Patterns of DNA Variability at X-Linked Loci in Mus Domesticus

Introns of four X-linked genes (Hprt, Plp, Glra2, and Amg) were sequenced to provide an estimate of nucleotide diversity at nuclear genes within the house mouse and to test the neutral prediction that the ratio of intraspecific polymorphism to interspecific divergence is the same for different loci. Hprt and Plp lie in a region of the X chromosome that experiences relatively low recombination rates, while Glra2 and Amg lie near the telomere of the X chromosome, a region that experiences higher recombination rates. A total of 6022 bases were sequenced in each of 10 Mus domesticus and one M. caroli. Average nucleotide diversity (π) for introns within M. domesticus was quite low (π = 0.078%). However, there was substantial variation in the level of heterozygosity among loci. The two telomeric loci, Glra2 and Amg, had higher ratios of polymorphism to divergence than the two loci experiencing lower recombination rates. These results are consistent with the hypothesis that heterozygosity is reduced in regions with lower rates of recombination, although sampling of additional genes is needed to establish whether there is a general correlation between heterozygosity and recombination rate as in Drosophila melanogaster.     

DNA variation in a conifer,Cryptomeria japonica (Cupressaceae sensu lato)

We investigated the nucleotide variation of a conifer, Cryptomeria japonica, and the divergence between this species and its closest relative, Taxodium distichum, at seven nuclear loci (Acl5, Chi1, Ferr, GapC, HemA, Lcyb, and Pat). Samples of C. japonica were collected from three areas, Kantou-Toukai, Hokuriku, and Iwate. No apparent geographic differentiation was found among these samples. However, the frequency spectrum of the nucleotide polymorphism revealed excesses of intermediate-frequency variants, which suggests that the population was not panmictic and a constant size in the past. The average nucleotide diversity, pi, for silent sites was 0.00383. However, values of pi for silent sites vary among loci. Comparisons of polymorphism to divergence among loci (the HKA test) showed that the polymorphism at the Acl5 locus was significantly lower. We also observed a nearly significant excess of replacement polymorphisms at the Lcyb locus. These results suggested possibilities of natural selection acting at some of the loci. Intragenic recombination was detected only once at the Chi1 locus and was not detected at the other loci. The low level of population recombination rate, 4Nr, seemed to be due to both low level of recombination, r, and small population size, N.     

Microsatellite variation and recombination rate in the human genome

Background (purifying) selection on deleterious mutations is expected to remove linked neutral mutations from a population, resulting in a positive correlation between recombination rate and levels of neutral genetic variation, even for markers with high mutation rates. We tested this prediction of the background selection model by comparing recombination rate and levels of microsatellite polymorphism in humans. Published data for 28 unrelated Europeans were used to estimate microsatellite polymorphism (number of alleles, heterozygosity, and variance in allele size) for loci throughout the genome. Recombination rates were estimated from comparisons of genetic and physical maps. First, we analyzed 61 loci from chromosome 22, using the complete sequence of this chromosome to provide exact physical locations. These 61 microsatellites showed no correlation between levels of variation and recombination rate. We then used radiation-hybrid and cytogenetic maps to calculate recombination rates throughout the genome. Recombination rates varied by more than one order of magnitude, and most chromosomes showed significant suppression of recombination near the centromere. Genome-wide analyses provided no evidence for a strong positive correlation between recombination rate and polymorphism, although analyses of loci with at least 20 repeats suggested a weak positive correlation. Comparisons of microsatellites in lowest-recombination and highest-recombination regions also revealed no difference in levels of polymorphism. Together, these results indicate that background selection is not a major determinant of microsatellite variation in humans.     

Single nucleotide polymorphisms and recombination rate in humans

Levels of heterozygosity for single nucleotide polymorphisms vary by more than one order of magnitude in different regions of the human genome. Regional differences in the rate of recombination explain a substantial fraction of the variation in levels of nucleotide polymorphism, consistent with the widespread action of natural selection at the molecular level.     

Reduced Levels of DNA Polymorphism and Fixed between-Population Differences in the Centromeric Region of Drosophila Ananassae

We have estimated DNA sequence variation within and between two populations of Drosophila ananassae, using six-cutter restriction site variation at vermilion (v) and furrowed (fw). These two gene regions are located close to the centromere on the left and right X chromosome arms, respectively. In the fw region, no DNA polymorphism was detected within each population. In the v region, average heterozygosity per nucleotide was very low in both populations (pi = 0.0005 in the Burma population, and 0.0009 in the India population). These estimates are significantly lower than those from loci in more distal gene regions. The distribution of DNA polymorphisms between both populations was also striking. At fw, three fixed differences between the Burma and India populations were detected (two restriction site differences and one insertion/deletion of approximately 2 kb). At v, each DNA polymorphism in high frequency in the total sample was nearly fixed in one or the other population, although none of them reached complete fixation. The observed pattern of reduced variation within populations and fixed differences between populations appears to correlate with recombination rate. We conclude that recent hitchhiking associated with directional selection is the best explanation for this pattern. The data indicate that different selective sweeps have occurred in the two populations. The possible role of genetic hitchhiking in rapid population differentiation in gene regions of restricted recombination is discussed.     

Contrasting evolutionary histories of two introns of the duchenne muscular dystrophy gene, Dmd, in humans

The Duchenne muscular dystrophy (Dmd) locus lies in a region of the X chromosome that experiences a high rate of recombination and is thus expected to be relatively unaffected by the effects of selection on nearby genes. To provide a picture of nucleotide variability at a high-recombination locus in humans, we sequenced 5. 4 kb from two introns of Dmd in a worldwide sample of 41 alleles from Africa, Asia, Europe, and the Americas. These same regions were also sequenced in one common chimpanzee and one orangutan. Dramatically different patterns of genetic variation were observed at these two introns, which are separated by >500 kb of DNA. Nucleotide diversity at intron 44 pi = 0.141% was more than four times higher than nucleotide diversity at intron 7 pi = 0.034% despite similar levels of divergence for these two regions. Intron 7 exhibited significant linkage disequilibrium extending over 10 kb and also showed a significant excess of rare polymorphisms. In contrast, intron 44 exhibited little linkage disequilibrium and no skew in the frequency distribution of segregating sites. Intron 7 was much more variable in Africa than in other continents, while intron 44 displayed similar levels of variability in different geographic regions. Comparison of intraspecific polymorphism to interspecific divergence using the HKA test revealed a significant reduction in variability at intron 7 relative to intron 44, and this effect was most pronounced in the non-African samples. These results are best explained by positive directional selection acting at or near intron 7 and demonstrate that even genes in regions of high recombination may be influenced by selection at linked sites.     

Evolution of the genomic recombination rate in murid rodents

Although very closely related species can differ in their fine-scale patterns of recombination hotspots, variation in the average genomic recombination rate among recently diverged taxa has rarely been surveyed. We measured recombination rates in eight species that collectively represent several temporal scales of divergence within a single rodent family, Muridae. We used a cytological approach that enables in situ visualization of crossovers at meiosis to quantify recombination rates in multiple males from each rodent group. We uncovered large differences in genomic recombination rate between rodent species, which were independent of karyotypic variation. The divergence in genomic recombination rate that we document is not proportional to DNA sequence divergence, suggesting that recombination has evolved at variable rates along the murid phylogeny. Additionally, we document significant variation in genomic recombination rate both within and between subspecies of house mice. Recombination rates estimated in F(1) hybrids reveal evidence for sex-linked loci contributing to the evolution of recombination in house mice. Our results provide one of the first detailed portraits of genomic-scale recombination rate variation within a single mammalian family and demonstrate that the low recombination rates in laboratory mice and rats reflect a more general reduction in recombination rate across murid rodents.     

Using multilocus sequence data to assess population structure, natural selection, and linkage disequilibrium in wild tomatoes

We employed a multilocus approach to examine the effects of population subdivision and natural selection on DNA polymorphism in 2 closely related wild tomato species (Solanum peruvianum and Solanum chilense), using sequence data for 8 nuclear loci from populations across much of the species' range. Both species exhibit substantial levels of nucleotide variation. The species-wide level of silent nucleotide diversity is 18% higher in S. peruvianum (pi(sil) approximately 2.50%) than in S. chilense (pi(sil) approximately 2.12%). One of the loci deviates from neutral expectations, showing a clinal pattern of nucleotide diversity and haplotype structure in S. chilense. This geographic pattern of variation is suggestive of an incomplete (ongoing) selective sweep, but neutral explanations cannot be entirely dismissed. Both wild tomato species exhibit moderate levels of population differentiation (average F(ST) approximately 0.20). Interestingly, the pooled samples (across different demes) exhibit more negative Tajima's D and Fu and Li's D values; this marked excess of low-frequency polymorphism can only be explained by population (or range) expansion and is unlikely to be due to population structure per se. We thus propose that population structure and population/range expansion are among the most important evolutionary forces shaping patterns of nucleotide diversity within and among demes in these wild tomatoes. Patterns of population differentiation may also be impacted by soil seed banks and historical associations mediated by climatic cycles. Intragenic linkage disequilibrium (LD) decays very rapidly with physical distance, suggesting high recombination rates and effective population sizes in both species. The rapid decline of LD seems very promising for future association studies with the purpose of mapping functional variation in wild tomatoes.     

The influence of recombination on SNP diversity in chickens

The association between recombination rate and diversity, but not divergence is considered to be driven mainly by natural selection: fixation of positively selected variants and associated hitchhiking effects and/or background selection eliminating deleterious alleles. In the present study, we investigated the relationship between recombination rate, SNP diversity and interspecies divergence for 29 loci in chickens. We found that recombination rate is positively correlated with nucleotide diversity but is not correlated with interspecies divergence. It appears that variation in recombination rate explains over 30% of the variation in levels of diversity among 29 loci. Our data suggested that natural selection is a main factor in shaping SNP diversity in chickens. Since SNP diversity is significantly lower at Z-linked than at autosomal loci, we argued that genetic hitchhiking might be more important than background selection in producing the observed correlation.     

Nucleotide polymorphism at the Atmyb2 locus of the wild plant Arabidopsis thaliana

DNA variation was studied in a 2.2 kb region of the regulatory gene Atmyb2 using 20 ecotypes of Arabidopsis thaliana and one accession each of Arabis gemmifera and Arabidopsis himalaica. Nucleotide diversity (pi) in the region was 0.0027, which was lower than for other loci in A. thaliana. The MYB domain of the Atmyb2 gene (pi = 0.0036) had a larger variation than the non-MYB region (pi = 0.0013). Tajima's test and Fu and Li's test did not give a significant result. In contrast to the low level of polymorphism, the degree of divergence of the Atmyb2 region was higher between A. thaliana and A. gemmifera (K = 0.0730) than for other loci. The MYB domain (K = 0.0436) had smaller divergence than the non-MYB region (K = 0.0939). The HKA test detected significant discordance in the ratio of polymorphism to divergence in some comparisons. The pattern of low polymorphism and high divergence, which is mainly observed in the non-MYB region of the gene, is inconsistent with the neutral mutation theory. Strong purifying selection after establishment of A. thaliana and a species-specific adaptive process could be invoked to account for this pattern of polymorphism and divergence of Atmyb2.     

Variation in mutation rate and polymorphism among mitochondrial genes of Silene vulgaris

The prevailing wisdom of the plant mitochondrial genome is that it has very low substitution rates, thus it is generally assumed that nucleotide diversity within species will also be low. However, recent evidence suggests plant mitochondrial genes may harbor variable and sometimes high levels of within-species polymorphism, a result attributed to variance in the influence of selection. However, insufficient attention has been paid to the effect of among-gene variation in mutation rate on varying levels of polymorphism across loci. We measured levels of polymorphism in seven mitochondrial gene regions across a geographically wide sample of the plant Silene vulgaris to investigate whether individual mitochondrial genes accumulate polymorphisms equally. We found that genes vary significantly in polymorphism. Tests based on coalescence theory show that the genes vary significantly in their scaled mutation rate, which, in the absence of differences among genes in effective population size, suggests these genes vary in their underlying mutation rate. Further evidence that among-gene variance in polymorphism is due to variation in the underlying mutation rate comes from a significant positive relationship between the number of segregating sites and silent site divergence from an outgroup. Contrary to recent studies, we found unconvincing evidence of recombination in the mitochondrial genome, and generally confirm the standard model of plant mitochondria characterized by low substitution rates and no recombination. We also show no evidence of significant variation in the strength or direction of selection among genes; this result may be expected if there is no recombination. The present study provides some of the most thorough data on plant mitochondrial polymorphism, and provides compelling evidence for mutation rate variation among genes. The study also demonstrates the difficulty in establishing a null model of mitochondrial genome polymorphism, and thus the difficulty, in the absence of a comparative approach, in testing the assumption that low substitution rates in plant mitochondria lead to low polymorphism.     

The Rosy Region of Drosophila Melanogaster and Drosophila Simulans. I. Contrasting Levels of Naturally Occurring DNA Restriction Map Variation and Divergence

A 40-kb region around the rosy and snake loci was analyzed for restriction map variation among 60 lines of Drosophila melanogaster and 30 lines of Drosophila simulans collected together at a single locality in Raleigh, North Carolina. DNA sequence variation in D. simulans was estimated to be 6.3 times greater than in D. melanogaster (heterozygosities per nucleotide of 1.9% vs. 0.3%). This result stands in marked contrast to results of studies of phenotypic variation including proteins (allozymes), morphology and chromosome arrangements which are generally less variable and less geographically differentiated in D. simulans. Intraspecific polymorphism is not distributed uniformly over the 40-kb region. The level of heterozygosity per nucleotide varies more than 12-fold across the region in D. simulans, being highest over the hsc2 gene. Similar, though less extreme, variation in heterozygosity is also observed in D. melanogaster. Average interspecific divergence (corrected for intraspecific polymorphism) averaged 3.8%. The pattern of interspecific divergence over the 40-kb region shows some disparities with the spatial distribution of intraspecific variation, but is generally consistent with selective neutrality predictions: the most polymorphic regions within species are generally the most divergent between species. Sequence-length polymorphism is observed for D. melanogaster to be at levels comparable to other gene regions in this species. In contrast, no sequence length variation was observed among D. simulans chromosomes (limit of resolution approximately 100 bp). These data indicate that transposable elements play at best a minor role in the generation of naturally occurring genetic variation in D. simulans compared to D. melanogaster. We hypothesize that differences in species effective population size are the major determinant of the contrasting levels and patterns of DNA sequence and insertion/deletion variation that we report here and the patterns of allozyme and morphological variation and differentiation reported by other workers for these two species.     

Nuclear gene genealogies reveal historical,demographic and selective factors associated with speciation in field crickets

Population genetics theory predicts that genetic drift should eliminate shared polymorphism, leading to monophyly or exclusivity of populations, when the elapsed time between lineage-splitting events is large relative to effective population size. We examined patterns of nucleotide variation in introns at four nuclear loci to relate processes affecting the history of genes to patterns of divergence among natural populations and species. Ancestral polymorphisms were shared among three recognized species, Gryllus firmus, G. pennsylvanicus, and G. ovisopis, and genealogical patterns suggest that successive speciation events occurred recently and rapidly relative to effective population size. High levels of shared polymorphism among these morphologically, behaviorally, and ecologically distinct species indicate that only a small fraction of the genome needs to become differentiated for speciation to occur. Among the four nuclear gene loci there was a 10-fold range in nucleotide diversity, and patterns of polymorphism and divergence suggest that natural selection has acted to maintain or eliminate variation at some loci. While nuclear gene genealogies may have limited applications in phylogeography or other approaches dependent on population monophyly, they provide important insights into the historical, demographic, and selective forces that shape speciation.     

Nucleotide variation at the runt locus in Drosophila melanogaster and Drosophila simulans.

Intra- and interspecific nucleotide variation for the major developmental gene runt in Drosophila was studied in D. melanogaster and D. simulans. The 1.5-kb protein-coding region and the 0.4-kb intron of the runt gene were sequenced for 11 alleles in each species. The D. melanogaster alleles originated from east Africa. Estimated parameters of intraspecific variation in D. melanogaster (exons: theta = 0.018, pi = 0.018; intron: theta = 0.014, pi = 0.014) and D. simulans (exons: theta = 0.007, pi = 0.005; intron: theta = 0.008, pi = 0.005) were below average for other X-linked genes, while divergence between species (exons: D = 0.094; intron: D = 0.069) fell within the normal range for both silent and replacement changes. This estimate for runt, along with published values for three other genes in regions of normal recombination, show east African D. melanogaster to be roughly twice as polymorphic as D. simulans. The majority of nucleotide variation, silent and replacement, in both species was found to be selectively neutral using various statistical tests (HKA, McDonald-Kreitman, Tajima, and Fu and Li tests). Monte Carlo simulations of the coalescent process significantly rejected a Wright-Fisher model with respect to an amino acid polymorphism and the distribution of polymorphic sites among the D. simulans lines. This indicated an old lineage and may reflect ancestral population substructuring in D. simulans.     

A Method for Calibrating Molecular Clocks and Its Application to Animal Mitochondrial DNA

A generalized least-squares procedure is introduced for the calibration of molecular clocks and applied to the complete mitochondrial DNA sequences of 13 animal species. The proposed technique accounts for both nonindependence and heteroscedasticity of molecular-distance data, problems that have not been taken into to account in such analyses in the past. When sequence-identity data are transformed to account for multiple substitutions/site, the molecular divergence scales linearly with time, but with substantially more variation in the substitution rate than expected under a Poisson model. Significant levels of divergence are predicted at zero divergence time for most loci, suggesting high levels of site-specific heterozygosity among mtDNA molecules establishing in sister taxa. For nearly all loci, the baseline heterozygosity is lower and the substitution rate is higher in mammals relative to other animals. There is considerable variation in the evolutionary rate among loci but no compelling evidence that the average rate of mtDNA evolution is elevated with respect to that of nuclear DNA. Using the observed patterns of interspecific divergence, empirical estimates are derived for the mean coalescence times of organelles colonizing sister taxa.     

Development of a single nucleotide polymorphism map of porcine chromosome 2

Single nucleotide polymorphism markers are developed on SSC2, predominantly on the p-arm. Several studies reported a quantitative trait loci (QTL) for backfat thickness in this region. Single nucleotide polymorphisms were identified by comparative re-sequencing of polymerase chain reaction (PCR) products from a panel of eight individuals. The panel consisted of five Large Whites (each from a different Dutch breeding company), a Meishan, a Pietrain and a Wild Boar. In total, 67 different PCR products were sequenced and 301 SNPs were identified in 32,429 bp of consensus sequence, an average of one SNP in every 108 bp. After correction for sample size, this polymorphism rate corresponds to a heterozygosity value of one SNP in every 357 bp. For 63% of the SNPs, there was variation among the five Large Whites, and these SNPs are relevant for linkage and association studies in commercial populations. Comparing the Whites with other breeds revealed higher variation rates with: (i) Meishan, 89%; (ii) Pietrain, 69%; (iii) Wild Boar, 70%. Because many of the experimental populations to identify QTL are based on crosses between these breeds, these SNPs are relevant for the fine mapping of the QTL identified within these crosses.     

Mechanisms of genetic exchange within the chromosomal inversions of Drosophila pseudoobscura

We have used the inversion system of Drosophila pseudoobscura to investigate how genetic flux occurs among the gene arrangements. The patterns of nucleotide polymorphism at seven loci were used to infer gene conversion events between pairs of different gene arrangements. We estimate that the average gene conversion tract length is 205 bp and that the average conversion rate is 3.4 x 10(-6), which is 2 orders of magnitude greater than the mutation rate. We did not detect gene conversion events between all combinations of gene arrangements even though there was sufficient nucleotide variation for detection and sufficient opportunity for exchanges to occur. Genetic flux across the inverted chromosome resulted in higher levels of differentiation within 0.1 Mb of inversion breakpoints, but a slightly lower level of differentiation in central inverted regions. No gene conversion events were detected within 17 kb of an inversion breakpoint suggesting that the formation of double-strand breaks is reduced near rearrangement breakpoints in heterozygotes. At least one case where selection rather than proximity to an inversion breakpoint is responsible for reduction in polymorphism was identified.     

Genetic variation of Avicennia marina (Forsk.) Vierh. (Avicenniaceae) in Vietnam revealed by microsatellite and AFLP markers

Genetic variation of Avicennia marina in the costal area of Vietnam was examined using microsatellite and AFLP markers. By using five microsatellite loci a total of 21 alleles were detected. The average number of alleles per locus per population ranged from 1.667 to 3.000. The observed heterozygosity varied from 0.180 to 0.263, with an average of 0.210 indicating relatively low level of genetic variation comparing to the previous studies on A. marina in the worldwide range. The expected heterozygosity was larger than the observed heterozygosity leading to positive inbreeding coefficients in all the six populations. Highly significant departures from Hardy-Weinberg Equilibrium were detected in four populations. AFLP analysis revealed a total of 386 loci, of which 232 (60.1%) were polymorphic. In congruent with microsatellite markers relatively low levels of genetic variation were detected at both gene and nucleotide levels (H = 0.086; pi = 0.0054). Reduced level of genetic variation was found in the central population, and in the southern populations. Both microsatellite and AFLP markers revealed large genetic differentiation (F(ST) = 0.262 and 0.338, respectively) indicating strong genetic structure among regional populations. Pairwise genetic distance by AFLP showed two populations in the north and the other two in the south are closely related each other.     

Nucleotide diversity and linkage disequilibrium in wild avocado (Persea americana Mill.)

Resequencing studies provide the ultimate resolution of genetic diversity because they identify all mutations in a gene that are present within the sampled individuals. We report a resequencing study of Persea americana, a subtropical tree species native to Meso- and Central America and the progenitor of cultivated avocado. The sample includes 21 wild accessions from Mexico, Costa Rica, Ecuador, and the Dominican Republic. Estimated levels of nucleotide polymorphism and linkage disequilibrium (LD) are obtained from fully resolved haplotype data from 4 nuclear loci that span 5960 nucleotide sites. Results show that, although avocado is a subtropical tree crop and a predominantly outcrossing plant, the overall level of genetic variation is not exceptionally high (nucleotide diversity at silent sites, pi(sil) = 0.0102) compared with available estimates from temperate plant species. Intralocus LD decays rapidly to half the initial value within about 1 kb. Estimates of recombination rate (based on the sequence data) show that the rate is not exceptionally high when compared with annual plants such as wild barley or maize. Interlocus LD is significant owing to substantial population structure induced by mixing of the 3 botanical races of avocado.