Linkage Analysis of the Multilocus Glucosephosphate Isomerase Isozyme System in Sunfish (Centrarchidae, Teleostii)

The purpose of the present investigation is to determine whether the two duplicated glucosephosphate isomerase (EC 5.3.1.9) loci Gpi-A and Gpi-B reside on the same chromosome in teleostean fishes. Interspecific sunfish hybrids were employed for the cross because of the different species-specific electrophoretic mobilities of the allelic isozymes at each GPI locus and because of their genomic compatibility. F1 sunfish hybrids, formed from a male warmouth (Lepomis gulosus) X female green sunfish (L. cyanellus) cross, were mated to form the F2 generation. The number of each of the nine different isozyme phenotypes, revealed by starch gel electrophoresis, was determined using 256 F2 individuals. The high frequency of recombinant phenotypes in the F2 generation indicated that the two GPI loci are not linked. An excess of F2 individuals heterozygous at both loci was observed and is interpreted as being caused by heterosis. The absence of linkage for the homologous loci encoding GPI subunits and for other multilocus isozyme systems is consistent with the postulate that the genomes of present-day vertebrates arose through one or more polyploidization events early in vertebrate evolution.     

Multilocus Behavior in Random Environments. II. Linkage Disequilibrium in an Additive Model

The effect of a stochastic environment on an additive, two-locus model of a diploid population is examined. The appropriate diffusion equation is derived and its asymptotic properties are approximated by an Orstein-Uhlenbeck process. The first and second order moments of this approximating process are given. The mean linkage disequilibrium will be nonzero if the alleles at the different loci are correlated. The sign of the mean disequilibrium is determined by the sign of the correlation.     

The Effects of Selection on Linkage Analysis for Quantitative Traits

The effects of within-sample selection on the outcome of analyses detecting linkage between genetic markers and quantitative traits were studied. It was found that selection by truncation for the trait of interest significantly reduces the differences between marker genotype means thus reducing the power to detect linked quantitative trait loci (QTL). The size of this reduction is a function of proportion selected, the magnitude of the QTL effect, recombination rate between the marker locus and the QTL, and the allele frequency of the QTL. Proportion selected was the most influential of these factors on bias, e.g., for an allele substitution effect of one standard deviation unit, selecting the top 80%, 50% or 20% of the population required 2, 6 or 24 times the number of progeny, respectively, to offset the loss of power caused by this selection. The effect on power was approximately linear with respect to the size of gene effect, almost invariant to recombination rate, and a complex function of QTL allele frequency. It was concluded that experimental samples from animal populations which have been subjected to even minor amounts of selection will be inefficient in yielding information on linkage between markers and loci influencing the quantitative trait under selection.     

Multilocus Patterns of Nucleotide Diversity, Population Structure and Linkage Disequilibrium in Boechera stricta, a Wild Relative of Arabidopsis

Information about polymorphism, population structure, and linkage disequilibrium (LD) is crucial for association studies of complex trait variation. However, most genomewide studies have focused on model systems, with very few analyses of undisturbed natural populations. Here, we sequenced 86 mapped nuclear loci for a sample of 46 genotypes of Boechera stricta and two individuals of B. holboellii, both wild relatives of Arabidopsis. Isolation by distance was significant across the species range of B. stricta, and three geographic groups were identified by structure analysis, principal coordinates analysis, and distance-based phylogeny analyses. The allele frequency spectrum indicated a genomewide deviation from an equilibrium neutral model, with silent nucleotide diversity averaging 0.004. LD decayed rapidly, declining to background levels in ~10 kb or less. For tightly linked SNPs separated by <1 kb, LD was dependent on the reference population. LD was lower in the specieswide sample than within populations, suggesting that low levels of LD found in inbreeding species such as B. stricta, Arabidopsis thaliana, and barley may result from broad geographic sampling that spans heterogeneous genetic groups. Finally, analyses also showed that inbreeding B. stricta and A. thaliana have ~45% higher recombination per kilobase than outcrossing A. lyrata.     

Toward a Wolbachia Multilocus Sequence Typing System: Discrimination of Wolbachia Strains Present in Drosophila Species

Among the diverse maternally inherited symbionts in arthropods, Wolbachia are the most common and infect over 20% of all species. In a departure from traditional genotyping or phylogenetic methods relying on single Wolbachia genes, the present study represents an initial Multilocus Sequence Typing (MLST) analysis to discriminate closely related Wolbachia pipientis strains, and additional data on sequence diversity in Wolbachia. We report a new phylogenetic characterization of four genes (aspC, atpD, sucB, and pdhB), and provide an expanded analysis of markers described in previous studies (16S rDNA, ftsZ, groEL, dnaA, and gltA). MLST analysis of the bacterial strains present in 16 different Drosophila–Wolbachia associations detected four distinct clonal complexes that also corresponded to maximum-likelihood identified phylogenetic clades. Among the 16 associations analyzed, six could not be assigned to MLST clonal complexes and were also shown to be in conflict with relationships predicted by maximum-likelihood phylogenetic inferences. The results demonstrate the discriminatory power of MLST for identifying strains and clonal lineages of Wolbachia and provide a robust foundation for studying the ecology and evolution of this widespread endosymbiont.     

Pleiotropic Overdominance and the Maintenance of Genetic Variation in Polygenic Characters

A model of selection is described in which optimizing phenotypic selection is combined with pleiotropic overdominance. Thus, the role that mutation commonly plays in models of phenotypic evolution is replaced by balancing selection. Expressions are provided for the equilibrium genetic variance in phenotype and for the heterozygosity. An approximate analysis of the transient properties of the model shows that, in certain circumstances, the behavior is quite similar to that of models based on the interaction of mutation and selection.     

用超显性模型估计遗传多态平衡的种群参数

基于超显性模型,采用数学方法阐述了种群遗传多样性的延续机制及平衡条件,从双等位基因导出的定律有较大的局限性,在许多教科书中常用的估计平衡态基因频数的定律只适用于双等位基因,本文用另一种方法导出一些公式,将其扩展到多等位基因,讨论了种群中基因数ｎ,遗传负载荷（Ｌ）,杂合频ＹＸ（Ｈｅ）和纯合频率（Ｈｏｍ）之间的关系。     

Discontinuity and Limited Linkage in the Homologous Recombination System of a Hyperthermophilic Archaeon

Genetic transformation of Sulfolobus acidocaldarius by a multiply marked pyrE gene provided a high-resolution assay of homologous recombination in a hyperthermophilic archaeon. Analysis of 100 Pyr⁺ transformants revealed that this recombination system could transfer each of 23 nonselected base pair substitutions to the recipient chromosome along with the selected marker. In 30% of the recombinants, donor markers were transferred as multiple blocks. In at least 40% of the recombinants, donor markers separated by 5 or 6 bp segregated from each other, whereas similar markers separated by 2 bp did not segregate. Among intermarker intervals, the frequency of recombination tract endpoints varied 40-fold, but in contrast to other recombination systems, it did not correlate with the length of the interval. The average length of donor tracts (161 bp) and the frequent generation of multiple tracts seemed generally consistent with the genetic properties observed previously in S. acidocaldarius conjugation. The efficiency with which short intervals of diverged pyrE sequence were incorporated into the genome raises questions about the threat of ectopic recombination in Sulfolobus spp. mediated by this apparently efficient yet permissive system.All cells and some viruses encode systems of homologous recombination (HR) which support the successful replication of their genomes. In eukaryotic cells, HR systems repair double-strand breaks and ensure proper chromosome segregation during meiosis (1, 2, 17). Double-strand break repair by eukaryotic HR has been studied intensively in yeast, where it has been shown to cause the net transfer of a short section of sequence from the intact DNA to the broken DNA in a unilateral, i.e., nonreciprocal, manner. Outside this central zone, the flanking segments of the two DNAs may also be exchanged, generating a crossover. The relative yields of noncrossover versus crossover events vary in different situations, and this appears to reflect the different ways in which displacement loops formed by strand invasion are ultimately resolved (1, 17, 36).In bacteria, HR helps reassemble replication forks disrupted by encounters with various DNA lesions (6, 20, 27). For practical reasons, however, genetic assays of bacterial HR typically follow the process of replacing a segment of a recipient chromosome or plasmid with a corresponding (i.e., homologous) donor DNA segment introduced into the cell. This “ends-out” mode of HR underlies the classical techniques of genetic mapping and strain construction of Escherichia coli and other bacteria. It also occurs in natural populations and contributes to genome evolution, as indicated by the “mosaic” patterns of sequence polymorphisms documented in various E. coli lineages (28). Thus, the functional properties of the host HR system combine with those of the DNA transfer systems to influence the rate of genetic exchange and the nature (including the abundance and average length) of the homologous DNA segments incorporated as a result.The importance of HR for genetic exchange and genome stability raises questions about its role in microorganisms highly diverged from model microbial species and adapted to extreme environments. Many archaea meet these criteria, but HR has not been examined extensively in archaea, particularly with respect to functional properties in vivo. The archaeal homologues of the RecA and Rad51/Dmc1 proteins, termed “RadA,” share a distinct motif structure which more closely resembles the eukaryotic than the bacterial consensus (32). In vitro, the RadA proteins of various hyperthermophilic archaea have been shown to bind single-stranded DNA (ssDNA) preferentially, thereby forming nucleoprotein filaments. This binding has been reported to stimulate ATP hydrolysis, displacement loop formation, and strand exchange (19, 26, 34). Evidence for HR in vivo includes observations that DNA of Pyrococcus and related archaea fragmented by gamma irradiation reassembles quickly in vivo (8) and that Sulfolobus species can be genetically transformed by linear DNA (7, 16, 22). In other archaea (methanogens and extreme halophiles), small, nonreplicating, circular DNAs have been observed to integrate into recipient genomes by reciprocal crossovers (11), and deletion of the radA gene of the extreme halophile Haloferax volcanii has demonstrated that the protein is essential for HR in that species but not for viability (37).In Sulfolobus spp., which grow optimally at about pH 3 and 80°C (12), auxotrophic mutants provide sensitive assays of recombination at particular chromosomal loci. Several studies suggest that in Sulfolobus acidocaldarius, many of these events require only short DNA segments. In conjugation assays, for example, more than 90% of randomly chosen pairs of 5-fluoroorotic acid (FOA)-resistant S. acidocaldarius mutants generated Pyr⁺ recombinants (31), despite the fact that about 95% of such mutations normally arise in the 594-bp pyrE coding sequence (14). Furthermore, when pyrE mutations of known positions were tested in pairwise combinations, the relative yield of recombinants did not decrease significantly until the separation became less than about 50 bp, indicating that donor sequences were transferred to the recipient chromosome as small segments (18).Although these results reveal genetic consequences of conjugation in S. acidocaldarius, they do not clarify whether these consequences primarily reflect the DNA transfer process or, alternatively, subsequent HR. For example, transfer of short DNA fragments from a donor cell to a recipient cell seems able to explain both the facile resolution of very closely spaced pyrE mutations (31) and the inefficient replacement of large pyrE deletions (18) in S. acidocaldarius conjugation. However, other Sulfolobus spp. transfer relatively large conjugative plasmids between cells (33), so a similar transfer capability must be considered for S. acidocaldarius. In that case, the observed “short-patch” nature of the exchanges would reflect processing of longer intervals of transferred DNA by the HR system to yield short replacement tracts; this would resemble noncrossover events in eukaryotic double-strand break repair (1, 17, 36) or the patches of DNA incorporated by transformation of Helicobacter pylori (21, 25). An apparent failure of circular DNA containing a full-length pyrE gene with a promoter to integrate into the S. acidocaldarius genome by a single reciprocal crossover raises other questions. Despite protection of the circular DNA against the host restriction system and scoring by PCR within several generations of transformation, no sequences of the nonreplicating vector could be detected in any of a number of independent transformants (22). Possible explanations include an extremely high frequency of reciprocal crossovers, leading to stochastic elimination of the nonselectable vector sequences within a short time, or an intrinsically nonreciprocal mode of the initial recombination, which simply copied the functional pyrE sequence onto the host chromosome. While the mechanistic basis remains unresolved, the observations themselves combine with the presence of unusual DNA enzymes (10) and the absence of otherwise conserved DNA repair proteins (13) to suggest that HR and related DNA transactions of Sulfolobus spp. and other hyperthermophilic archaea may have unusual functional features.As an important step toward understanding Sulfolobus HR in molecular terms, we developed a quantitative assay to analyze individual recombination events in S. acidocaldarius to high resolution. The results show that this HR process transfers markers from a short donor DNA to the recipient genome to generate a diversity of configurations in which both the length and number of replacement tracts vary widely. In addition, the HR system exhibited limited genetic linkage and readily resolved certain markers spaced only 5 or 6 bp apart.     

Overdominance interacts with linkage to determine the rate of adaptation to a new optimum

Overdominance, or a fitness advantage of a heterozygote over both homozygotes, can occur commonly with adaptation to a new optimum phenotype. We model how such overdominant polymorphisms can reduce the evolvability of diploid populations, uncovering a novel form of epistatic constraint on adaptation. The fitness load caused by overdominant polymorphisms can most readily be ameliorated by evolution at tightly linked loci; therefore, traits controlled by multiple loosely linked loci are predicted to be strongly constrained. The degree of constraint is also sensitive to the shape of the relationship between phenotype and fitness, and the constraint caused by overdominance can be strong enough to overcome the effects of clonal interference on the rate of adaptation for a trait. These results point to novel influences on evolvability that are specific to diploids and interact with genetic architecture, and they predict a source of stochastic variability in eukaryotic evolution experiments or cases of rapid evolution in nature.     

The Evolution of Multilocus Systems under Weak Selection

The evolution of multilocus systems under weak selection is investigated. Generations are discrete and nonoverlapping; the monoecious population mates at random. The number of multiallelic loci, the linkage map, dominance, and epistasis are arbitrary. The genotypic fitnesses may depend on the gametic frequencies and time. The results hold for s << c(min), where s and c(min) denote the selection intensity and the smallest two-locus recombination frequency, respectively. After an evolutionarily short time of t(1) ~ (ln s)/ln(1 - c(min)) generations, all the multilocus linkage disequilibria are of the order of s [i.e., O(s) as s -> 0], and then the population evolves approximately as if it were in linkage equilibrium, the error in the gametic frequencies being O(s). Suppose the explicit time dependence (if any) of the genotypic fitnesses is O(s(2)). Then after a time t(2) ~ 2t(1), the linkage disequilibria are nearly constant, their rate of change being O(s(2)). Furthermore, with an error of O(s(2)), each linkage disequilibrium is proportional to the corresponding epistatic deviation for the interaction of additive effects on fitness. If the genotypic fitnesses change no faster than at the rate O(s(3)), then the single-generation change in the mean fitness is ΔW = W(-1)V(g) + O(s(3)), where V(g) designates the genic (or additive genetic) variance in fitness. The mean of a character with genotypic values whose single-generation change does not exceed O(s(2)) evolves at the rate ΔZ = W(-1)C(g) + O(s(2)), where C(g) represents the genic covariance of the character and fitness (i.e., the covariance of the average effect on the character and the average excess for fitness of every allele that affects the character). Thus, after a short time t(2), the absolute error in the fundamental and secondary theorems of natural selection is small, though the relative error may be large.     

Multilocus analysis of hypertension: a hierarchical approach

While hypertension is a complex disease with a well-documented genetic component, genetic studies often fail to replicate findings. One possibility for such inconsistency is that the underlying genetics of hypertension is not based on single genes of major effect, but on interactions among genes. To test this hypothesis, we studied both single locus and multilocus effects, using a case-control design of subjects from Ghana. Thirteen polymorphisms in eight candidate genes were studied. Each candidate gene has been shown to play a physiological role in blood pressure regulation and affects one of four pathways that modulate blood pressure: vasoconstriction (angiotensinogen, angiotensin converting enzyme - ACE, angiotensin II receptor), nitric oxide (NO) dependent and NO independent vasodilation pathways and sodium balance (G protein-coupled receptor kinase, GRK4). We evaluated single site allelic and genotypic associations, multilocus genotype equilibrium and multilocus genotype associations, using multifactor dimensionality reduction (MDR). For MDR, we performed systematic reanalysis of the data to address the role of various physiological pathways. We found no significant single site associations, but the hypertensive class deviated significantly from genotype equilibrium in more than 25% of all multilocus comparisons (2,162 of 8,178), whereas the normotensive class rarely did (11 of 8,178). The MDR analysis identified a two-locus model including ACE and GRK4 that successfully predicted blood pressure phenotype 70.5% of the time. Thus, our data indicate epistatic interactions play a major role in hypertension susceptibility. Our data also support a model where multiple pathways need to be affected in order to predispose to hypertension.     

Two Modes of Balancing Selection in Drosophila Melanogaster: Overcompensation and Overdominance

Overdominance is often invoked to account for the extensive polymorphisms found in natural populations of organisms; overcompensation, however, may be equally or more important. Overcompensation occurs when limiting resources are better exploited by a genetically mixed than by a uniform population, and is often causally related to frequency-dependent selection. We have designed experiments to test whether overcompensation occurs in Drosophila melanogaster, using the Sod locus as a marker. Tests are made at each of two densities and two temperatures for cultures with desired genetic compositions. Both temperature and density have statistically significant effects on the per-female productivity of the cultures. More important, there are strong effects due to overcompensation. Cultures that are more polymorphic are also more productive than less polymorphic ones even when the level of individual heterozygosity is the same in all. There is also overdominance for the Sod locus: the heterozygotes are more productive than either homozygote at every temperature and density, and the differences are statistically significant in several cases. These results corroborate previous studies showing that overdominance may contribute to the maintenance of the Sod polymorphisms. Moreover, our results indicate that the significance of overcompensation as a mechanism to account for polymorphism in natural populations deserves further investigation.     

The Admixture Linkage Disequilibrium and Genetic Linkage Inference on the Gradual Admixture Population

Through the theoretical analysis of the admixture linkage disequilibrium (ALD) in the gradual admixture (GA) model, in which admixture occurs in every generation, the ALD is found to be proportional to the difference in marker allele frequencies, p₁-p₂, between two subpopulations. Based on this property, we can employ a strict monotonic function (Δ_ker=Δ/(p₁-p₂), where Δ denotes the linkage disequilibrium (LD)) of the recombination fraction between the marker locus and the disease locus to infer the true genetic linkage. We construct a quasi likelihood ratio test (LRT) for the case-only data utilizing the information of unlinked markers in the human genome. The simulation results show that our tests can be used to fine map a disease locus. The effects of parameter values in the ALD mapping are also discussed.