Polymix breeding with paternity analysis in Populus: a test for differential reproductive success (DRS) among pollen donors

Polymix breeding with paternity analysis (PMX/WPA) has been proposed as an alternative to traditional full-sib breeding and testing schemes. To fully capture the benefits of PMX/WPA, differential reproductive success (DRS) of pollen parents used in the polymix must be modest. DRS was evaluated in an operational test of PMX/WPA for a hybrid poplar breeding program. A 16-parent pollen polymix (Populus nigra L.) was used to pollinate seven clones of Populus deltoides (Bartr. ex. Marshall) under greenhouse breeding conditions. Progeny were grown out briefly and randomly sampled (357) prior to out-planting in field trials. Twenty-eight simple sequence repeat (SSR) loci were evaluated and 15 were selected for genetic characterization in small populations of three Populus spp (P. nigra, P. deltoides, and P. balsamifera spp trichocarpa Torr. & Gray). Seven loci were ultimately selected for paternity analysis of progeny. The average exclusion probability of the seven loci in P. nigra was 0.604; combined, the theoretical exclusion probability was 0.9999. However, only 95% of sampled progeny were unambiguously assigned a single paternal parent. Missing data likely accounted for most of the ambiguity. DRS was statistically significant though not prohibitive for practical utility of PMX/WPA as a breeding system. Of the 112 potential crosses in this study, 92 were represented. Eight of the 16 pollen parents contributed 83% of the progeny. Good pollen vigor, as measured by germination percent, did not ensure paternal success, but poor vigor was associated with lack of paternal success. PMX/WPA appears to be logistically and economically attractive for hybrid poplar breeding and testing.     

Impact of reconstructed pedigrees on progeny-test breeding values in red spruce

Pedigrees reconstructed through DNA marker assigned paternities in polymix (PMX) and open pollinated (OP) progeny tests were analyzed using mixed models to test the effect of unequal male reproductive success and pedigree errors on quantitative genetic parameters. The reconstructed pedigree increased heritabilities in the larger PMX test. Increased heritability resulted from adding the paternities to the pedigree per se, not by correcting the male reproductive bias by specifying the exact pedigree. Removing hypothesized pedigree errors had no effect on quantitative parameters, either because the magnitude of the errors was too small (PMX) or the progeny test was too small to detect variance components reliably (OP). Although there was no advantage in backwards selection, the increased additive variance, heritabilities and accuracy of progeny with assigned paternities in the pedigree, should permit forward selection of offspring with greater genetic gain and complete control of coancestry for future breeding decisions. Some possible breeding population structures with the new genetic information are discussed.     

Impact of reconstructed pedigrees on progeny-test breeding values in red spruce

Pedigrees reconstructed through DNA marker assigned paternities in polymix (PMX) and open pollinated (OP) progeny tests were analyzed using mixed models to test the effect of unequal male reproductive success and pedigree errors on quantitative genetic parameters. The reconstructed pedigree increased heritabilities in the larger PMX test. Increased heritability resulted from adding the paternities to the pedigree per se, not by correcting the male reproductive bias by specifying the exact pedigree. Removing hypothesized pedigree errors had no effect on quantitative parameters, either because the magnitude of the errors was too small (PMX) or the progeny test was too small to detect variance components reliably (OP). Although there was no advantage in backwards selection, the increased additive variance, heritabilities and accuracy of progeny with assigned paternities in the pedigree, should permit forward selection of offspring with greater genetic gain and complete control of coancestry for future breeding decisions. Some possible breeding population structures with the new genetic information are discussed.     

Testing for linkage disequilibrium in the New Zealand radiata pine breeding population

Linkage analysis is commonly used to find marker-trait associations within the full-sib families of forest tree and other species. Study of marker-trait associations at the population level is termed linkage-disequilibrium (LD) mapping. A female-tester design comprising 200 full-sib families generated by crossing 40 pollen parents with five female parents was used to assess the relationship between the marker-allele frequency classes obtained from parental genotypes at SSR marker loci and the full-sib family performance (average predicted breeding value of two parents) in radiata pine (Pinus radiata D. Don). For alleles (at a marker locus) that showed significant association, the copy number of that allele in the parents was significantly correlated, either positively or negatively, with the full-sib family performance for various economic traits. Regression of parental breeding value on its genotype at marker loci revealed that most of the markers that showed significant association with full-sib family performance were not significantly associated with the parental breeding values. This suggests that over-representation of the female parents in our sample of 200 full-sib families could have biased the process of detecting marker-trait associations. The evidence for the existence of marker-trait LD in the population studied is rather weak and would require further testing. The exact test for genotypic disequilibrium between pairs of linked or unlinked marker loci revealed non-significant LD. Observed genotypic frequencies at several marker loci were significantly different from the expected Hardy-Weinberg equilibrium. The possibilities of utilising marker-trait associations for early selection, among-family selection and selecting parents for the next generation of breeding are also discussed.     

Spatial variation in local pollen flow and mating success in a Picea abies clone archive and their implications for a novel ��breeding without breeding�� strategy

Currently, Norway spruce (Picea abies) breeding in Sweden is based on crosses between the best clones followed by clonal testing of the progenies to select for the long-term breeding population. An alternative breeding strategy called “Breeding without Breeding” (BwB) is proposed, which, in principle, relies on the DNA marker-based pedigree reconstruction from wind-pollinated progenies instead of controlled crosses. To test whether the pedigree structure could be established from progenies of clonal trials, we investigated the spatial pattern of local pollen flow and paternity assignment in a clone archive of Norway spruce. The results showed that 42% of the progeny can be assigned to fathers within 30-m distance with high confidence. Effective pollen dispersal decreased rapidly with distance and followed exponential distribution on local scale. The extent of close-neighbor (within 6 m) mating ranged from 0% to 48% among grafts with an average of 13%. Distance explained 25% deviance in mating success, and other factors such as phenology and spatial configuration of the clones should have contributed the rest. The success of parentage assignment in clone archive opens up the possibility to apply BwB in clonal trials of species that are easy to propagate vegetatively. This procedure could substantially shorten the breeding cycle and still give similar gain per year as the conventional breeding.     

Paternity analysis in a progeny test of Cryptomeria japonica revealed adverse effects of pollen contamination from outside seed orchards on morphological traits

To evaluate the effects of pollen contamination from outside of Cryptomeria japonica seed orchard on the growth performance (height and diameter at breast height, DBH) and morphological traits (stem straightness and basal stem straightness), paternity testing using seven microsatellite markers was performed in a progeny test. In the studied progeny test, high rates of inconsistency were found between the observed and expected genotypes. The average rates of pollen contamination from outside the orchard and self-fertilization were 58.47% and 0.65%, respectively. We divided the individuals of the studied progeny test into two groups based on their genotype data, for which: (1) both parents were elite trees and (2) only the mother trees were elite trees, and then compared them with respect to the growth performance and morphological traits of progenies using data at 20 and 30 years old. Significant adverse effects of contaminating pollen were detected in relation to straightness, but not tree height and DBH. The results suggest that the genetic gains for straightness generally show higher narrow-sense heritability than growth traits, which should be increased by reduction of pollen contamination. Breeding with paternal analysis (BWPA) is an effective approach for evaluating breeding materials based on maternal and paternal information revealed by DNA markers. The use of BWPA in progeny test allows effective forward- and backward selection without laborious and time-consuming tasks. In this study, we also suggest that the significant pollen contamination and paternal deviation found in the open-pollinated progeny test are serious impediments for BWPA.     

Comparison of ancestral and current-generation inbreeding in an experimental strawberry breeding population

Progenies from first-generation self, half-sib, full-sib, and cross fertilizations were generated to evaluate the magnitude of inbreeding depression for vegetative and production traits in strawberry. Tests were conducted to determine the linearity of trait mean depression with inbreeding rate (F) over this range of inbreeding values, as an indication of the presence of non-additive epistasis. A control population, for which a similar range of coancestry had accumulated over several cycles of breeding and selection, was also generated to compare the consequences of ancestral and current-generation inbreeding. Trait means for crosses among current-generation half-sibs, full-sibs, and selfs were 2–17%, 3–12%, and 14–45% lower than for unrelated crosses among the same set of parents, respectively. Linear regression of progeny means on current generation F was significantly negative for all traits and explained 17–44% of the variance among progeny means. Mean depression was largely linear over the range of inbreeding rates tested in this population, indicating the absence of epistasis for the traits evaluated. Conversely, (F) regressions of progeny means on pedigree inbreeding coefficients, where coancestry had accumulated over several cycles of breeding and selection, were uniformly non-significant and explained 0–10% of the variance among cross means. Further, multiple regression of progeny means for current-generation relatives on pedigree F failed to improve fit significantly over regression on current-generation F alone for all traits. Together, these results suggest that pedigree inbreeding coefficients are poor predictors of changes in homozygosity when populations are developed through multiple cycles of breeding and selection. They also imply that inbreeding depression will be of minor importance for strawberry breeding populations managed with adequate population sizes and strong directional selection.     

Group coancestry-controlled selection in a Pinus sylvestris L. breeding program

 Integer Linear Programming was used to maximize genetic gain from selection at a given level of relatedness. Variances and breeding values for total height were available for 296 plus-trees of Pinus sylvestris which had been evaluated by open-pollinated progeny testing at a single test site in northern Sweden. Second-generation breeding and selection scenarios for this breeding population were evaluated using simulated data derived deterministically from normal distributions of estimated breeding values of progeny around mid-parent family means. The study considered two mating designs, assortative and non-assortative single-pair mating, and two selection criteria, individual phenotype and performance of half-sib progeny. Relatedness (group coancestry) was restricted to a level equivalent to that given by within-family selection of 2 trees per family from each of 25 families (the current standard in Sweden). Selection that allows the best-performing families to contribute a greater number of progeny was superior, both when the breeding population size was limited to 50 individuals and when it was allowed to be larger. The selected set giving the greatest average breeding value under restricted group coancestry included the best individual from families that would have been rejected under application of standard within-family selection. We made a comparison of the present value on retrieved gain between phenotypic selection and evaluation by progeny testing. Received: 24 November 1998 / Accepted: 14 December 1998     

Chloroplast DNA inheritance in the orchid Anacamptis palustris using single-seed polymerase chain reaction

The modality of chloroplast inheritance in orchids has been investigated only in a few species due to the difficulties associated with the analysis of large progeny numbers from experimental crosses. To test chloroplast DNA inheritance in the orchid Anacamptis palustris, we took advantage of the presence of a highly variable minisatellite repeat located in the tRNA(LEU) intron in the chloroplast genome. Seed progeny obtained from experimental crosses between parental individuals carrying different chloroplast DNA (cpDNA) minisatellite repeat numbers were analyzed using a single-seed polymerase chain reaction (PCR) protocol. All examined seeds displayed the maternal cpDNA haplotypes, indicating that cpDNA inheritance is strictly maternal in this Mediterranean orchid species. No evidence for paternal leakage was found. This finding concurs with results obtained from PCR amplifications of pollen massulae that exclude the presence of chloroplast DNA in the pollen tetrads.     

CROSS ABILITY OF MIMULUS GUTTATUS IN RELATION TO COMPONENTS OF GENE FIXATION

The joint effects of parental gene fixation and consanguinity of mates upon the fitness of matings was examined in Mimulus guttatus. Plants from four populations were crossed at five levels of genetic relatedness, and five viability characters were scored in progeny. Parental gene fixation at 12 polymorphic allozyme loci was partitioned into local, subpopulation, and population components. A model is proposed wherein parental gene fixation influences distance-dependent crossing success. At a fixed locus, inbreeding is favored if natural selection caused allele fixation, or is disfavored if gene fixation was random. The distance between mates required to eliminate gene fixation depends upon patch size of fixation. When selective fixation and patch size differ among loci, an optimal crossing distance is possible. In M. guttatus, progeny viability generally decreased with greater relatedness between mates, but this decrease was often heterogeneous among populations. The highest pollen viability and the lowest seed set were found at an intermediate relatedness between mates. To determine whether parental gene fixation influences these crossing patterns, a type of mutational-load analysis was performed. Progeny fitness was regressed on parent F and fitness estimated at F = 1. This was done for each component of F, for a) crosses that maintain gene fixation and b) crosses that eliminate gene fixation. A multiplicative, composite measure of fitness indicates that, in M. guttatus, genes fixed during local or population differentiation favor outbreeding, while genes fixed during subpopulation differentiation favor inbreeding. This predicts that random mating within subpopulations confers highest progeny fitness, exclusive of between-population matings. However, predictions did not fit the observed patterns of crossing success very well, perhaps because gene fixation was relatively low or was not adequately measured at loci influencing fitness.     

Evidence for inbreeding depression and post-pollination selection against inbreeding in the dioecious plant Silene latifolia

In many species, inbred individuals have reduced fitness. In plants with limited pollen and seed dispersal, post-pollination selection may reduce biparental inbreeding, but knowledge on the prevalence and importance of pollen competition or post-pollination selection after non-self pollination is scarce. We tested whether post-pollination selection favours less related pollen donors and reduces inbreeding in the dioecious plant Silene latifolia. We crossed 20 plants with pollen from a sibling and an unrelated male, and with a mix of both. We found significant inbreeding depression on vegetative growth, age at first flowering and total fitness (22% in males and 14% in females). In mixed pollinations, the unrelated male sired on average 57% of the offspring. The greater the paternity share of the unrelated sire, the larger the difference in relatedness of the two males to the female. The effect of genetic similarity on paternity is consistent with predictions for post-pollination selection, although paternity, at least in some crosses, may be affected by additional factors. Our data show that in plant systems with inbreeding depression, such as S. latifolia, pollen or embryo selection after multiple-donor pollination may indeed reduce inbreeding.     

FITNESS CONSEQUENCES OF MIXED-DONOR POLLEN LOADS IN THE ANNUAL LEGUME CHAMAECRISTA FASCICULATA

We experimentally examined the effects of pollen composition on progeny fitness in the self-compatible, annual plant Chamaecrista fasciculata. Plants were hand-pollinated with single- and mixed-donor pollen loads and with various combinations of self- and outcross pollen. For outcrosses, pollen was obtained from two plants at each of two different distances within the same subpopulation as the female parent. Seedlings from all crosses were planted back into the maternal site. For single-donor crosses, seed weight, progeny fruit production, and overall relative fitness were significantly higher for outcross, as compared to self-treatments, but we found no significant differences among outcross sources. For all fitness components, the value observed for crosses derived from mixed loads was intermediate between the values for the singledonor crosses that comprised the mixed load. In a parallel experiment, an analysis of seed paternity of progeny which resulted from pollen mixtures of self- and outcross pollen showed random paternity in two maternal families, and significant excess of outcross in one family. Our results demonstrate that mixed pollen loads do not confer a fitness advantage to the maternal plant in this species, and that the fitness observed for progeny derived from mixed loads is generally consistent with a hypothesis of random paternity.     

Inheritance of self-compatibility in almond: breeding strategies to assure self-compatibility in the progeny

To assure self-compatibility in the progenies, three different crosses were conducted for the first time in an almond breeding programme: self-pollination (266 descendants from 30 families), crosses between parents sharing an S-allele (108 descendants from five families) and crosses with homozygous self-compatible parents (62 descendants from five families). Depending on the cross, self-compatibility in the progenies was determined by observing pollen tube growth (by means of fluorescence microscopy), stylar S-RNases analysis or allele-specific PCR. The results obtained fit with the accepted hypothesis of inheritance of self-compatibility and the three crossing strategies used ensured 100% of self-compatible descendants. These strategies increase the efficiency of the breeding programme and avoid the laborious task of evaluating this characteristic. From the breeding point of view, self-fertilisation and crosses between relatives tend to produce inbreeding. Furthermore, these methods reduce the possibilities of choosing the parental combination. The use of homozygous self-compatible parents does not have any of these disadvantages. As far as we know, this is the first time that allele-specific PCR has been used for early selection of self-compatible seedlings. The advantages and disadvantages of the three methodologies used to determine self-compatibility are discussed.     

Use of mid-parent values and progeny tests to increase the efficiency of potato breeding for combined processing quality and disease and pest resistance

A potato breeding strategy is presented which avoids the common but ineffective practice of intense early-generation visual selection between seedlings in a glasshouse and spaced plants at a seed site. Once pair crosses have been made, progeny tests are used to discard whole progenies before starting conventional within-progeny selection at the unreplicated small-plot stage. Clones are also visually selected from the best progenies for use as parents in the next cycle of crosses whilst they are multiplied to provide enough tubers for assessment of their yield and quality. Mid-parent values, as well as progeny tests, are then used to select between the resultant crosses. Material from other breeding programmes can be included in the parental assessments and used in the next cycle of crosses if superior. Finally, in seeking new cultivars, the number of clones on which to practise selection is increased by sowing more true seed of the best progenies, but without selection until the small-plot stage. Traits considered are resistance to late blight [ Phytophthora infestans (Mont.) de Bary] and to the white potato cyst nematode [ Globodera pallida (Stone)], fry colour and tuber yield and appearance, as visually assessed by breeders. The theoretical superiority of the strategy for seeking new cultivars lies in being able to practise between-cross selection for a number of economically important traits within 1 or 2 years of making crosses, something that is not possible on individuals as seedlings in the glasshouse or spaced plants at the seed site. This also means that full-sib family selection can be operated on a 3-year cycle, an improvement on current practice of clonal selection on what is often at least a nine-year cycle. New cultivars can be sought with more confidence from the best progenies in each cycle, and modern methods of rapid multiplication used to reduce the number of clonal generations required to find the best clones.     

Unravelling first-generation pedigrees in wild endangered salmon populations using molecular genetic markers

There is increasing interest in the use of molecular genetic data to infer genealogical relationships among individuals in the absence of parental information. Such analyses can provide insight into mating systems and estimations of heritability in the wild. In addition, accurate pedigree reconstruction among the founders of endangered populations being reared in captivity would be invaluable. Many breeding programs for endangered species attempt to minimize loss of genetic variation and inbreeding through strategies designed to minimize global co-ancestry, but they assume a lack of relatedness among the founders. Yet populations that are the target of such programs are generally in serious demographic decline, and many of the available founders may be closely related. Here we demonstrate determination of full and half-sib relationships among the wild founders of a captive breeding program involving two endangered Atlantic salmon populations using two different approaches and associated software, pedigree and colony. A large portion of the juveniles collected in these two rivers appear to be derived from surprisingly few females mating with a large number of males, probably small precocious parr. Another group of potential founders, obtained from a local hatchery, clearly originated from a small number of full-sib crosses. These results allowed us to prioritize individuals on the basis of conservation value, and are expected to help minimize loss of genetic variation through time. In addition, insight is provided into the number of contributing parents and the mating systems that produced this last generation of endangered wild Atlantic salmon.     

PATERNAL CONDITION DRIVES PROGENY SEX‐RATIO BIAS IN A LIZARD THAT LACKS PARENTAL CARE

Sex‐allocation theory predicts that females in good condition should preferentially produce offspring of the sex that benefits the most from an increase in maternal investment. However, it is generally assumed that the condition of the sire has little effect on progeny sex ratio, particularly in species that lack parental care. We used a controlled breeding experiment and molecular paternity analyses to examine the effects of both maternal and paternal condition on progeny sex ratio and progeny fitness in the brown anole (Anolis sagrei), a polygynous lizard that lacks parental care. Contrary to the predictions of sex‐allocation theory, we found no relationship between maternal condition and progeny sex ratio. By contrast, progeny sex ratio shifted dramatically from female‐biased to male‐biased as paternal condition increased. This pattern was driven entirely by an increase in the production of sons as paternal condition improved. Despite strong natural selection favoring large size and high condition in both sons and daughters, we found no evidence that progeny survival was related to paternal condition. Our results emphasize the importance of considering the paternal phenotype in studies of sex allocation and highlight the need for further research into the pathways that link paternal condition to progeny fitness.     

Transmission of a cyclical translocation in two cranberry cultivars

The occurrence of cyclical translocation involving three non-homologous chromosomes and affecting pollen stainability has been observed in two cranberry cultivars: Howes, and Wilcox, a progeny of Howes. These cultivars were crossed with six normal cranberry cultivars to study the transmission of the cyclical translocation to their progeny. The translocational progeny were determined by pollen tetrad analysis. A total of 102 individuals (6 crosses) were analyzed in the progeny of Wilcox and 116 individuals (5 crosses) in the progeny of Howes. The ratios observed in the progeny of Wilcox and Howes were 71 translocated: 31 normal, and 79 translocated: 37 normal, respectively. The segregations deviated from the expected 1 translocated: 1 normal progeny ratio, but fit either a 3:1 or 2:1 ratio. The altered segregations may indicate the presence of a balanced lethal system located in the translocated segments of both Howes and Wilcox. Sterile individuals were found in the progeny of WilcoxxHowes, which could indicate that the two parents have non-identical translocations. The translocated progeny of both cultivars had a normal distribution for pollen stainability, which indicated that both the occurrence of crossing over in the interstitial region and the segregation of chromosomes are under polygenic control.     

适用于种群遗传管理的小熊猫微卫星亲子鉴定体系的建立

由于标记缺失、生产记录不详、亲权关系不明及部分个体来源不清等历史问题,中国小熊猫圈养种群存在谱系错漏、近亲繁殖等风险。近年来,随着小熊猫种群规模不断扩大,管理者们对谱系的准确性提出了更高的需求,亲子鉴定工作也成为了研究的重点。本文采用26个微卫星标记,对国内3个小熊猫圈养种群进行了亲缘关系运算,完成了相关谱系的查错与整理。26个位点多态性与稳定性良好,联合非亲排除概率达到0.9999以上,可解决国内小熊猫圈养种群的各类亲子鉴定需求。在单亲未知或双亲未知的情况下,8或11个位点组合可实现亲子鉴定。5个位点组合可进行个体识别。在小熊猫圈养管理过程中,应用一套亲子鉴定体系对小熊猫圈养的谱系进行查漏补缺,有利于制定科学的配对计划、避免近亲繁殖,对小熊猫种群保护有着重要意义。     

Indirect genetic estimates of breeding population size in the polyploid green sturgeon (Acipenser medirostris)

The utility of genetic measures for kinship reconstruction in polysomic species is not well evaluated. We developed a framework to test hypotheses about estimating breeding population size indirectly from collections of outmigrating green sturgeon juveniles. We evaluated a polysomic dataset, in allelic frequency and phenotypic formats, from green sturgeon to describe the relationship among known progeny from experimental families. The distributions of relatedness values for kin classes were used for reconstructing green sturgeon pedigrees from juveniles of unknown relationship. We compared three rarefaction functions that described the relationship between the number of kin groups and number of samples in a pedigree to estimate the annual abundance of spawners contributing to the threatened green sturgeon Southern Distinct Population Segment in the upper Sacramento River. Results suggested the estimated abundance of breeding green sturgeon remained roughly constant in the upper Sacramento River over a 5‐year period, ranging from 10 to 28 individuals depending on the year and rarefaction method. These results demonstrate an empirical understanding for the distribution of relatedness values among individuals is a benefit for assessing pedigree reconstruction methods and identifying misclassification rates. Monitoring of rare species using these indirect methods is feasible and can provide insight into breeding and ontogenetic behaviour. While this framework was developed for specific application to studying fish populations in a riverscape, the framework could be advanced to improve genetic estimation of breeding population size and to identify important breeding habitats of rare species when combined with finer‐scaled sampling of offspring.     

Exclusion probabilities for pedigree testing farm animals

Pedigree testing, using genetic markers, may be undertaken for a variety of situations, of which the classical paternity testing is only one. This has not always been made clear in the literature. Exclusion probabilities associated with various testing situations, including the use of autosomal or X-linked codominant marker systems with any number of alleles, are presented. These formulae can be used to determine the appropriate exclusion probability for the situation being investigated. One such situation is where sire groups of progeny are to be verified without knowledge of the dams' genotypes, in which case the classical paternity exclusion probability is too high, and if used may result in an optimistic declaration about the progeny that have not been excluded. On the other hand, if mating pairs are known then incorrect progeny can be excluded at a higher rate than suggested by paternity exclusion calculations. The formulae also assist in determining the usefulness of X-linked markers, particularly if the pedigree checks involve progeny of only one sex. A system of notation that is useful for the algebraic manipulation of genetic probabilities, including exclusion probabilities as presented here, is also given.