient gus expression was influenced by the co-cultivation period, maize genotype and Agrobacterium strain. The expression was highest after the 3rd day of co-culture compared to the 4th day with intense blue staining was detected for IZEs which were infected with Agrobacterium strains EHA105 harbouring pCAMBIA2301 and EHA101 harbouring pTF102 vector. Putative transformants (T_o) were regenerated from bialaphos resistant callus. Differences were detected on the number of putative transformants regenerated among the maize lines. Polymerase chain reaction (PCR) amplification of Phosphinothricin acetyltransferase (bar) and gus gene confirmed the transfer of the transgenes into the maize cells. Southern blot hybridization confirmed stable integration of gus into PTL02 maize genome and segregation analysis confirmed the inheritance of the gus. A transformation efficiency of 1.4 % was achieved. This transformation system can be used to introduce genes of interest into tropical maize lines for genetic improvement.     

Performance prediction of F1 hybrids between recombinant inbred lines derived from two elite maize inbred lines

Selection of recombinant inbred lines (RILs) from elite hybrids is a key method in maize breeding especially in developing countries. The RILs are normally derived by repeated self-pollination and selection. In this study, we first investigated the accuracy of different models in predicting the performance of F₁ hybrids between RILs derived from two elite maize inbred lines Zong3 and 87-1, and then compared these models through simulation using a wider range of genetic models. Results indicated that appropriate prediction models depended on genetic architecture, e.g., combined model using breeding value and genome-wide prediction (BV+GWP) has the highest prediction accuracy for high V _D/V _A ratio (>0.5) traits. Theoretical studies demonstrated that different components of genetic variance were captured by different prediction models, which in turn explained the accuracy of these models in predicting the F₁ hybrid performance. Based on genome-wide prediction model (GWP), 114 untested F₁ hybrids possibly having higher grain yield than the original F₁ hybrid Yuyu22 (the single cross between Zong3 and 87-1) have been identified and recommended for further field test.