Emperical evidence for intraclonal genetic variation is described here for clonal systems using a variety of molecular techniques and implicating a diversity of mechanisms. However, clonal systems are still generally perceived as having strict genetic fidelity. As concepts of genetic variability move from primary sequence data to include epigenetic and structural influences on genetic expression, the ability to detect changes in the genome at short intervals allows precedence to be given to inherent biological variation that is often analytically ignored. Therefore, the advent of powerful molecular techniques, like genome mapping, mean that our concepts of genetic fidelity within eukaryotic clones and the whole philosophy of the ‘clone’ needs to be re-evaluated and re-defined to replace old unproven dogma in this aspect of science.

The co-occurrence of three chromosome-wide phenomena – imprinting, facultative heterochromatization and diffuse centromere – in the mealybug Planococcus lilacinus makes investigation of the genomics of this species an attractive prospect. In order to estimate the complexity of the genome of this species, 300 random stretches of its DNA, constituting ∼0·1% of the genome, were sequenced. Coding sequences appear to constitute ∼53·5%, repeat sequences ∼44·5% and non-coding single-copy sequences ∼2% of the genome. The proportion of repetitive sequences in the mealybug is higher than that in the fruit fly Drosophila melanogaster (∼30%). The mealybug genome (∼220 Mb) is about 1·3 times the size of the fly genome (∼165 Mb) and its GC content (∼35%) less than that of the fly genome (∼40%). The relative abundance of various dinucleotides, as analysed by the method of Gentles and Karlin, shows that the dinucleotide signatures of the two species are moderately similar and that in the mealybug there is neither over-representation nor under-representation of any dinucleotide.

We used polymorphic microsatellite markers to look for recombination during parthenogenetic oogenesis between the X chromosomes of aphids of the tribe Macrosiphini. We examined the X chromosome because it comprises ∼25% of the genome and previous cytological observations of chromosome pairing and nucleolar organizer (NOR) heteromorphism suggest recombination, although the same is not true for autosomes. A total of 564 parthenogenetic females of Myzus clones with three distinct reproductive modes (cyclical parthenogenesis, obligate parthenogenesis and obligate parthenogenesis with male production) were genotyped at three informative X-linked loci. Also, parthenogenetically produced males from clones encompassing the full range of male-producing reproductive strategies were genotyped. These included 391 Myzus persicae males that were genotyped at three X-linked loci and 538 males from Sitobion clones that were genotyped at five informative X-linked loci. Our results show no departure from clonality in parthenogenetic generations of aphids of the tribe Macrosiphini: no recombinant genotypes were observed in parthenogenetically produced males or females.

The nature of forces maintaining variation for quantitative traits can only be assessed at the level of individual genes affecting variation in the traits. Identification of single-nucleotide polymorphisms (SNPs) associated with variation in Drosophila sensory bristle number at the Delta (Dl) locus provides us with the opportunity to test a model for the maintenance of variation in bristle number by genotype by environment interaction (GEI). Under this model, genetic variation is maintained at a locus under stabilizing selection if phenotypic values of heterozygotes are more stable than homozygotes across a range of environments, and the mean allelic effect is much smaller than the standard deviation of allelic effects across environments. Homozygotes and heterozygotes for two SNPs at Dl, one affecting sternopleural and the other abdominal bristle number, were reared in five different environments. There was significant GEI for both bristle traits. Neither condition of the model was satisfied for Dl SNPs exhibiting GEI for sternopleural bristle number. Heterozygotes for the abdominal bristle number SNPs were indeed the most stable genotype for two of the three environment pairs exhibiting GEI, but the mean genotypic effect was greater than the standard deviation of effects across environments. Therefore, this mechanism of GEI seems unlikely to be responsible for maintaining the common bristle number polymorphisms at Dl.

Most aphids are cyclic parthenogens, so are ideal models in studies of the mechanisms and consequences of sex and recombination. However, owing to a shortage of physical and genetic markers, there have been few studies of the most fundamental genetic processes in these organisms. For example, it is not known whether autosomal segregation during male spermatogenesis is in Mendelian proportions: we address that question here. The aphid Myzus persicae has a typical karyotype of 2n = 12 in females (XX), while males are XO (2n = 11). During male meiosis, only the spermatocytes with an X chromosome are viable. We hypothesized that assortment of autosomes might be non-random because chromosomal imprinting leading to elimination of the paternal autosomes is seen in the closely related coccoids. In other aphid models, we have observed segregation distortions at single microsatellite loci (Wilson, 2000). Such distortions may have nothing to do with ‘selfish’ behaviour, but may be caused by mutation accumulation causing fitness differentials. Thus single-locus distortions might be predicted to be more likely to be detected via the male lines of clones that have lost the ability to reproduce sexually (male-producing obligate parthenogenesis (androcyclic)). Using microsatellites we show that genetic imprinting or selfish autosome behaviour does not occur in male M. persicae. Generally, loci segregated in Mendelian proportions in both sexes of cyclically parthenogenetic (holocyclic) clones. However, in androcyclic clones, segregation distortions consistently involved the same two autosomes. This is consistent with linkage of markers to deleterious mutations associated with a loss of sexual reproduction.

Chromatin integrity is maintained throughout the cell cycle through repair mechanisms and intrinsically by the ordered packaging of DNA in association with histone proteins; however, aberrant rearrangements within and between chromosomes do occur. The role of the nuclear matrix protein topoisomerase II (TopoII) in generating chromosome breakpoints has been a focus of recent investigations. TopoII preferentially binds in vitro to scaffold-associated regions (SARs) and is involved in many DNA processing activities that require chromosome untangling. SARs, biochemically defined DNA elements rich in A+T, have been proposed to serve as structural boundaries for chromatin loops and to delineate functional domains. In our investigation of gene compartmentalization in a eukaryotic genome, SAR-associated nucleotide motifs from Drosophila were mapped in the regions of three histone gene clusters in an in silico analysis of the genome of Caenorhabditis elegans. Sites with similarity to the 15 bp consensus for TopoII cleavage were found predominantly in A+T enriched intergenic regions. Reiteration of sites matching the TopoII core consensus led to the identification of a novel core histone gene on chromosome IV and provided evidence for duplication and inversion in each of the three histone gene clusters. Breakpoint analysis of DNA flanking reiterated regions revealed potential sites for TopoII cleavage and a base composition phenomenon suggestive of a trigger for inversion events.

Natural populations are structured spatially into local populations and genetically into diverse ‘genetic backgrounds’ defined by different combinations of selected alleles. If selection maintains genetic backgrounds at constant frequency then neutral diversity is enhanced. By contrast, if background frequencies fluctuate then diversity is reduced. Provided that the population size of each background is large enough, these effects can be described by the structured coalescent process. Almost all the extant results based on the coalescent deal with a single selected locus. Yet we know that very large numbers of genes are under selection and that any substantial effects are likely to be due to the cumulative effects of many loci. Here, we set up a general framework for the extension of the coalescent to multilocus scenarios and we use it to study the simplest model, where strong balancing selection acting on a set of n loci maintains 2n backgrounds at constant frequencies and at linkage equilibrium. Analytical results show that the expected linked neutral diversity increases exponentially with the number of selected loci and can become extremely large. However, simulation results reveal that the structured coalescent approach breaks down when the number of backgrounds approaches the population size, because of stochastic fluctuations in background frequencies. A new method is needed to extend the structured coalescent to cases with large numbers of backgrounds.

Rutidosis leptorrynchoides is a perennial forb endemic to grasslands and grassy woodlands in southeastern Australia. Studies of seed dispersal, spatial genetic structure and clonality were carried out in four populations around the Canberra region that varied in levels of correlated paternity to examine: (1) whether R. leptorrhynchoides populations exhibit fine-scale spatial genetic structure and whether this varies between populations as a function of correlated paternity; (2) whether there is a correlation between seed dispersal distance and genetic relatedness within populations; and (3) whether clonal reproduction occurs in this species and to what degree this could account for the observed spatial genetic structure. The results show that there is variation in the magnitude and extent of spatial genetic structure between R. leptorrhynchoides populations. The three larger populations, with low to moderate full-sib proportions, showed significant patterns of coancestry between plants over scales of up to one metre, whereas the smallest population, with a high full-sib proportion, had erratically high but non-significant coancestry values. The observed patterns of genetic clumping could be explained by a combination of limited seed dispersal and correlated mating owing to limited mate availability resulting from the species' sporophytic self-incompatibility system. Clonality does not appear to be an important factor contributing to genetic structure in this species.

In Drosophila melanogaster, the main cuticular hydrocarbons (HCs) are some of the pheromones involved in mate discrimination. These are sexually dimorphic in both their occurrence and their effects. The production of predominant HCs has been measured in male and female progeny of 220 PGal4 lines mated with the feminising UAS-transformer transgenic strain. In 45 lines, XY flies were substantially or totally feminised for their HCs. Surprisingly, XX flies of 14 strains were partially masculinised. Several of the PGal4 enhancer-trap variants screened here seem to interact with sex determination mechanisms involved in the control of sexually dimorphic characters. We also found a good relationship between the degree of HC transformation and GAL4 expression in oenocytes. The fat body was also involved in the switch of sexually dimorphic cuticular hydrocarbons but its effect was different between the sexes.

Markov chain Monte Carlo procedures allow the reconstruction of full-sibships using data from genetic marker loci only. In this study, these techniques are extended to allow the reconstruction of nested full- within half-sib families, and to present an efficient method for calculating the likelihood of the observed marker data in a nested family. Simulation is used to examine the properties of the reconstructed sibships, and of estimates of heritability and common environmental variance of quantitative traits obtained from those populations. Accuracy of reconstruction increases with increasing marker information and with increasing size of the nested full-sibships, but decreases with increasing population size. Estimates of variance component are biased, with the direction and magnitude of bias being dependent upon the underlying errors made during pedigree reconstruction.

Clines for size and stress resistance traits have been described for several Drosophila species and replicable clines across different species may indicate climatic selection. Here we consider clines in stress resistance traits in an Australian endemic species, D. serrata, by comparing levels of variation within and among isofemale lines initiated with flies collected from the eastern coast of Australia. We also consider clinal variation in chill coma recovery, a trait that has recently been shown to exhibit high levels of variation among Drosophila species. Patterns were compared with those in the cosmopolitan species D. melanogaster from the same area. Both desiccation and starvation resistance showed no clinal pattern despite heritable variation among isofemale lines. In contrast chill coma resistance exhibited a linear cline in the anticipated direction, resistance increasing with latitude. Body size was measured as wing length and body weight. Both traits showed geographic variation and strong non-linear clines with a sharp reduction in size in the tropics. These results are discussed in the context of climatic selection and evolutionary processes limiting species borders.

Resistance to organophosphorus insecticides (OP) in Culex pipiens mosquitoes represents a convenient model for investigating the fitness cost of resistance genes and its origin, since both the environmental changes in nature and the adaptive genes are clearly identified. Two loci are involved in this resistance – the super-locus Ester and the locus Ace.1 – each displaying several resistance alleles. Population surveys have shown differences in fitness cost between these resistance genes and even between resistance alleles of the same locus. In order to better understand this fitness cost and its variability, the effects of these resistance genes on several fitness-related traits are being studied. Here, through competition experiments between two males for the access to one female, we analysed the effect on paternity success associated with three resistance alleles – Ester4, Ester1 and Ace.1R – relative to susceptible males and relative to one another. The eventual effect of female genotype on male mating success was also studied by using susceptible and resistant females. The strains used in this experiment had the same genetic background. Susceptible males had a mating advantage when competing with any of the resistant males, suggesting a substantial cost of resistance genes to this trait. When competing against susceptible males, the paternity success did not vary among resistant males, whatever the genotype of the female. When competing against other resistant males, no difference in paternity success was apparent, except when the female was Ester1.

The logistic or S-shaped curve of growth is one of the few universal laws in biology. It is certain that there exist specific genes affecting growth curves, but, due to a lack of statistical models, it is unclear how these genes cause phenotypic differentiation in growth and developmental trajectories. In this paper we present a statistical model for detecting major genes responsible for growth trajectories. This model is incorporated with pervasive logistic growth curves under the maximum likelihood framework and, thus, is expected to improve over previous models in both parameter estimation and inference. The power of this model is demonstrated by an example using forest tree data, in which evidence of major genes affecting stem growth processes is successfully detected. The implications for this model and its extensions are discussed.

When 41 populations from Africa (south of the Sahara) and Indian Ocean islands were analysed for their chromosomal inversion polymorphism, 34 rearrangements were found, including the four common cosmopolitans (In(2L)t, In(2R)NS, In(3L)P and In(3R)P), four rare cosmopolitans (In(2L)NS, In(3R)C, In(3R)Mo and In(3R)K) and six African polymorphic (‘recurrent’) endemics. Mean inversion frequencies per major autosome arm were positively and, generally, highly correlated to each other. There was no altitudinal nor latitudinal cline of inversion frequency, except for one African polymorphic endemic. Significant longitudinal clines were detected for In(2L)t, In(3L)P and In(3R)K; in all cases, inversion frequencies decreased eastward. Principal components analysis and ANOVA made it possible to distinguish three groups of populations. A high level of polymorphism was found in populations from west tropical Africa. The other low altitude populations from the mainland were moderately polymorphic, whereas the lowest levels of polymorphism were those of high altitude populations and of Indian Ocean islands. Moreover, some regional and local differentiation was also found. The frequency of unique autosomal inversions was not different from those found in Asia, Australia and America, but was significantly higher than that in Europe and North Africa. A West–East differentiation was also observed for the African polymorphic endemics. The present geographic pattern suggests a long, patchy evolution with restricted gene flow, followed by the modern period with numerous recent migrations linked to human transportation.

Reproduction in individual animals of sexual species depends largely upon their ability to detect and distinguish specific signal(s) among those produced by various potential sexual partners. In Drosophila melanogaster males, there is a natural polymorphism for discrimination of female and male principal pheromones that segregates with chromosome 3. We have mapped two loci on chromosome 3 that change sex-pheromone discrimination in males. We successively exploited meiotic recombination, deficiencies and enhancer-trap strains; excision of the transposon in two selected enhancer-trap strains clearly reverted the discrimination phenotype. These results indicate that pheromonal discrimination is a character that can be genetically manipulated, and provide further insights into the evolution of the specific mate recognition system.

The progeny of Drosophila females doubly-mated to males from the same and a closely related species are mostly sired by conspecific males. We examined the genetic basis for conspecific mating preference and sperm precedence by using 186 Drosophila lines in which random chromosomal fragments of D. sechellia were introgressed into D. simulans. Sperm competition was measured for each of these lines by crossing ebony D. simulans female with ebony D. simulans males followed by wild-type males from the introgressed lines. Variation in sperm competition (proportion of progeny sired by the second male), mating discrimination (proportion of introgressed males that failed to remate), and male fecundity (proportion of progeny sired by introgressed males) were scored. The introgressed lines exhibited highly significant heterogeneity in the three phenotypes scored, motivating an analysis to locate quantitative trait loci (QTLs) responsible for the differences. Applying composite interval mapping, we found eight QTLs that explain a significant level of variation among introgressed lines in the phenotypes scored. Cytological position overlapped among some QTLs suggesting possible pleiotropic effects. Analysis of the joint effects of simulans/sechellia genetic composition at different QTLs and markers suggests that complex interactions among alleles are partially responsible for interspecific differences in sexual traits.

Amplified fragment length polymorphisms (AFLPs) are a widely used marker system: the technique is very cost-effective, easy and rapid, and reproducibly generates hundreds of markers. Unfortunately, AFLP alleles are typically scored as the presence or absence of a band and, thus, heterozygous and dominant homozygous genotypes cannot be distinguished. This results in a significant loss of information, especially as regards mapping of quantitative trait loci (QTLs). We present a Monte Carlo Markov Chain method that allows us to compute the identity by descent probabilities (IBD) in a general pedigree whose individuals have been typed for dominant markers. The method allows us to include the information provided by the fluorescent band intensities of the markers, the rationale being that homozygous individuals have on average higher band intensities than heterozygous individuals, as well as information from linked markers in each individual and its relatives. Once IBD probabilities are obtained, they can be combined into the QTL mapping strategy of choice. We illustrate the method with two simulated populations: an outbred population consisting of full sib families, and an F2 cross between inbred lines. Two marker spacings were considered, 5 or 20 cM, in the outbred population. There was almost no difference, for the practical purpose of QTL estimation, between AFLPs and biallelic codominant markers when the band density is taken into account, especially at the 5 cM spacing. The performance of AFLPs every 5 cM was also comparable to that of highly polymorphic markers (microsatellites) spaced every 20 cM. In economic terms, QTL mapping with a dense map of AFLPs is clearly better than microsatellite QTL mapping and little is lost in terms of accuracy of position. Nevertheless, at low marker densities, AFLPs or other biallelic markers result in very inaccurate estimates of QTL position.

We investigate how sampling of parents or children based on their extreme phenotypic values selected from clinical databases would affect the power of identification of quantitative trait loci (QTL) by a transmission disequilibrium test (TDT). We consider three selective sampling schemes based on the selection of phenotypic values of parents or children in nuclear families: (1) two children, one of extreme value, the other random; (2) two children extremely discordant; (3) one parent of extreme value. Other family members not specified will be recruited randomly with regard to phenotypic values. Our study shows that the second sampling scheme can always enhance the power for QTL identification, sometimes dramatically so. The increase in the statistical power of the TDT is particularly dramatic when h2 at the QTL under test is small or intermediate (e.g. 0·05 or 0·10). For the other two sampling schemes, under dominant effects at the QTL, the power is always increased relative to random sampling; however, under recessive or additive genetic effects, the power gain is generally minor or even decreased a little sometimes. Allele frequencies at the QTL and the selection stringency are important for determining the effect of selective sampling on the power of QTL identification. Our study is useful as a practical guideline on how to perform the TDT efficiently in practice by taking advantage of the extensive databases accumulated that are enriched with people of extreme phenotypic values.

Here, we introduce the idea of probabilities of line origins for alleles in general pedigrees as found in crosses between outbred lines. We also present software for calculating these probabilities. The proposed algorithm is based on the linear regression method of Haley, Knott and Elsen (1994) combined with the Markov chain Monte Carlo (MCMC) method for estimating quantitative trait locus coefficients used as regressors. We compared the relative precision of our method and the original method as proposed by Haley et al. (1994). The scenarios studied varied in the allelic distribution of marker alleles in parental lines and in the frequency of missing marker genotypes. We found that the MCMC method achieves a higher accuracy in all scenarios considered. The benefits of using MCMC approximation are substantial if the frequency of missing marker data is high or the number of marker alleles is low and the allelic frequency distribution is similar in both parental lines.

Here, we describe a randomization testing strategy for mapping interacting quantitative trait loci (QTLs). In a forward selection strategy, non-interacting QTLs and simultaneously mapped interacting QTL pairs are added to a total genetic model. Simultaneous mapping of epistatic QTLs increases the power of the mapping strategy by allowing detection of interacting QTL pairs where none of the QTL can be detected by their marginal additive and dominance effects. Randomization testing is used to derive empirical significance thresholds for every model selection step in the procedure. A simulation study was used to evaluate the statistical properties of the proposed randomization tests and for which types of epistasis simultaneous mapping of epistatic QTLs adds power. Least squares regression was used for QTL parameter estimation but any other QTL mapping method can be used. A genetic algorithm was used to search for interacting QTL pairs, which makes the proposed strategy feasible for single processor computers. We believe that this method will facilitate the evaluation of the importance at epistatic interaction among QTLs controlling multifactorial traits and disorders.