Apomixis in agriculture: the quest for clonal seeds

Apomixis, or asexual reproduction through seeds, is a natural trait that could have an immense positive impact on crop production. Apomictic breeding strategies could allow the fixation and indefinite propagation of any desired genotype, however complex. Apomicts display a wide variety of developmental mechanisms, which can be viewed as a short-circuiting of sexual development. Gametophytic and sporophytic apomixis are distinguished by the developmental origin of apomictically derived embryos. Genetic studies suggest that individual elements of gametophytic apomixis, such as apomeiosis and parthenogenesis, are either controlled by one or two dominant Mendelian factors. As recombination around apomeiosis loci is suppressed, it is currently not known how complex these loci are. Much less is known regarding the genetic control of sporophytic apomixis but initial studies suggest a complex genetic control. Genetic analyses of sexual reproduction in plant model systems have identified genes that, when mutated, display elements of apomixis. Such studies help in the identification of candidate genes and promoters that can be used for the de novo engineering of apomixis through biotechnology. Molecular genetic studies in apomictic and sexual systems will generate the knowledge necessary for the engineering of conditional apomixis technology. Approaches encouraging collaboration and widespread dissemination of the acquired knowledge will constitute the most innovative route to the development, deployment and acceptance of apomixis technology in agriculture.     

Genetic linkage maps of the chromosomal regions associated with apomictic and sexual modes of reproduction in Cenchrus ciliaris

Cenchrus ciliaris reproduces by apomixis, an asexual mode of reproduction through seeds. Genetic analysis of apomixis in this species revealed that this trait is dominant and that a chromosomal region of more than 11?Mb controls this trait, which is hemizygous, heterochromatic and recombinationally suppressed. A novel F₂ mapping population comprising 86 individuals segregating for apomictic and sexual modes of reproduction, generated after crossing a new set of obligate apomictic and sexual parents (IG-96-3108 and IG-96-443), was used in this study to identify a large number of amplified fragment length polymorphism (AFLP) and sequence characterized amplified region (SCAR) markers linked to these traits. Out of 180 polymorphic AFLP markers, 42 and 29 markers associated with apomixis and sexuality were mapped around Apo and Sexual loci, respectively. Markers 20G, 18G and 19G showed close linkage to Apo locus at map distance of only 1.1?cM, while 12FS, 4HS and 12b showed tight linkage to Sexual locus at map distance of 1.7?cM. Markers clustered around Apo and Sexual loci on either side. A large number of recombining AFLP markers were mapped around both loci, indicating a minor role of suppression of recombination. Four anchor markers from earlier studies also clustered around Apo locus, validating the present genetic linkage map. In addition, seven and one SCAR markers closely linked to Apo and Sexual loci were also developed, which could be used for fine mapping of the loci.     

Sexual and apomictic plant reproduction in the genomics era: exploring the mechanisms potentially useful in crop plants

Arabidopsis, Mimulus and tomato have emerged as model plants in researching genetic and molecular basis of differences in mating systems. Variations in floral traits and loss of self-incompatibility have been associated with mating system differences in crops. Genomics research has advanced considerably, both in model and crop plants, which may provide opportunities to modify breeding systems as evidenced in Arabidopsis and tomato. Mating system, however, not recombination per se, has greater effect on the level of polymorphism. Generating targeted recombination remains one of the most important factors for crop genetic enhancement. Asexual reproduction through seeds or apomixis, by producing maternal clones, presents a tremendous potential for agriculture. Although believed to be under simple genetic control, recent research has revealed that apomixis results as a consequence of the deregulation of the timing of sexual events rather than being the product of specific apomixis genes. Further, forward genetic studies in Arabidopsis have permitted the isolation of novel genes reported to control meiosis I and II entry. Mutations in these genes trigger the production of unreduced or apomeiotic megagametes and are an important step toward understanding and engineering apomixis.     

Evolution of genetic variability and the advantage of sex and recombination in changing environments.

The role of recombination and sexual reproduction in enhancing adaptation and population persistence in temporally varying environments is investigated on the basis of a quantitative-genetic multilocus model. Populations are finite, subject to density-dependent regulation with a finite growth rate, diploid, and either asexual or randomly mating and sexual with or without recombination. A quantitative trait is determined by a finite number of loci at which mutation generates genetic variability. The trait is under stabilizing selection with an optimum that either changes at a constant rate in one direction, exhibits periodic cycling, or fluctuates randomly. It is shown by Monte Carlo simulations that if the directional-selection component prevails, then freely recombining populations gain a substantial evolutionary advantage over nonrecombining and asexual populations that goes far beyond that recognized in previous studies. The reason is that in such populations, the genetic variance can increase substantially and thus enhance the rate of adaptation. In nonrecombining and asexual populations, no or much less increase of variance occurs. It is explored by simulation and mathematical analysis when, why, and by how much genetic variance increases in response to environmental change. In particular, it is elucidated how this change in genetic variance depends on the reproductive system, the population size, and the selective regime, and what the consequences for population persistence are.     

Mapping quantitative trait loci in selected breeding populations: A segregation distortion approach

Quantitative trait locus (QTL) mapping is often conducted in line-crossing experiments where a sample of individuals is randomly selected from a pool of all potential progeny. QTLs detected from such an experiment are important for us to understand the genetic mechanisms governing a complex trait, but may not be directly relevant to plant breeding if they are not detected from the breeding population where selection is targeting for. QTLs segregating in one population may not necessarily segregate in another population. To facilitate marker-assisted selection, QTLs must be detected from the very population which the selection is targeting. However, selected breeding populations often have depleted genetic variation with small population sizes, resulting in low power in detecting useful QTLs. On the other hand, if selection is effective, loci controlling the selected trait will deviate from the expected Mendelian segregation ratio. In this study, we proposed to detect QTLs in selected breeding populations via the detection of marker segregation distortion in either a single population or multiple populations using the same selection scheme. Simulation studies showed that QTL can be detected in strong selected populations with selected population sizes as small as 25 plants. We applied the new method to detect QTLs in two breeding populations of rice selected for high grain yield. Seven QTLs were identified, four of which have been validated in advanced generations in a follow-up study. Cloned genes in the vicinity of the four QTLs were also reported in the literatures. This mapping-by-selection approach provides a new avenue for breeders to improve breeding progress. The new method can be applied to breeding programs not only in rice but also in other agricultural species including crops, trees and animals.     

Genetic analysis of apomixis in Citrus and Poncirus by molecular markers

Propagation of citrus rootstocks depends upon the production of clonal plants from nucellar seedlings. This makes apomixis one of the host important traits in breeding programs for citrus rootstocks. The genetic control of apomixis was studied in a 50-tree progeny derived from the cross C. volkameriana×P. trifoliata using 69 molecular markers and bulked segregant analysis. The proportion of nucellar seedlings was estimated by isoenzymatic analysis of 25 seedlings per tree for 2 consecutive years. The type of embryony (polyembryonic versus monoembryonic seeds) was also determined for fruit-yielding trees. Separate genetic maps for each parental species were developed. The integration and comparison of these maps could be accomplished using common multiallelic segregant loci. Differences in gene synteny between the two species-specific genetic maps were shown. Important distortions in the segregation of markers at several genomic regions, some of them also involving differences in the C-methylation pattern, have been observed, especially for the pollen parent. Analysis of quantitative trait loci (QTLs) revealed the presence of six genomic positions (two in P. trifoliata and four in C. volkameriana) contributing individually up to 24% of the total variation for apomixis. Within the same species, QTLs with positive and negative allele effects were present, even in the same linkage group. One of the markers associated to apomixis (Apo2) is also associated to embryony type. Therefore, the genetic control of apomictic reproduction found in citrus (nucellar embryony) is quite complex compared to what has been reported for gametophytic apomixis. Molecular markers linked to QTLs governing apomixis will be useful to assist selection of future apomictic rootstocks for citrus varieties. Received: 30 November 1998 / Accepted: 28 December 1998     

Partial isolation of the genomic region linked with apomixis in <Emphasis Type="Italic">Paspalum simplex</Emphasis>

Apomixis is a form of asexual reproduction through seed and has the potential to be applied, to great benefit, to agriculture. Understanding the genetic control of apomixis has proven to be a challenging task because the trait is mainly present in wild species and genetic mapping is often impaired by a block of recombination. A physical mapping approach has therefore been undertaken to unlock the genetic control of apomixis in Paspalum simplex Morong, a species with a relatively small genome and which exhibits a degree of genetic synteny with rice. In this paper, we report on the construction of a bacterial artificial chromosome library for Paspalum simplex with a coverage of approximately three genome equivalents and an average insert size of 94 kb. The BAC library was screened with 19 sequence characterized amplified region markers which were 100% linked to apomixis and a recombinant SCAR marker, all developed through a bulked segregant analysis strategy. A mini-sequencing procedure reported in the literature greatly aided the direct development of SCAR markers from amplified fragment length polymorphism bands. Several BAC clones linked to apomixis were identified and assembled into seven contigs and 18 singletons. Two of the BAC clones identified contained independently isolated markers. This is the first such report in an apomictic model that lacks recombination at the locus. We believe that extension of the contigs coupled to high-throughput sequencing will help the understanding of the genomic structure of the apomixis locus in P. simplex.     

A molecular map of the apomixis-control locus in <Emphasis Type="Italic">Paspalum procurrens</Emphasis> and its comparative analysis with other species of <Emphasis Type="Italic">Paspalum</Emphasis>

Since apomixis was first mapped in Paspalum, the absence of recombination that characterizes the related locus appeared to be the most difficult bottleneck to overcome for the dissection of the genetic determinants that control this trait. An approach to break the block of recombination was developed in this genus through an among-species comparative mapping strategy. A new apomictic species, P. procurrens (Q4094) was crossed with a sexual plant of P. simplex and their progeny was classified for reproductive mode with the aid of morphological, embryological and genetic analyses. On this progeny, a set of heterologous rice RFLP markers strictly co-segregating in coupling phase with apomixis was identified. These markers were all located on the telomeric region of the long arm of the chromosome 12 of rice. In spite of the lack of recombination exhibited by the apomixis-linked markers in P. procurrens, a comparative mapping analysis among P. simplex, P. malacophyllum, P. notatum and P. procurrens, allowed us to identify a small group of markers co-segregating with apomixis in all these species. These markers bracketed a chromosome region that likely contains all the genetic determinants of apomictic reproduction in Paspalum. The implications of this new inter-specific approach for overcoming the block of recombination to isolate the genetic determinants of apomixis and gain a better comprehension of genome structure of apomictic chromosome region are discussed.     

Molecular and genetic regulation of apomixis

Apomixis is defined as the asexual plant reproduction through seeds that results in the production of genetically uniform progeny. In fact, apomixis could be considered as a natural way of cloning. Currently there are more than 400 plant species known to use apomixis as a strategy for their propagation. The primary fundamental aspects of apomixis are the bypassing of meiosis and parthenogenetic development of the embryo without fertilization Apomixis attracts special attention because of its potential value for agriculture, as it could be harnessed for plant breeding programs enabling the permanent fixation of heterosis in crop plants. A better understanding of the molecular and genetic regulation of apomixis is important for developmental and evolutionary perspectives but also for implementation of engineering of apomixis traits into agricultural crop plants. Despite apomixis is considered as one of the key technologies for the improving agriculture, it is currently not fully known how the genetic and molecular regulation of this important trait occurs. In this review, an up to date information on the biology of apomixis and the known genes and genetic loci associated with regulation of different components of apomixis is provided.     

Deleterious Mutations as an Evolutionary Factor. II. Facultative Apomixis and Selfing

A population with u deleterious mutations per genome per generation is considered in which only those individuals that carry less than a critical number k of mutations are viable. Besides a large number of loci subject to mutation and selection, the genome contains one or two special loci responsible for the mode of reproduction. Amphimixis vs. apomixis and amphimixis vs. selfing are considered separately. In the first case, the genome degradation rate v (= u/square root k) is found to play the decisive role, as in the case of recombination. When v greater than 1.25, obligate amphimixis is established. If v decreases below this value, the alleles with first low and then larger penetrance are fixed, until alleles conferring obligate asexual reproduction become advantageous. The proportion of resources allocated to produce seeds also increases with decrease of v. These results are unlikely to depend on the genetic basis of the mode of reproduction. The result of competition between outcrossing and selfing depends on both u and k, as well as on whether the mutations are recessive. The alleles for selfing with low penetrance are selected against if the mutations are at all recessive. The fitness of alleles with high penetrance depends primarily on u, decreasing when u increases. There may exist conditions when only the alleles providing intermediate selfing rates can be fixed in a population. In other cases a population may exist with either obligate outcrossing or selfing at a high rate. Thus, truncation selection against deleterious mutations may be a factor supporting obligate or facultative sex despite the twofold advantage of apomixis or selfing.     

Are female reproductive status and male morphotypes of the giant freshwater prawn Macrobrachium rosenbergii altered by selection for high growth?

Understanding genetic changes in the hierarchical population structure of the giant freshwater prawn (GFP) Macrobrachium rosenbergii is important to improve productivity and profitability of prawn farming. We examined correlated responses in male morphotype and female reproduction status resulting from a selection programme for high growth in a GFP population undergoing four generations of selection. Two contemporary lines (selection and control) were maintained and produced simultaneously; the selection line was selected for high breeding values for harvest body weight and the control for average breeding values of the population. A total of 21,459 individual prawns were measured for body traits and morphotypes. Individual morphotype was recorded as a binary trait (0 and 1) and were analysed using a generalised threshold mixed model with a logit link function. Correlated genetic changes in three important male morphotypes (small, orange and blue claw males) were significant (p < 0.05–0.001) only in generation 2010 but not in the last generation (2012). Selection for high growth did not have an adverse effect on reproduction status of females, except that the selection line had a greater proportion of spawned females than the control (p = 0.01). Our results pointed out that selection for high growth may have had both positive and negative impacts on the social population structure of male morphotypes in GFP. Undesired changes in important morphotypes should be taken account of in genetic improvement programmes for GFP in order to minimise social competition effects and increase total biomass and consequently economic returns of the sector.     

Genetic fine-mapping of <Emphasis Type="Italic">DIPLOSPOROUS</Emphasis> in <Emphasis Type="Italic">Taraxacum</Emphasis> (dandelion; Asteraceae) indicates a duplicated <Emphasis Type="Italic">DIP</Emphasis>-gene

Background  

DIPLOSPOROUS (DIP) is the locus for diplospory in Taraxacum, associated to unreduced female gamete formation in apomicts. Apomicts reproduce clonally through seeds, including apomeiosis, parthenogenesis, and autonomous or pseudogamous endosperm formation. In Taraxacum, diplospory results in first division restitution (FDR) nuclei, and inherits as a dominant, monogenic trait, independent from the other apomixis elements. A preliminary genetic linkage map indicated that the DIP-locus lacks suppression of recombination, which is unique among all other map-based cloning efforts of apomeiosis to date. FDR as well as apomixis as a whole are of interest in plant breeding, allowing for polyploidization and fixation of hybrid vigor, respectively. No dominant FDR or apomixis genes have yet been isolated. Here, we zoom-in to the DIP-locus by largely extending our initial mapping population, and by analyzing (local) suppression of recombination and allele sequence divergence (ASD).     

不同QTL增效基因初始频率下标记辅助选择的效果

采用随机模拟方法模拟了在一个闭锁群体内连续对单个性状选择10个世代的情形。在假定选择性状受一个位于常染色体上的QTL和多基因共同控制的情况下,采用动物模型标记辅助最佳线性无偏预测方法估计个体育种值并据此进行种畜的选留,并在此基础上系统地比较了QTL增效基因初始频率对标记辅助选择效果的影响。结果表明：当群体中QTL增效基因的初始频率较低时,选择所获得的QTL基因型值的进展会更大,标记辅助选择在单位时间内可获得较大的遗传进展;此时,尽管QTL增效基因在群体中固定所需的世代数会更长一些,但其频率上升的速度却更快。而QTL增效基因初始频率的高低对群体近交增量的影响不是很大。     

High-resolution physical mapping in Pennisetum squamulatum reveals extensive chromosomal heteromorphism of the genomic region associated with apomixis

Gametophytic apomixis is asexual reproduction as a consequence of parthenogenetic development of a chromosomally unreduced egg. The trait leads to the production of embryos with a maternal genotype, i.e. progeny are clones of the maternal plant. The application of the trait in agriculture could be a tremendous tool for crop improvement through conventional and nonconventional breeding methods. Unfortunately, there are no major crops that reproduce by apomixis, and interspecific hybridization with wild relatives has not yet resulted in commercially viable germplasm. Pennisetum squamulatum is an aposporous apomict from which the gene(s) for apomixis has been transferred to sexual pearl millet by backcrossing. Twelve molecular markers that are linked with apomixis coexist in a tight linkage block called the apospory-specific genomic region (ASGR), and several of these markers have been shown to be hemizygous in the polyploid genome of P. squamulatum. High resolution genetic mapping of these markers has not been possible because of low recombination in this region of the genome. We now show the physical arrangement of bacterial artificial chromosomes containing apomixis-linked molecular markers by high resolution fluorescence in situ hybridization on pachytene chromosomes. The size of the ASGR, currently defined as the entire hemizygous region that hybridizes with apomixis-linked bacterial artificial chromosomes, was estimated on pachytene and mitotic chromosomes to be approximately 50 Mbp (a quarter of the chromosome). The ASGR includes highly repetitive sequences from an Opie-2-like retrotransposon family that are particularly abundant in this region of the genome.     

Both costs and benefits of sex correlate with relative frequency of asexual reproduction in cyclically parthenogenic Daphnia pulicaria populations

Sexual reproduction is generally believed to yield beneficial effects via the expansion of expressed genetic variation, which increases the efficiency of selection and the adaptive potential of a population. However, when nonadditive gene action is involved, sex can actually impede the adaptive progress of a population. If selection promotes coupling disequilibria between genes of similar effect, recombination and segregation can result in a decrease in expressed genetic variance in the offspring population. In addition, when nonadditive gene action underlies a quantitative trait, sex can produce a change in trait means in a direction opposite to that favored by selection. In this study we measured the change in genotypic trait means and genetic variances across a sexual generation in four populations of the cyclical parthenogen Daphnia pulicaria, which vary predictably in their incidence of sexual reproduction. We show that both the costs and benefits of sex, as measured by changes in means and variances in life-history traits, increase substantially with decreasing frequency of sex.     

Quantitative genetic variation in Daphnia: temporal changes in genetic architecture

Nonadditive genetic variation and genetic disequilibrium are two important factors that influence the evolutionary trajectory of natural populations. We assayed quantitative genetic variation in a temporary-pond-dwelling population of Daphnia pulex over a full season to examine the role of nonadditive genetic variation and genetic disequilibrium in determining the short-term evolutionary trajectory of a cyclic parthenogen. Quantitative traits were influenced by three factors: (1) clonal selection significantly changed the population mean phenotype during the course of the growing season; (2) sexual reproduction and recombination led to significant changes in life-history trait means and the levels of expressed genetic variation, implying the presence of substantial nonadditive genetic variation and genetic disequilibrium; and (3) Egg-bank effects were found to be an important component of the realized year-to-year change. Additionally, we examined the impact of genetic disequilibria induced by clonal selection on the genetic (co)variance structure with a common principal components model. Clonal selection caused significant changes in the (co)variance structure that were eliminated by a single bout of random mating, suggesting that a build-up of disequilibria was the primary source of changes in the (co)variance structure. The results of this study highlight the complexity of natural selection operating on populations that undergo alternating phases of sexual and asexual reproduction.     

Selection for litter size and litter birthweight in Large White pigs: Maximum,mean and variability of reproduction traits

Gradually increasing trend of litter size poses a challenge to pig farmers in terms of managing larger litters. Therefore, it seems that a balanced approach that optimises litter size, litter birthweight, and uniformity of those traits is needed in order to address animal welfare and farm management concerns. This study aimed to investigate this issue by defining several traits for total number born (TNB), number born alive (NBA) and litter birthweight (LW). First, the highest value from at least five records per sow was selected as maximum (max) value for each reproduction trait. Second, a mean (mean) for each reproduction trait was calculated per sow. Last, the variability of reproduction traits between parities of the sow was calculated as log-transformed variance of residuals of all observations per sow for each reproduction trait (LnVar). In total, 23 193 Large White sows from Topigs Norsvin with 152 282 litter records were used for analysis in ASReml 4.1. Also, a simulation of breeding schemes was performed with the use of SelAction 2.1 and estimates from genetic analysis. Maximum value of reproductive traits had a much higher heritability than repeated observations or mean of reproduction traits, e.g., 0.31 for maxTNB vs. 0.12 for TNB and 0.07 for meanTNB, which allows for a faster response under selection. The maximum value traits, however, were found to carry more risks, i.e. higher ratio of stillborn (not for maxNBA) and increased variability of traits. Thus, using them in breeding programme should be carefully considered. The genetic coefficient of variation on SD level estimated to indicate the genetic magnitude for variability phenotypes indicated a maximum change of 6–9% in genetic SD of TNB, NBA and LW. The genetic correlations between mean and corresponding variability traits varied from 0.66 to 0.74, whereas the correlation between other mean and variability traits ranged from 0.33 to 0.99. The simulation indicated that even with selection targeted against the variability of reproduction traits, a very limited change should be expected due to a complex genetic and phenotypic relationship between the traits. In the scenarios with selection against LnVarTNB and LnVarLW, this was a decrease of 0.1–0.6% per year, whereas in scenario with selection against LnVarNBA, the range was 0.6–1.1% per year. It is still possible to increase litter size and birthweight further, however, a balance between mean and variability of reproduction traits is required, which can be obtained only by a very well designed breeding programme.     

An AFLP marker tightly linked to apomixis reveals hemizygosity in a portion of the apomixis-controlling locus in Paspalum simplex.

A mapping population of Paspalum simplex segregating for apomixis (asexual reproduction through seeds) was screened with AFLPs to find apomixis-linked markers. Four AFLPs linked to apomixis in coupling phase were found. Three of them did not show recombinants among the 87 individuals of the mapping population, whereas the other was more loosely linked. Integrating the AFLP data with those obtained previously with rice RFLP anchor markers, a map was drawn for the chromosome region of P. simplex encompassing apomixis. We cloned the three AFLPs tightly linked with apomixis into plasmid vectors and used them as probes to hybridize the restriction digested DNA of the mapping population. Two of them revealed RFLP bands linked to apomixis together with other alleles, whereas one was proven to belong to a hemizygous portion of the apomixis locus. The total picture resulting from AFLP and RFLP analyses was that a cluster of markers tightly linked with apomixis was detected in P simplex together with two other markers that were more loosely linked. These two markers enclosed a relatively large chromosome segment characterized by strong repression of recombination. The block of recombination may have caused sequence divergence and, therefore, hemizygosity of some regions belonging to the apomixis-controlling chromosome segment of P. simplex. The potential of developing an apomixis-specific sequence for screening large-fragment libraries for the physical isolation of the locus encompassing apomixis is discussed.     

Apomixis for crop improvement

Summary Apomixis is a genetically controlled reproductive process by which embryos and seeds develop in the ovule without female meiosis and egg cell fertilization. Apomixis produces seed progeny that are exact replicas of the mother plant. The major advantage of apomixis over sexual reproduction is the possibility to select individuals with desirable gene combinations and to propagate them as clones. In contrast to clonal propagation through somatic embryogenesis or in vitro shoot multiplication, apomixis avoids the need for costly processes, such as the production of artificial seeds and tissue culture. It simplifies the processes of commercial hybrid and cultivar production and enables a large-scale seed production economically in both seed- and vegetatively propagated crops. In vegetatively reproduced plants (e.g., potato), the main applications of apomixis are the avoidance of phytosanitary threats and the spanning of unfavorable seasons. Because of its potential for crop improvement and global agricultural production, apomixis is now receiving increasing attention from both scientific and industrial sectors. Harnessing apomixis is a major goal in applied plant genetic engineering. In this regard, efforts are focused on genetic and breeding strategies in various plant species, combined with molecular methods to analyze apomictic and sexual modes of reproduction and to identify key regulatory genes and mechanisms underlying these processes. Also, investigations on the components of apomixis, i.e., apomeiosis, parthenogenesis, and endosperm development without fertilization, genetic screens for apomictic mutants and transgenic approaches to modify sexual reproduction by using various regulatory genes are receiving a major effort. These can open new avenues for the transfer of the apomixis trait to important crop species and will have far-reaching potentials in crop improvement regarding agricultural production and the quality of the products.     

Insight into the genetic bases of climatic adaptation in experimentally evolving wheat populations

Experimental populations evolving under natural selection represent an interesting tool to study genetic bases of adaptation. Evolution of genes possibly involved in adaptive response can be followed together with the corresponding phenotypic traits. Using experimental populations of hexaploid wheat, we studied the evolution of flowering time, a major adaptive trait that synchronizes the initiation of reproduction and the occurrence of favourable environmental conditions. During 12 generations, three populations were grown in contrasted environments (Vervins North France, Le Moulon near Paris, Toulouse South France) under the influence of natural selection, drift, mutation and recombination. Evolution of diversity at the major gene VRN-1 involved in wheat vernalization response has been analysed jointly with earliness estimated in controlled conditions. Whatever the population, rapid phenotypic changes as well as parallel genotypic variations were observed in the first seven generations, probably as the result of selection acting on this major gene which explains 80% of the trait variation overall. Different allelic combinations at physically unlinked copies of VRN-1 located on distinct genomes (A, B and D) were selected between populations. As theoretically expected, due to population differentiation, a high level of genetic diversity was maintained overall in generation 12. Surprisingly, in two populations out of three, the emergence of new alleles by mutation or migration, coupled with temporal variable selection or frequency-dependent selection, allowed to maintain within-population diversity despite local genetic drift and natural selection. This result may plead for an evolutionary approach of wheat genetic resource conservation.