Crop-wild Hybrids Research Articles

Morphology and fitness components of wild × crop F1hybrids ofSorghum bicolor(L.) in Ethiopia: implications for survival and introgression of crop genes in the wild pool

Natural hybridization between wild/weedy and crop species often results in rare hybrids, which can be more weedy and difficult to control. Moreover, the advent of transgenic crop plants raises questions of biosafety risk assessment on the consequences of rare hybrids with possible fitness enhancing genes on the environment. This study aimed at measuring the fitness components of wild–crop sorghum hybrids for various juvenile survival and adult morphological and fertility characters as part of the risk assessment of transgenic sorghum in Africa where the crop was believed to have first domesticated and serves as the major staple. Out of a pool of hybrids made in 2010 from 23 wild sorghum accessions and two released cultivated sorghum varieties using hand emasculation techniques, seven were selected for the field study of their fitness components in 2011. The study confirmed that crop–wild hybrids of sorghum are fertile. Two approaches were followed (relative fitness and mid-parent heterosis) which showed that most of the hybrids were as fit as their wild parents, and in some cases they showed mid-parent heterosis for the measured traits. The results of this study highlighted a potential risk that hybrids carrying crop genes (including herbicide resistance transgenes) could pose because they could be more weedy than their wild/weedy parents if transgenic sorghum is deployed in regions where the wild and cultivated sorghum populations coexist, such as in Ethiopia and in other parts of Africa.

Genomic and environmental selection patterns in two distinct lettuce crop–wild hybrid crosses

Genomic selection patterns and hybrid performance influence the chance that crop (trans)genes can spread to wild relatives. We measured fitness(-related) traits in two different field environments employing two different crop–wild crosses of lettuce. We performed quantitative trait loci (QTL) analyses and estimated the fitness distribution of early- and late-generation hybrids. We detected consistent results across field sites and crosses for a fitness QTL at linkage group 7, where a selective advantage was conferred by the wild allele. Two fitness QTL were detected on linkage group 5 and 6, which were unique to one of the crop–wild crosses. Average hybrid fitness was lower than the fitness of the wild parent, but several hybrid lineages outperformed the wild parent, especially in a novel habitat for the wild type. In early-generation hybrids, this may partly be due to heterosis effects, whereas in late-generation hybrids transgressive segregation played a major role. The study of genomic selection patterns can identify crop genomic regions under negative selection across multiple environments and cultivar–wild crosses that might be applicable in transgene mitigation strategies. At the same time, results were cultivar-specific, so that a case-by-case environmental risk assessment is still necessary, decreasing its general applicability.

The red queen in the corn: agricultural weeds as models of rapid adaptive evolution

Weeds are among the greatest pests of agriculture, causing billions of dollars in crop losses each year. As crop field management practices have changed over the past 12 000 years, weeds have adapted in turn to evade human removal. This evolutionary change can be startlingly rapid, making weeds an appealing system to study evolutionary processes that occur over short periods of time. An understanding of how weeds originate and adapt is needed for successful management; however, relatively little emphasis has been placed on genetically characterizing these systems. Here, we review the current literature on agricultural weed origins and their mechanisms of adaptation. Where possible, we have included examples that have been genetically well characterized. Evidence for three possible, non-mutually exclusive weed origins (from wild species, crop-wild hybrids or directly from crops) is discussed with respect to what is known about the microevolutionary signatures that result from these processes. We also discuss what is known about the genetic basis of adaptive traits in weeds and the range of genetic mechanisms that are responsible. With a better understanding of genetic mechanisms underlying adaptation in weedy species, we can address the more general process of adaptive evolution and what can be expected as we continue to apply selective pressures in agroecosystems around the world.

How to escape from crop‐to‐weed gene flow: phenological variation and isolation‐by‐time within weedy sunflower populations

The evolution of crop-related weeds may be constrained by recurrent gene flow from the crop. However, flowering time variation within weedy populations may open the way for weed adaptation by allowing some weeds to escape from this constraint. We investigated this link between phenology, gene flow and adaptation in weedy sunflower populations that have recently emerged in Europe from crop-wild hybridization. We studied jointly flowering phenology and genetic diversity for 15 microsatellite loci in six cultivated sunflower (Helianthus annuus) fields infested by weedy sunflower populations. The flowering overlap of cultivated and weedy sunflowers varied between and within populations: some weedy individuals were found to be completely isolated from the crop, the frequency of these plants being higher in populations from highly infested fields. Within weedy populations, we detected a pattern of isolation-by-time: the genetic divergence between individuals was positively correlated with their divergence in flowering period. In addition, earlier weeds, which flowered synchronously with the crop, were genetically more similar than late-flowering weeds to the cultivated varieties. Overall, our results suggest that crop-to-weed gene flow occurred, but was limited by divergent phenologies. We discuss the roles of weed adaptation and population history in the generation of this partial reproductive isolation.

Morphological markers for the detection of introgression from cultivated into wild carrot (Daucus carotaL.) reveal dominant domestication traits

Hybridisation and subsequent introgression have recently received much attention in the context of genetically modified crops. But crop-wild hybrid detection in the field can be difficult, as most domestication traits seem to be recessive, and the hybrid phenotype may also depend on the direction of the cross or environmental factors. Our aim was to develop a reliable set of morphological markers that differ between two wild and 13 cultivated carrots (Daucus carota L.) and to evaluate their inheritance in hybrid lines. We then examined these morphological markers in four F1 hybrids obtained by fertilising plants from the two wild accessions with pollen from two common carrot cultivars. Of the 16 traits that differed between the two carrot subspecies, three took intermediate values in the hybrids, eight resembled the cultivar parent (dominant domestication traits), two resembled the wild parent (domestication traits recessive), and three were not significant or growth condition-dependent. Root:shoot ratio was seven times higher for cultivars than for wild plants, while still attaining equivalent total dry weight, which shows that dry matter production by the shoot is much higher in cultivars than in wild plants. High root:shoot ratios were also present in the hybrids. While we found no maternal effects, the type of cultivar used for pollination had an impact on hybrid characteristics. The morphological markers developed here provide insights into the mode of inheritance of ecologically relevant traits and can be useful for pre-screening wild populations for hybrid detection prior to genetic analysis.

Population genetic structure of in situ wild Sorghum bicolor in its Ethiopian center of origin based on SSR markers

In situ population studies of wild relatives of crops are crucial for the conservation of plant genetic resources, especially in regions with high genetic diversity and a risk of local extinction. Ethiopia is the center of origin for sorghum, yet little is known about the genetic structure of extant wild populations. Using 9 Simple Sequence Repeat loci, we characterized 19 wild populations from five regions, 8 local cultivar populations from three regions, and 10 wild sorghum accessions from several African countries. To our knowledge, this is the most comprehensive study to date of in situ wild sorghum populations in Africa. Genetic diversity corrected for sample size was significantly greater in the wild populations in situ than in local cultivars or the accessions. Approximately 41 % of the genetic variation in the wild plants was partitioned among populations, indicating a high degree of differentiation and potential value for germplasm conservation, and the average number of migrants (Nm) per generation was 0.43. Cluster analyses showed that some wild populations were grouped by geographic region, whereas others were not, presumably due to long-distance seed movement. Four wild populations from disjunct regions formed a unique cluster with an Ethiopian accession of subsp. drummondii and probably represent a weedy race. STRUCTURE and other analyses detected evidence for crop-wild hybridization, consistent with previous molecular marker studies in Kenya, Mali, and Cameroon. In summary, in situ wild sorghum populations in Ethiopia harbor substantial genetic diversity and differentiation, despite their close proximity to conspecific cultivars in this crop/wild/weedy complex.

Les tournesols adventices : un exemple d’évolution d’une mauvaise-herbe apparentée à une espèce cultivée – Synthèse des recherches menées sur les populations adventices de tournesol en France depuis 2004

Weedy sunflowers have been officially reported in South-Western France in 2004. They display a combination of phenotypic traits of the wild and domesticated forms of the species and infest between 15 and 20% of fields of sunflower crop in this area, although at variable levels. When the infestation is strong, it affects seed yield and oil quality. Molecular studies showed that weedy sunflowers most probably resulted from accidental crop-wild hybridization during the seed production process and from the introduction of the resulting hybrids into commercial seed lots. Multiple independent introductions were at the source of the number of infested fields observed nowadays. The temporal dynamics of the infestation of a field and the detailed conditions for its success remain largely unknown. The flowering synchrony between crops and weeds is substantial, even if a variable fraction of the weeds (15-55%) flower completely outside the crop flowering period. Molecular survey of adult weedy plants and their progenies showed that crop-toweed gene flow occurs, at a rate that can reach 35 %on average, at the peak of the crop flowering period. We draw perspectives on the durability of herbicide-tolerant sunflower varieties as a solution to control these weeds, and call for more studies tackling weed management from an evolutionary lens.

Hybridization between crops and wild relatives: the contribution of cultivated lettuce to the vigour of crop-wild hybrids under drought, salinity and nutrient deficiency conditions.

With the development of transgenic crop varieties, crop–wild hybridization has received considerable consideration with regard to the potential of transgenes to be transferred to wild species. Although many studies have shown that crops can hybridize with their wild relatives and that the resulting hybrids may show improved fitness over the wild parents, little is still known on the genetic contribution of the crop parent to the performance of the hybrids. In this study, we investigated the vigour of lettuce hybrids using 98 F2:3 families from a cross between cultivated lettuce and its wild relative Lactuca serriola under non-stress conditions and under drought, salinity and nutrient deficiency. Using single nucleotide polymorphism markers, we mapped quantitative trait loci associated with plant vigour in the F2:3 families and determined the allelic contribution of the two parents. Seventeen QTLs (quantitative trait loci) associated with vigour and six QTLs associated with the accumulation of ions (Na+, Cl− and K+) were mapped on the nine linkage groups of lettuce. Seven of the vigour QTLs had a positive effect from the crop allele and six had a positive effect from the wild allele across treatments, and four QTLs had a positive effect from the crop allele in one treatment and from the wild allele in another treatment. Based on the allelic effect of the QTLs and their location on the genetic map, we could suggest genomic locations where transgene integration should be avoided when aiming at the mitigation of its persistence once crop–wild hybridization takes place.Electronic supplementary materialThe online version of this article (doi:10.1007/s00122-012-1897-4) contains supplementary material, which is available to authorized users.

Crop to wild introgression in lettuce: following the fate of crop genome segments in backcross populations.

BackgroundAfter crop-wild hybridization, some of the crop genomic segments may become established in wild populations through selfing of the hybrids or through backcrosses to the wild parent. This constitutes a possible route through which crop (trans)genes could become established in natural populations. The likelihood of introgression of transgenes will not only be determined by fitness effects from the transgene itself but also by the crop genes linked to it. Although lettuce is generally regarded as self-pollinating, outbreeding does occur at a low frequency. Backcrossing to wild lettuce is a likely pathway to introgression along with selfing, due to the high frequency of wild individuals relative to the rarely occurring crop-wild hybrids. To test the effect of backcrossing on the vigour of inter-specific hybrids, Lactuca serriola, the closest wild relative of cultivated lettuce, was crossed with L. sativa and the F1 hybrid was backcrossed to L. serriola to generate BC1 and BC2 populations. Experiments were conducted on progeny from selfed plants of the backcrossing families (BC1S1 and BC2S1). Plant vigour of these two backcrossing populations was determined in the greenhouse under non-stress and abiotic stress conditions (salinity, drought, and nutrient deficiency).ResultsDespite the decreasing contribution of crop genomic blocks in the backcross populations, the BC1S1 and BC2S1 hybrids were characterized by a substantial genetic variation under both non-stress and stress conditions. Hybrids were identified that performed equally or better than the wild genotypes, indicating that two backcrossing events did not eliminate the effect of the crop genomic segments that contributed to the vigour of the BC1 and BC2 hybrids. QTLs for plant vigour under non-stress and the various stress conditions were detected in the two populations with positive as well as negative effects from the crop.ConclusionAs it was shown that the crop contributed QTLs with either a positive or a negative effect on plant vigour, we hypothesize that genomic regions exist where transgenes could preferentially be located in order to mitigate their persistence in natural populations through genetic hitchhiking.

Genomic regions in crop-wild hybrids of lettuce are affected differently in different environments: implications for crop breeding.

Many crops contain domestication genes that are generally considered to lower fitness of crop–wild hybrids in the wild environment. Transgenes placed in close linkage with such genes would be less likely to spread into a wild population. Therefore, for environmental risk assessment of GM crops, it is important to know whether genomic regions with such genes exist, and how they affect fitness. We performed quantitative trait loci (QTL) analyses on fitness(-related) traits in two different field environments employing recombinant inbred lines from a cross between cultivated Lactuca sativa and its wild relative Lactuca serriola. We identified a region on linkage group 5 where the crop allele consistently conferred a selective advantage (increasing fitness to 212% and 214%), whereas on linkage group 7, a region conferred a selective disadvantage (reducing fitness to 26% and 5%), mainly through delaying flowering. The probability for a putative transgene spreading would therefore depend strongly on the insertion location. Comparison of these field results with greenhouse data from a previous study using the same lines showed considerable differences in QTL patterns. This indicates that care should be taken when extrapolating experiments from the greenhouse, and that the impact of domestication genes has to be assessed under field conditions.

No effect of transgene and strong wild parent effects on seed dormancy in crop-wild hybrids of rice: implications for transgene persistence in wild populations

Soil seed banks act as a gene pool for local plant species and, as such, can buffer local populations, especially those experiencing challenging environmental conditions. Seed dormancy has important implications to dynamics of soil seed banks. Therefore, estimating the seed dormancy of transgenic crop–wild hybrids could shed light on the persistence of transgenes in wild-plant soil seed banks. Individuals from eight populations of wild rice Oryza rufipogon were crossed with those of three insect-resistant transgenic rice lines. Selfed (F2–F4) and backcrossed populations (BC1, BC1F2 and BC1F3) were then made from the hybrids. Seed germination was tested under three treatments: (a) normal; (b) overwintering in soil; and (c) one-week heat-shocking. The effects of transgene, wild parent and hybrid generation on hybrid seed germination were examined. No significant effect of insect-resistant transgenes (Bt and CpTI) was detected on the seed dormancy of crop–wild hybrids, while a significant wild parent effect was found. The seeds of advanced generation hybrids have higher germination percentages and lower dormancy than do those of F1 and BC1 generations. The study showed that the dormancy of hybrid seeds was determined mainly by their genetic backgrounds. All hybrid seeds have higher germination percentages and lower dormancy (and, consequently, a poorer overwintering ability), compared with wild seeds, and reduce dormancy would contribute to a fitness disadvantage, compared with wild types. Therefore, such seeds might form part of naturally occurring soil seed banks, through which crop genes would persist in wild populations.

Metapopulation structure and fine-scaled genetic structuring in crop-wild hybrid weed beets

This study explores the microspatial and temporal genetic variation in crop-wild hybrid weed beets that emerged from the seed bank in a cultivated field surveyed over two successive years. We demonstrate the occurrence of demes highly genetically differentiated, kin-structured, characterized by moderate effective population sizes, differing in propensity for selfing, and arising from nonrandom genetic subsets of the seed bank. Only one deme identified in the first survey year significantly contributed to the weed beets that emerged in the second year. Spatial structuring appears to be primarily due to gravity seed dispersal and limited pollen flow among weed beet demes. Within each genetic cluster identified by Bayesian assignments and multivariate analyses, F(IS) estimates and level of biparental inbreeding--revealed by progeny analyses--dropped to non-significant values. This suggests that random mating occurs at the scale of genetically distinct demes over a very short scale. Our results highlight the need to carefully depict genetic discontinuities in weed species, when attempting to describe their local genetic neighborhoods within which genetic drift and selective processes occur.

Sunflower crop-wild hybrids: Identification and risks

Wild sunflower, Helianthus annuus, is an open-pollinated species that can be a major weed in several crops. If it hybridizes with cultivated sunflower the following generations can generate crop-wild hybrids that could evolve into invasive populations. The objective of this study was to morphologically characterize F1 hybrids between inbred lines and wild sunflower, their F2, BC1 and BC2 progenies with domestic sunflower to assess the risk of these plants evolving into invasive biotypes. Crop-wild hybrids were easily identified as off-type plants. F1 plants were taller, branched with several heads, smaller disks and phyllaries, stigma and pale anthocyanins, and a longer flowering period compared to cultivated sunflower. None of the F2 plants were similar to wild sunflower plants, but morphologically closer to the cultivated materials. These results showed the improbability that crop-wild off-type plants give rise to plants morphologically similar to wild sunflowers and therefore the risk of developing invasive populations by their introduction.

Construction of a crop—wild hybrid population for broadening genetic diversity in cultivated sunflower and first evaluation of its combining ability: the concept of neodomestication

The genetic base of sunflower elite lines is very narrow, due to many years of selection and breeding. To broaden the genetic diversity of the cultivated sunflower, in 1995 73 wild sunflower populations were crossed with 3 cultivated lines (Testers), and 219 hybrid offspring’s were evaluated in the field. GCA and SCA effects were computed suggesting for all traits a genetic potential for improvement through selection. Study of the hybrids revealed that the wild accessions bear different genetic abilities to combine with the testers for traits of morphological architecture, phenology and yield (seed weight and seed oil). The variance due to GCA and SCA showed that gene action was additive for days to flowering, branching and plant height. Genotypes derived from the same geographic origin may have either good or poor general combing ability. The correlation between GCA and per se genotype performance was positive for all traits except for seed oil content. This was the first attempt to evaluate wild-cultivated hybrids in sunflower on a large scale and will be the starting point for the management of hybrid Helianthus annuus populations for breeding. GCA and SCA estimations will facilitate the definition of strategies to manage and exploit the natural diversity for this crop.

The origin and evolution of a recent agricultural weed: population genetic diversity of weedy populations of sunflower (Helianthus annuus L.) in Spain and France.

The recurrent evolution of crop-related weeds during agricultural history raises serious economic problems and challenging scientific questions. Weedy forms of sunflower, a species native from America, have been reported in European sunflower fields for a few decades. In order to understand their origin, we analysed the genetic diversity of a sample of weedy populations from France and Spain, and of conventional and ornamental varieties. A crop-specific maternally inherited marker was present in all weeds. At 16 microsatellite loci, the weedy populations shared most of their diversity with the conventional varieties. But they showed a large number of additional alleles absent from the cultivated pool. European weedy populations thus most probably originated from the unintentional pollination of maternal lines in seed production fields by wild plants growing nearby, resulting in the introduction of crop-wild hybrids into the farmers’ fields. The wide diversity and the low population structure detected were indicative of a multiplicity of introductions events rather than of field-to-field propagation. Further studies are required to understand the local evolutionary dynamics of a weedy population, and especially the respective roles of crop-to-weed gene flow and selection in the fate of an initial source of crop-wild hybrids.

Ecological patterns and genetic analysis of post-dispersal seed predation in sunflower (Helianthus annuus) crop-wild hybrids

Crop-wild hybridization has been documented in many cultivated species, but the ecological and genetic factors that influence the likelihood or rate that cultivar alleles will introgress into wild populations are poorly understood. Seed predation is one factor that could mitigate the spread of otherwise advantageous cultivar alleles into the wild by reducing seedling recruitment of crop-like individuals in hybrid populations. Seed predation has previously been linked to several seed characters that differ between cultivated and wild sunflower, such as seed size and oil content. In this study, seed morphological and nutritional characters were measured in a segregating population of sunflower crop-wild hybrids and wild and cultivated lines. Seed predation rates among lines were then assessed in the field. The relationship between seed predation and seed characters was investigated and quantitative trait loci (QTL) were mapped for all traits. There was no effect of seed type (hybrid vs. parents) on seed predation, although a trend toward more early predation of wild seeds was observed. Within the hybrids, seed predators preferred seeds that contained more oil and energy but were lower in fibre. The relationship between seed predation and oil content was supported by co-localized QTL for these traits on one linkage group. These results suggest that oil content may be a more important determinant of seed predation than seed size and provide molecular genetic evidence for this relationship. The cultivar allele was also found to increase predation at all QTL, indicating that post-dispersal seed predation may mitigate the spread of cultivar alleles into wild populations.

Populations of weedy crop–wild hybrid beets show contrasting variation in mating system and population genetic structure

Reproductive traits are key parameters for the evolution of invasiveness in weedy crop–wild hybrids. In Beta vulgaris, cultivated beets hybridize with their wild relatives in the seed production areas, giving rise to crop–wild hybrid weed beets. We investigated the genetic structure, the variation in first-year flowering and the variation in mating system among weed beet populations occurring within sugar beet production fields. No spatial genetic structure was found for first-year populations composed of F1 crop–wild hybrid beets. In contrast, populations composed of backcrossed weed beets emerging from the seed bank showed a strong isolation-by-distance pattern. Whereas gametophytic self-incompatibility prevents selfing in wild beet populations, all studied weed beet populations had a mixed-mating system, plausibly because of the introgression of the crop-derived Sf gene that disrupts self-incompatibility. No significant relationship between outcrossing rate and local weed beet density was found, suggesting no trends for a shift in the mating system because of environmental effects. We further reveal that increased invasiveness of weed beets may stem from positive selection on first-year flowering induction depending on the B gene inherited from the wild. Finally, we discuss the practical and applied consequences of our findings for crop-weed management.

A comparative study of invasive Helianthus annuus populations in their natural habitats of Argentina and Spain

Wild Helianthus annuus is native to North America but it naturalized in other parts of the world as well. Although the origin of exotic populations is uncertain, they have probably evolved very differently in different countries. To unravel the origin of invasive populations from Argentina and Spain, morphological and agro-ecological data of nine populations from central Argentina, six from Andalusia and one from Gerona were collected in their natural habitats during three exploration trips in 2007 and 2008. In Argentina wild H. annuus was found mainly in disturbed areas between roads and fences. In a few cases the populations were located on the margins of cultivated fields. The Argentinean populations are spread across more than 50,000 m2 at a density of about 25 plants m-2. In Spain, the populations were found mainly in croplands. The largest population covered about 1,500 m2 and comprised no more than 200 plants. The Argentinean populations had taller plants with a higher number of heads of small size, while the Spanish populations were characterized by bigger heads with wider ligules and bracts. Plants were shorter and leaf size was larger in Gerona than in Andalusia. Multivariate analysis differentiated populations from Argentina and Spain by many traits. Wild-crop gene flow is likely the source of genetic variation among them. In Argentina, the populations keep the appearance of early wild introductions, while the Spanish populations are weedier and probably originated from pollen contamination of commercial seed with wild plants or crop-wild hybrids.

Existence of vigorous lineages of crop-wild hybrids in Lettuce under field conditions.

Plant to plant gene flow is a route of environmental exposure for GM plants specifically since crosses with wild relatives could lead to the formation of more vigorous hybrids, which could increase the rate of introgression and the environmental impact. Here, we test the first step in the process of potential transgene introgression: whether hybrid vigor can be inherited to the next generation, which could lead to fixation of altered, i.e., elevated, quantitative traits. The potential for a permanent elevated fitness was tested using individual autogamous progeny lineages of hybrids between the crop Lactuca sativa (Lettuce) and the wild species Lactuca serriola (Prickly Lettuce). We compared progeny from motherplants grown under either greenhouse or field conditions. The survival of young plants depended strongly on maternal environment. Furthermore, we observed that offspring reproductive fitness components were correlated with maternal fitness. Our study demonstrates that post-zygotic genotypic sorting at the young plants stage reduces the number of genotypes non-randomly, leading to inheritance of high levels of reproductive traits in the surviving hybrid lineages, compared to the pure wild relatives. Consequently, directional selection could lead to displacement of the pure wild relative and fixation of more vigorous genome segments originating from crops, stabilizing plant traits at elevated levels. Such information can be used to indentify segments which are less likely to introgress into wild relative populations as a target for transgene insertion.

Fitness effects and genetic architecture of plant–herbivore interactions in sunflower crop–wild hybrids

*Introgression of cultivar alleles into wild plant populations via crop-wild hybridization is primarily governed by their fitness effects as well as those of linked loci. The fitness of crop-wild hybrids is often dependent on environmental factors, but less is understood about how aspects of the environment affect individual cultivar alleles. *This study investigated the effects of naturally occurring herbivory on patterns of phenotypic selection and the genetic architecture of plant-herbivore interactions in an experimental sunflower crop-wild hybrid population in two locales. *Phenotypic selection analyses suggested that cultivar alleles conferring increased size were generally favored, but at one site cultivar-like flowering time was favored only if three types of herbivory were included in the selection model. Quantitative trait locus (QTL) mapping identified three regions in which the cultivar allele conferred a selective advantage for a number of co-localized traits. Quantitative trait loci for several measures of insect herbivory were detected and, although the cultivar allele increased herbivory damage at the majority of these QTLs, they rarely colocalized with advantageous cultivar alleles for morphological traits. *These results suggest that a subset of cultivar traits/alleles are advantageous in natural environments but that herbivory may mitigate the selective advantage of some cultivar alleles.