Heterosis: current advances in the search for molecular mechanisms

Heterosis as the increased performance of hybrid progeny compared to their inbred parents is one of the most intriguing phenomena in genetics. The first attempts to find out about underlying mechanisms were based on theoretical models, which were useful, but could not characterize this unique phenomenon as a whole. With the advent of molecular markers great efforts were made to identify genomic regions causing heterotic response and clarify prospects of using information about molecular divergence of parental forms as a criterion for the prediction of F1 performance. Despite some achievements, the effec-tiveness of both molecular divergence and prospective heterotic QTL for practical goals was limited, confirming that genetic heterogeneity is necessary, but not sufficient to produce perfect phenotype. Current methodological tools of functional genomics and related disciplines have provided new opportunities for searching for mechanisms of heterosis at different levels in the context of relative importance of dominance, overdominance and epistasis. To date, differences in genome organization, gene expression and epigenetic status have been found between hybrids and their parents. At the genomic level, some QTLs associated with heterosis were identified and the impact of DNA divergence on F1 performance was evaluated. At the level of transcriptome, it was shown that heterosis in hybrids occurs along with changes in gene expression regulation under the influence of circadian clock genes. Several studies have been conducted to clarify the role of epigenetic DNA modification and genomic imprinting in the manifestation of heterosis. Taken together, data indicates that heterosis cannot been explained by a single common mechanism, because this complex phenomenon involves many components, a cumulative effect of which leads to the formation of an outstanding phenotype

Theoretical aspects of heterosis

The phenomenon of heterosis, known as superior performance of hybrid organism compared with either of their parents, has been exploited by agricultural practices in the production of various crops since the beginning of the last century; however, its genetic basis has remained obscure. With experimental data obtained from the study of maize hybrids, and mathematical calculations, some genetic models have been proposed to explain heterosis from various types of gene interaction, such as dominance, over-dominance and epistasis. However, any of the proposed concepts has weak points, which place limitations on the possibility of overall interpretation of heterotic response in F1. In this review we gather a brief account of findings from various studies for critical evaluation of the main theoretical concepts based on the information accumulated to date by genetics and molecular biology and focused on particular mechanisms acting for specific traits. We discussed some aspects concerning the role of mutation loads in the formation of heterotic phenotype. Also, we gathered a brief account of findings for interpretation of genetic effects due to linkage and non-allelic genes’ interactions that make nuances to analysis of dominance and over-dominance. We have provided information about combining ability, its practical application in the context of the concept of heterotic groups. Here we also discussed some aspects of “genotype–environment” interaction. Recent advancements in genetics and molecular biology indicate the importance of various types of gene action for heterosis and confirm the necessity of systemlevel approaches to understanding this unique phenomenon

Mineral composition of bread wheat lines with introgressions of alien genetic material

Background. Modern bread wheat (Triticum aestivum L.) cultivars developed mainly to increase productivity often contain low concentrations of minerals in their grain. Wild and primitive wheats demonstrate rich genetic diversity, including the content of minerals in the grain, and can be used to improve the wheat gene pool for this trait. The aim of this work was to study the mineral composition in the grain of bread wheat lines with introgressions of the genetic material from Triticum L. spp.Materials and methods. We studied parental spring bread wheat cultivars, accessions of tetraploid and hexaploid Triticum spp., and 20 introgressive lines obtained on their basis in 2018 and 2020. Concentrations of macro- (K, P, Ca, and Mg) and micronutrients (Zn, Fe, Cu, and Mn) were measured using atomic emission spectrometry with inductively coupled plasma, and total protein content in wheat grain according to GOST 10846-91. The data were processed using the Statistica 10.0 and MS Excel software packages.Results and conclusion. The content of minerals in the grain of Triticum spp. was higher than in T. aestivum cultivars (the twoyear average difference was 1,02 to 2,13-fold, depending on the studied mineral). Most of the lines with alien genetic material exceeded their parent bread wheat cultivars in Zn, Fe, Cu and Mn content and came close to them in the levels of N, P, Mg and K. ANOVA established a statistically significant impact of the genotype, environment, and genotype × environment interaction on the variation of the grain macro- and micronutrient content. Lines with a consistently high concentration of grain minerals and high productivity were identified. These lines are of interest for wheat breeding for grain quality

SSR loci potentially associated with high amylopectine content in maize kernel endosperm

As a component of functional nutrition, maize cultivars with “non-traditional” kernel composition (waxy, oilbearing, sugar, opaque, etc. phenotypic variants) are promising. Mutations in the waxy gene, which break down the structure and function of the enzyme for amylose biosynthesis, lead to a waxy (with a high content of amylopectin) endosperm formation. High variability of the waxy gene limits the use of microsatellite loci in marker associated selection of waxy maize genotypes. The increased frequency of gene rearrangements within the waxy locus facilitated the origination of many high-amylopectin corn lines carrying different SSR allelic variants. The purpose of this study was to evaluate the effectiveness of using waxy locus microsatellite sequences for identification and labeling of waxy maize genotypes. To this end, a complex of biochemical (calorimetry, bichromate method), molecular-genetic (SSR-PCR, capillary gel electrophoresis with fluorescent detection of fragments) and statistical (descriptive statistics, cluster analysis, χ2) analysis methods was used. Plant material used were 33 samples of corn kernels including mutant forms with a high content of amylose, amylopectin, short-chain starches, were kindly provided by VIR genetic collection (Russian Federation) and Maize Genetics Cooperation Stock Center (USA). The contents of starch, short-chain soluble carbohydrates, amylose, amylopectin in the grain of 33 maize samples were evaluated. Compositionally similar (to endosperm carbohydrates content) groups of samples were identified. They include 13 high-amylopectin samples carriers of waxy (wx) gene mutations and 20 samples with wild-type character (Wx). Molecular genetic screening of the collection included an analysis of the polymorphism of the microsatellite loci phi022, phi027, phi061 associated with the waxy gene sequence. Allelic composition of individual loci and their combinations were analyzed in relation to the accumulation of reserve carbohydrates in the kernel endosperm. Only the analysis of the phi022/phi027 combination or all three markers in the complex allows differentiating the wild Wx and mutant wx phenotypes of maize. It was shown that not the individual allelic polymorphisms of the phi022, phi027, phi061 loci are efficient for the markerassociated selection of high-amylopectin maize, but their unique combinations

Marker-assisted identification of maize genotypes with improved protein quality

Currently, more than 70 % of maize is used for food and fodder; therefore, grain quality improvement can increase its nutritive and energy value. Deficiency of two essential amino acids (lysine and tryptophan) significantly reduces the nutritional quality of maize proteins. However, in comparison to conventional maize varieties, opaque2 (o2) mutants have greater contents of lysine and tryptophan in their endosperm proteins and their bioavailability is better. The aim of the study was identification of maize accessions with high-quality protein. A collection of maize accessions of various ecogeographical origins was studied by molecular methods. This approach was expected to improve maize breeding efficiency. We collected 54 maize genotypes differing in grain quality performance. Amplification with three specific markers to the opaque-2 gene (phi057, phi112 and umc1066) revealed homozygous recessive o2 genotypes, associated with improved nutritional quality of the protein. UREA-PAG electrophoresis of zein proteins was used for Quality Protein Maize (QPM) identification. In addition to the mutant o2 allele, QPM contains genetic modifiers that convert starchy endosperm of o2 mutant to the hard vitreous phenotype. The selected QPM accessions are of interest for maize breeding programs aimed at grain quality improvement. The use of the markers to o2 and modifier genes accelerates the development of QPM varieties and significantly reduces the labor and financial costs of their production

Allelic and epigenetic DNA variation in relation to F1 heterosis manifestation in F1 hybrids of Capsicum annuum L.

Managing F1 heterosis is one of the major objectives in hybrid crop breeding programs. The classical theory considers the heterozygosity in F1 hybrids to be the main factor contributing to heterosis and therefore presumes a linear relationship between the value of genetic polymorphisms in parental lines and the heterotic response of their F1 offspring. Therefore, the genetic diversity information is viewed as a tool for selection of promising cross-combinations, but results published by different researchers are inconsistent. In this work, we studied the contributions of structural and nonstructural DNA polymorphisms to F1 heterosis manifestation. We used SSR and methyl-sensitive AFLP (MSAP with HpaII and MspI izoshisomers) protocols for obtaining specific patterns for heterotic and nonheterotic F1 hybrids of sweet pepper (Capsicum annuum L.) from a Belarusian breeding program. We found out that a certain portion of heterosis for yield-related traits might be explained by the polymorphism revealed by SSR analysis. According to our data, the total number of polymorphic SSR loci and the ratio of polymorphic and nonpolymorphic loci demonstrate a significant predictive value and can serve as additional prognostic criteria for the selection of promising cross-combinations. From the MSAP assay, we found a relationship between heterosis and the numbers of methylated and nonmethylated DNA loci for yield traits. Our results indicate that cross-hybridization may favor epiallelic modifications in F1 hybrids, presumably responsible for heterosis. Thus, epigenetic DNA variation may explain the absence of a linear relationship between the level of structural DNA divergence and F1 heterosis, as well as the manifestation of heterosis in crosses of related (genetically similar) accessions

ВЛИЯНИЕ ИНТРОГРЕССИЙ ГЕНЕТИЧЕСКОГО МАТЕРИАЛА ЭГИЛОПСОВ В ГЕНОМЕ ГИБРИДНЫХ ЛИНИЙ ТРИТИКАЛЕ НА БИОХИМИЧЕСКИЙ СОСТАВ ЗЕРНА

The study of the influence of the genome composition of triticale lines with genetic material Aegilops diploid species on the biochemical composition of grain was made. The high content of protein and starch in the grain was detected in triticale lines, in which genomes aegilops foreign material is present either as an entire chromosome, or a translocation. The increase in the content of protein and essential amino acids is observed for the line carrying the 2R(2U) chromosome substitution. Thus, the triticale genome reorganization, which has occurred as a result of introgressive hybridization of hexaploid triticale and genome-substituted forms of wheat, contributes to the creation of new economically valuable forms with a high content of protein, essential amino acids, and starch in the grain.Проведено изучение влияния состава генома линий тритикале с генетическим материалом диплоидных видов Aegilops на биохимический состав зерна. Высокое содержание белка и крахмала в зерне выявлено у линий тритикале,, в геномах которых чужеродный материал эгилопса присутствует либо в виде целой хромосомы, либо в виде транслокации. Линия тритикале с замещением хромосомы 2R на хромосому 2U Ae. umbellulata превышала родительский сорт тритикале не только по содержанию белка, но по количеству незаменимых аминокислот. Таким образом, реорганизация генома тритикале, произошедшая в результате интрогрессивной гибридизации гексаплоидных тритикале с геномно-замещенными формами мягкой пшеницы, способствует созданию новых хозяйственно ценных форм с повышенным содержанием белка, незаменимых аминокислот и крахмала в зерне

Informative EST-SSR markers for genotyping and intraspecific differentiation of Brassica oleracea var. capitata L.

Brassica oleraceae var. capitata L. is characterized by a high level of intraspecific heterogeneity due to some biological features that cause difficulties for breeding creating genetically homogenous forms and maintaining their genetic purity. Microsatellites (SSR) are highly polymorphic markers of plant genomes and represent one of the most effective tools for assessing genetic polymorphism. Among microsatellites, EST-SSR are most interesting, because they are directly linked to the expressed sequences and for that reason are widely used for analysis of genetic diversity and population structure. In this work, we studied the effectiveness of the use of transferable EST-SSR markers for both analyzing white cabbage diversity and genotyping pure lines. As a result, 15 microsatellite loci were characterized for the information content, allelic frequencies and heterogeneity levels. The effective multiallelic markers (Bo20TR, BoDCTD4, BoPC34, BoPLD1, BoCalc, BoPC15) with high information content (PIC > 0.7) that could be successfully used for analysis of inter- and intravarietal polymorphism in B. oleracea var. capitata were identified. It has been shown that intervarietal polymorphism expressed as the allelic diversity of EST SSR loci greatly facilitates varietal identification and typing of individual plants for breeding purposes. Based on the SSR-evaluation and subsequent clustering, the genetic structure of the breeding collection was identified, which showed that most experimental forms, in spite of different origin, have a common ancestral genetic basis. The identified donors of rare alleles could potentially be a source of valuable genetic segregation for further B. oleracea breeding improvement

Molecular-cytogenetic analysis of triticale and wheat lines with introgressions of the tribe Triticeae species genetic material

There are a number of problems in selection of cultivated cereals associated with the requirements to create forms with resistance to diseases, pests and unfavorable environmental conditions. The genetic diversity of genes for resistance to biotic and abiotic stresses can be increased by means of the gene pool of wild and cultivated wheat relatives. To improve agronomic traits in cereals, we have developed common wheat hybrid lines T. aestivum/ T. durum, T. aestivum/ T. dicoccoides and triticale lines by crossing hexaploid triticale with common wheat forms with the substitution of genome D for the geno­me of diploid Aegilops species. The aim of the study was to identify the lines of common wheat and hexaploid triticale with alien introgression using cytological and molecular-genetic analyses and evaluation of their cytological stability. Comparative analysis of the structure of chromosomes by GISH and FISH methods, microsatellite- and chromosome-specific markers revealed that hybridization of triticale with genome-substitution forms of wheat leads to the reorganization of the genome, including both the introgression of foreign material and wheat chromosome rearrangements, which lead to new combinations of genetic loci. The efficiency of wheat microsatellite markers to characterize of the T. aestivum/ T. durum, T. aestivum/ T. dicoccum interspecific hybrid lines was shown. From 4 to 12 translocations of different lengths from T. durum and T. dicoccum were identified in the chromosomes of A and В genomes in the hybrid lines. Meiotic stability of wheat and triticale hybrids was found. It creates prerequisites for preservation of alien genetic material in subsequent generations

SNP-АНАЛИЗ ГЕНЕТИЧЕСКОГО РАЗНООБРАЗИЯ ПШЕНИЦЫ БЕЛАРУСИ

We used a high-throughput array to evaluate the diversity of hexaploid wheat growing in Belarus under the breeding program through 384 gene-associated SNPs. The gene pool of winter and spring varieties are significantly different in frequency of 248 variants from 174 SNPs. The genetic structure of the Belarusian population of wheat has appeared similar to the Russian and Ukrainian varieties and is essentially different from west-European varieties. But it has a high variability and, consequently, a good genetic potential for the improvement through breeding.Проведено изучение генетического разнообразия пшеницы Беларуси на основе высокопропускного SNP маркирования. В целом из 384 использованных маркеров в исследуемой коллекции типирован 331 локус. Генофонды озимых и яровых сортов достоверно различаются по частотам 248 вариантов 174 SNP. Генетическая структура белорусских сортов обнаруживает значительное сходство с образцами российской и украинской селекции, но при этом обладает значительным запасом разнообразия, которое представляет хороший потенциал для создания новых высокопродуктивных адаптированных форм