Genetic basis of heterosis for growth-related traits in Arabidopsis investigated by testcross progenies of near-isogenic lines reveals a significant role of epistasis

Epistasis seems to play a significant role in the manifestation of heterosis. However, the power of detecting epistatic interactions among quantitative trait loci (QTL) in segregating populations is low. We studied heterosis in Arabidopsis thaliana hybrid C24 x Col-0 by testing near-isogenic lines (NILs) and their triple testcross (TTC) progenies. Our objectives were to (i) provide the theoretical basis for estimating different types of genetic effects with this experimental design, (ii) determine the extent of heterosis for seven growth-related traits, (iii) map the underlying QTL, and (iv) determine their gene action. Two substitution libraries, each consisting of 28 NILs and covering approximately 61 and 39% of the Arabidopsis genome, were assayed by 110 single-nucleotide polymorphism (SNP) markers. With our novel generation means approach 38 QTL were detected, many of which confirmed heterotic QTL detected previously in the same cross with TTC progenies of recombinant inbred lines. Furthermore, many of the QTL were common for different traits and in common with the 58 QTL detected by a method that compares triplets consisting of a NIL, its recurrent parent, and their F(1) cross. While the latter approach revealed mostly (75%) overdominant QTL, the former approach allowed separation of dominance and epistasis by analyzing all materials simultaneously and yielded substantial positive additive x additive effects besides directional dominance. Positive epistatic effects reduced heterosis for growth-related traits in our materials.     

The role of epistasis in the manifestation of heterosis: a systems-oriented approach

Heterosis is widely used in breeding, but the genetic basis of this biological phenomenon has not been elucidated. We postulate that additive and dominance genetic effects as well as two-locus interactions estimated in classical QTL analyses are not sufficient for quantifying the contributions of QTL to heterosis. A general theoretical framework for determining the contributions of different types of genetic effects to heterosis was developed. Additive x additive epistatic interactions of individual loci with the entire genetic background were identified as a major component of midparent heterosis. On the basis of these findings we defined a new type of heterotic effect denoted as augmented dominance effect di* that comprises the dominance effect at each QTL minus half the sum of additive x additive interactions with all other QTL. We demonstrate that genotypic expectations of QTL effects obtained from analyses with the design III using testcrosses of recombinant inbred lines and composite-interval mapping precisely equal genotypic expectations of midparent heterosis, thus identifying genomic regions relevant for expression of heterosis. The theory for QTL mapping of multiple traits is extended to the simultaneous mapping of newly defined genetic effects to improve the power of QTL detection and distinguish between dominance and overdominance.     

Genetic expectations of quantitative trait loci main and interaction effects obtained with the triple testcross design and their relevance for the analysis of heterosis

Interpretation of experimental results from quantitative trait loci (QTL) mapping studies on the predominant type of gene action can be severely affected by the choice of statistical model, experimental design, and provision of epistasis. In this study, we derive quantitative genetic expectations of (i) QTL effects obtained from one-dimensional genome scans with the triple testcross (TTC) design and (ii) pairwise interactions between marker loci using two-way analyses of variance (ANOVA) under the F(2)- and the F(infinity)-metric model. The theoretical results show that genetic expectations of QTL effects estimated with the TTC design are complex, comprising both main and epistatic effects, and that genetic expectations of two-way marker interactions are not straightforward extensions of effects estimated in one-dimensional scans. We also demonstrate that the TTC design can partially overcome the limitations of the design III in separating QTL main effects and their epistatic interactions in the analysis of heterosis and that dominance x additive epistatic interactions of individual QTL with the genetic background can be estimated with a one-dimensional genome scan. Furthermore, we present genetic expectations of variance components for the analysis of TTC progeny tested in a split-plot design, assuming digenic epistasis and arbitrary linkage.     

A complete solution for dissecting pure main and epistatic effects of QTL in triple testcross design

Epistasis plays an important role in genetics, evolution and crop breeding. To detect the epistasis, triple test cross (TTC) design had been developed several decades ago. Classical procedures for the TTC design use only linear transformations Z(1), Z(2) and Z(3), calculated from the TTC family means of quantitative trait, to infer the nature of the collective additive, dominance and epistatic effects of all the genes. Although several quantitative trait loci (QTL) mapping approaches in the TTC design have been developed, these approaches do not provide a complete solution for dissecting pure main and epistatic effects. In this study, therefore, we developed a two-step approach to estimate all pure main and epistatic effects in the F(2)-based TTC design under the F(2) and F(∞) metric models. In the first step, with Z(1) and Z(2) the augmented main and epistatic effects in the full genetic model that simultaneously considered all putative QTL on the whole genome were estimated using empirical Bayes approach, and with Z(3) three pure epistatic effects were obtained using two-dimensional genome scans. In the second step, the three pure epistatic effects obtained in the first step were integrated with the augmented epistatic and main effects for the further estimation of all other pure effects. A series of Monte Carlo simulation experiments has been carried out to confirm the proposed method. The results from simulation experiments show that: 1) the newly defined genetic parameters could be rightly identified with satisfactory statistical power and precision; 2) the F(2)-based TTC design was superior to the F(2) and F(2:3) designs; 3) with Z(1) and Z(2) the statistical powers for the detection of augmented epistatic effects were substantively affected by the signs of pure epistatic effects; and 4) with Z(3) the estimation of pure epistatic effects required large sample size and family replication number. The extension of the proposed method in this study to other base populations was further discussed.     

Quantitative trait loci mapping and the genetic basis of heterosis in maize and rice

Despite its importance to agriculture, the genetic basis of heterosis is still not well understood. The main competing hypotheses include dominance, overdominance, and epistasis. NC design III is an experimental design that has been used for estimating the average degree of dominance of quantitative trait loci (QTL) and also for studying heterosis. In this study, we first develop a multiple-interval mapping (MIM) model for design III that provides a platform to estimate the number, genomic positions, augmented additive and dominance effects, and epistatic interactions of QTL. The model can be used for parents with any generation of selfing. We apply the method to two data sets, one for maize and one for rice. Our results show that heterosis in maize is mainly due to dominant gene action, although overdominance of individual QTL could not completely be ruled out due to the mapping resolution and limitations of NC design III. For rice, the estimated QTL dominant effects could not explain the observed heterosis. There is evidence that additive × additive epistatic effects of QTL could be the main cause for the heterosis in rice. The difference in the genetic basis of heterosis seems to be related to open or self pollination of the two species. The MIM model for NC design III is implemented in Windows QTL Cartographer, a freely distributed software.     

Gene actions at loci underlying several quantitative traits in two elite rice hybrids

To understand the gene activities controlling nine important agronomic quantitative traits in rice, we applied a North Carolina design 3 (NC III design) analysis to recombinant inbred lines (RILs) in highly heterotic inter- (IJ) and intra-subspecific (II) hybrids by performing the following tasks: (1) investigating the relative contribution of additive, dominant, and epistatic effects for performance traits by generation means analysis and variance component estimates; (2) detecting the number, genomic positions, and genetic effects of QTL for phenotypic traits; and (3) characterizing their mode of gene action. Under an F∞-metric, generation means analysis and variance components estimates revealed that epistatic effects prevailed for the majority of traits in the two hybrids. QTL analysis identified 48 and 66 main-effect QTL (M-QTL) for nine traits in IJ and II hybrids, respectively. In IJ hybrids, 20 QTL (41.7%) showed an additive effect of gene actions, 20 (41.7%) showed partial-to-complete dominance, and 8 (16.7%) showed overdominance. In II hybrids, 34 QTL (51.5%) exhibited additive effects, 14 (21.2%) partial-to-complete dominance, and 18 (27.3%) overdominance. There were 153 digenic interactions (E-QTL) in the IJ hybrid and 252 in the II hybrid. These results suggest that additive effects, dominance, overdominance, and particularly epistasis attribute to the genetic basis of the expression of traits in the two hybrids. Additionally, we determined that the genetic causes of phenotypic traits and their heterosis are different. In the plants we studied, the phenotypic traits investigated and their heterosis were conditioned by different M-QTL and E-QTL, respectively, and were mainly due to non-allelic interactions (epistasis).     

A dynamic and complex network regulates the heterosis of yield-correlated traits in rapeseed (Brassica napus L.)

Although much research has been conducted, the genetic architecture of heterosis remains ambiguous. To unravel the genetic architecture of heterosis, a reconstructed F(2) population was produced by random intercross among 202 lines of a double haploid population in rapeseed (Brassica napus L.). Both populations were planted in three environments and 15 yield-correlated traits were measured, and only seed yield and eight yield-correlated traits showed significant mid-parent heterosis, with the mean ranging from 8.7% (branch number) to 31.4% (seed yield). Hundreds of QTL and epistatic interactions were identified for the 15 yield-correlated traits, involving numerous variable loci with moderate effect, genome-wide distribution and obvious hotspots. All kinds of mode-of-inheritance of QTL (additive, A; partial-dominant, PD; full-dominant, D; over-dominant, OD) and epistatic interactions (additive × additive, AA; additive × dominant/dominant × additive, AD/DA; dominant × dominant, DD) were observed and epistasis, especially AA epistasis, seemed to be the major genetic basis of heterosis in rapeseed. Consistent with the low correlation between marker heterozygosity and mid-parent heterosis/hybrid performance, a considerable proportion of dominant and DD epistatic effects were negative, indicating heterozygosity was not always advantageous for heterosis/hybrid performance. The implications of our results on evolution and crop breeding are discussed.     

Identification of heterotic metabolite QTL in Arabidopsis thaliana RIL and IL populations

Two mapping populations of a cross between the Arabidopsis thaliana accessions Col-0 and C24 were cultivated and analyzed with respect to the levels of 181 metabolites to elucidate the biological phenomenon of heterosis at the metabolic level. The relative mid-parent heterosis in the F₁ hybrids was <20% for most metabolic traits. The first mapping population consisting of 369 recombinant inbred lines (RILs) and their test cross progeny with both parents allowed us to determine the position and effect of 147 quantitative trait loci (QTL) for metabolite absolute mid-parent heterosis (aMPH). Furthermore, we identified 153 and 83 QTL for augmented additive (Z₁) and dominance effects (Z₂), respectively. We identified putative candidate genes for these QTL using the aracyc database ( http://www.arabidopsis.org/biocyc ), and calculated the average degree of dominance, which was within the dominance and over-dominance range for most metabolites. Analyzing a second population of 41 introgression lines (ILs) and their test crosses with the recurrent parent, we identified 634 significant differences in metabolite levels. Nine per cent of these effects were classified as over-dominant, according to the mode of inheritance. A comparison of both approaches suggested epistasis as a major contributor to metabolite heterosis in Arabidopsis. A linear combination of metabolite levels was shown to significantly correlate with biomass heterosis ( r  = 0.62).     

Genetics of quantitative traits in Arabidopsis thaliana

The genetic control of 22 quantitative traits, including developmental rates and sizes, was examined in generations of Arabidopsis thaliana derived from the cross between the ecotypes, Columbia (Col) and Landsberg erecta (Ler). The data were obtained from three sets of families raised in the same trial: the 16 basic generations, that is, parents, F(1)'s, F(2)'s, backcrosses, recombinant inbred lines (RILs) and a triple test cross (TTC), the latter produced by crossing the RILs to Col, Ler and their F(1). The data were analysed by two approaches. The first (approach A) involved traditional generation mean and variance component analysis and the second (B), based around the RILs and TTC families, involved marker-based QTL analysis.From (A), genetic differences between Col and Ler were detected for all traits with moderate heritabilities. Height at flowering was the only trait to show heterosis. Dominance was partial to complete for all height traits, and there was no overdominance but there was strong evidence for directional dominance. For most other traits, dominance was ambidirectional and incomplete, with average dominance ratios of around 80%. Epistasis, particularly of the duplicate type that opposes dominance, was a common feature of all traits. The presence of epistasis must imply multiple QTL for all traits.The QTL analysis located 38 significant effects in four regions of chromosomes I, II, IV and V, but not III. QTL affecting rosette size and leaf number were identified in all four regions, with days to maturity on chromosomes IV and V. The only QTL for height was located at the expected position of the erecta gene (chromosome II; 50 cM), but the additive and dominance effects of this single QTL did not adequately explain the generation means. The possible involvement of other interacting height QTL is discussed.     

Population-based resequencing analysis of improved wheat germplasm at wheat leaf rust resistance locus Lr21

Little is known about the genetic control of heterosis in the complex polyploid crop species oilseed rape (Brassica napus L.). In this study, two large doubled-haploid (DH) mapping populations and two corresponding sets of backcrossed test hybrids (THs) were analysed in controlled greenhouse experiments and extensive field trials for seedling biomass and yield performance traits, respectively. Genetic maps from the two populations, aligned with the help of common simple sequence repeat markers, were used to localise and compare quantitative trait loci (QTL) related to the expression of heterosis for seedling developmental traits, plant height at flowering, thousand seed mass, seeds per silique, siliques per unit area and seed yield. QTL were mapped using data from the respective DH populations, their corresponding TH populations and from mid-parent heterosis (MPH) data, allowing additive and dominance effects along with digenic epistatic interactions to be estimated. A number of genome regions containing numerous heterosis-related QTL involved in different traits and at different developmental stages were identified at corresponding map positions in the two populations. The co-localisation of per se QTL from the DH population datasets with heterosis-related QTL from the MPH data could indicate regulatory loci that may also contribute to fixed heterosis in the highly duplicated B. napus genome. Given the key role of epistatic interactions in the expression of heterosis in oilseed rape, these QTL hotspots might harbour genes involved in regulation of heterosis (including fixed heterosis) for different traits throughout the plant life cycle, including a significant overall influence on heterosis for seed yield.     

Epistasis for fitness-related quantitative traits in Arabidopsis thaliana grown in the field and in the greenhouse

The extent to which epistasis contributes to adaptation, population differentiation, and speciation is a long-standing and important problem in evolutionary genetics. Using recombinant inbred (RI) lines of Arabidopsis thaliana grown under natural field conditions, we have examined the genetic architecture of fitness-correlated traits with respect to epistasis; we identified both single-locus additive and two-locus epistatic QTL for natural variation in fruit number, germination, and seed length and width. For fruit number, we found seven significant epistatic interactions, but only two additive QTL. For seed germination, length, and width, there were from two to four additive QTL and from five to eight epistatic interactions. The epistatic interactions were both positive and negative. In each case, the magnitude of the epistatic effects was roughly double that of the effects of the additive QTL, varying from -41% to +29% for fruit number and from -5% to +4% for seed germination, length, and width. A number of the QTL that we describe participate in more than one epistatic interaction, and some loci identified as additive also may participate in an epistatic interaction; the genetic architecture for fitness traits may be a network of additive and epistatic effects. We compared the map positions of the additive and epistatic QTL for germination, seed width, and seed length from plants grown in both the field and the greenhouse. While the total number of significant additive and epistatic QTL was similar under the two growth conditions, the map locations were largely different. We found a small number of significant epistatic QTL x environment effects when we tested directly for them. Our results support the idea that epistatic interactions are an important part of natural genetic variation and reinforce the need for caution in comparing results from greenhouse-grown and field-grown plants.     

Estimates of genetic variance in an F2 maize population

Maize (Zea mays L.) breeders have used several genetic-statistical models to study the inheritance of quantitative traits. These models provide information on the importance of additive, dominance, and epistatic genetic variance for a quantitative trait. Estimates of genetic variances are useful in understanding heterosis and determining the response to selection. The objectives of this study were to estimate additive and dominance genetic variances and the average level of dominance for an F2 population derived from the B73 x Mo17 hybrid and use weighted least squares to determine the importance of digenic epistatic variances relative to additive and dominance variances. Genetic variances were estimated using Design III and weighted least squares analyses. Both analyses determined that dominance variance was more important than additive variance for grain yield. For other traits, additive genetic variance was more important than dominance variance. The average level of dominance suggests either overdominant gene effects were present for grain yield or pseudo-overdominance because of linkage disequilibrium in the F2 population. Epistatic variances generally were not significantly different from zero and therefore were relatively less important than additive and dominance variances. For several traits estimates of additive by additive epistatic variance decreased estimates of additive genetic variance, but generally the decrease in additive genetic variance was not significant.     

The genetic basis of heterosis: multiparental quantitative trait loci mapping reveals contrasted levels of apparent overdominance among traits of agronomical interest in maize (Zea mays L.)

Understanding the genetic bases underlying heterosis is a major issue in maize (Zea mays L.). We extended the North Carolina design III (NCIII) by using three populations of recombinant inbred lines derived from three parental lines belonging to different heterotic pools, crossed with each parental line to obtain nine families of hybrids. A total of 1253 hybrids were evaluated for grain moisture, silking date, plant height, and grain yield. Quantitative trait loci (QTL) mapping was carried out on the six families obtained from crosses to parental lines following the "classical" NCIII method and with a multiparental connected model on the global design, adding the three families obtained from crosses to the nonparental line. Results of the QTL detection highlighted that most of the QTL detected for grain yield displayed apparent overdominance effects and limited differences between heterozygous genotypes, whereas for grain moisture predominance of additive effects was observed. For plant height and silking date results were intermediate. Except for grain yield, most of the QTL identified showed significant additive-by-additive epistatic interactions. High correlation observed between heterosis and the heterozygosity of hybrids at markers confirms the complex genetic basis and the role of dominance in heterosis. An important proportion of QTL detected were located close to the centromeres. We hypothesized that the lower recombination in these regions favors the detection of (i) linked QTL in repulsion phase, leading to apparent overdominance for heterotic traits and (ii) linked QTL in coupling phase, reinforcing apparent additive effects of linked QTL for the other traits.     

QTL and epistatic analyses of heterosis for seed yield and three yield component traits using molecular markers in rapeseed (Brassica napus L.)

Aiming to explore the basis of heterosis in rapeseed, QTLs for yield and three yield component traits were mapped and the digenic interactions were detected in an F₂ population derived from a cross between two elite rapeseed lines, SI-1300 and Eagle, in this study. Twenty-eight QTLs were detected for the four yield traits, with only two of them detected simultaneously in the Wuhan and Jingmen environments. Additive, partial dominance, dominance, and overdominance effects were all identified for the investigated traits. Dominance (including partial dominance) was shown by 55% of the QTLs, which suggests that dominance is a major genetic basis of heterosis in rapeseed. At the P ?? 0.01 level with 1000 random permutations, 108 and 104 significant digenic interactions were detected in Wuhan and Jingmen, respectively, for the four yield-related traits using all possible locus pairs of molecular markers. Digenic interactions, including additive by additive, additive by dominance, and dominance by dominance, were frequent and widespread in this population. In most cases (78.3%), the interactions occurred among marker loci for which significant effects were not detected by single-locus analysis. Some QTLs (57.1%) detected by single-locus analysis were involved in epistatic interactions. It was concluded that epistasis, along with dominance (including partial dominance), is responsible for the expression of heterosis in rapeseed.     

Unraveling epistasis with triple testcross progenies of near-isogenic lines

Libraries of near-isogenic lines (NILs) are a powerful plant genetic resource to map quantitative trait loci (QTL). Nevertheless, QTL mapping with NILs is mostly restricted to genetic main effects. Here we propose a two-step procedure to map additive-by-additive digenic epistasis with NILs. In the first step, a generation means analysis of parents, their F₁ hybrid, and one-segment NILs and their triple testcross (TTC) progenies is used to identify in a one-dimensional scan loci exhibiting QTL-by-background interactions. In a second step, one-segment NILs with significant additive-by-additive background interactions are used to produce particular two-segment NILs to test for digenic epistatic interactions between these segments. We evaluated our approach by analyzing a random subset of a genomewide Arabidopsis thaliana NIL library for growth-related traits. The results of our experimental study illustrated the potential of the presented two-step procedure to map additive-by-additive digenic epistasis with NILs. Furthermore, our findings suggested that additive main effects as well as additive-by-additive digenic epistasis strongly influence the genetic architecture underlying growth-related traits of A. thaliana.     

Something Old and Something New: Wedding Recombinant Inbred Lines with Traditional Line Cross Analysis Increases Power to Describe Gene Interactions

In this paper we present a novel approach to quantifying genetic architecture that combines recombinant inbred lines (RIL) with line cross analysis (LCA). LCA is a method of quantifying directional genetic effects (i.e. summed effects of all loci) that differentiate two parental lines. Directional genetic effects are thought to be critical components of genetic architecture for the long term response to selection and as a cause of inbreeding depression. LCA typically begins with two inbred parental lines that are crossed to produce several generations such as F1, F2, and backcrosses to each parent. When a RIL population (founded from the same P1 and P2 as was used to found the line cross population) is added to the LCA, the sampling variance of several nonadditive genetic effect estimates is greatly reduced. Specifically, estimates of directional dominance, additive x additive, and dominance x dominance epistatic effects are reduced by 92%, 94%, and 56% respectively. The RIL population can be simultaneously used for QTL identification, thus uncovering the effects of specific loci or genomic regions as elements of genetic architecture. LCA and QTL mapping with RIL provide two qualitatively different measures of genetic architecture with the potential to overcome weaknesses of each approach alone. This approach provides cross-validation of the estimates of additive and additive x additive effects, much smaller confidence intervals on dominance, additive x additive and dominance x dominance estimates, qualitatively different measures of genetic architecture, and the potential when used together to balance the weaknesses of LCA or RIL QTL analyses when used alone.     

家蚕茧质性状的QTL定位研究

采用QTLMapper 2．0 QTL作图软件,对F2群体的家蚕全茧量、茧层量、茧层率和蛹体重等性状进行了QTL定位分析,分别检测出7个、6个、2个、8个有显著效应分量的QTLs,分布于7个、5个、2个、7个不同的连锁群。控制全茧量、茧层量的QTLs一般存在复杂的上位性效应。对全茧量性状,有3对QTLs存在显著的加加上位性效应,其中1对还存在加显、显显互作;共有3个QTLs存在显著的显性效应,1个存在显著的加性效应。对茧层量QTLs,发现1对QTLs存在极显著的各项遗传效应,包括上位性效应;1对QTLs被检测到显著的显显互作,1个QTL具有显著的显性效应,并与另一个QTL存在显著的加加互作。茧层率、蛹体重主要受加性或显性的QTLs作用,没有发现茧层率QTLs的上位性效应,蛹体重的有效QTL大都呈现显著的负向显性效应,只有一对QTLs存在显著的加加上位性效应。第2、3、4、11、13、24、34、37、40连锁群是两个或多个性状QTLs分布的共同连锁群。全茧量和茧层量存在共同的QTL或染色体区域,育种上可通过适当选配,利用基因的互作效应,同步改良这两个性状。     

Quantitative trait loci affecting delta13C and response to differential water availibility in Arabidopsis thaliana

Phenotypic plasticity is an important response mechanism of plants to environmental heterogeneity. Here, we explored the genetic basis of plastic responses of Arabidopsis thaliana to water deficit by experimentally mapping quantitative trait loci (QTL) in two recombinant inbred populations (Cvi x Ler and Ler x Col). We detected genetic variation and significant genotype-by-environment interactions for many traits related to water use. We also mapped 26 QTL, including six for carbon isotope composition (delta13C). Negative genetic correlations between fruit length and fruit production as well as between flowering time and branch production were corroborated by QTL colocalization, suggesting these correlations are due to pleiotropy or physical linkage. Water-limited plants were more apically dominant with greater root:shoot ratios and higher delta13C (higher water-use efficiency) when compared to well-watered plants. Many of the QTL effects for these traits interacted significantly with the irrigation treatment, suggesting that the observed phenotypic plasticity is genetically based. We specifically searched for epistatic (QTL-QTL) interactions using a two-dimensional genome scan, which allowed us to detect epistasis regardless of additive genetic effects. We found several significant QTL-QTL interactions including three that exhibited environmental dependence. These results provide preliminary evidence for proposed genetic mechanisms underlying phenotypic plasticity.     

Epistasis plays an important role as genetic basis of heterosis in rice

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