Anthony D Long

• University of California, Irvine

The abundant and widely distributed deermice Peromyscus leucopus and P. maniculatus are important reservoirs for several different zoonotic agents in North America. For the pathogens they persistently harbor, these species are also examples of the phenomenon of infection tolerance. In the present study a prior observation of absent expression of th...

We use ATAC-seq to examine chromatin accessibility for four different tissues in Drosophila melanogaster: adult female brain, ovaries, and both wing and eye-antennal imaginal discs from males. Each tissue is assayed in eight different inbred strain genetic backgrounds, seven associated with a reference quality genome assembly. We develop a method f...

We carried out a 200 generation Evolve and Resequence (E&R) experiment initiated from an outbred diploid recombined 18-way synthetic base population. Replicate populations were evolved at large effective population sizes (>105 individuals), exposed to several different chemical challenges over 12 weeks of evolution, and whole-genome resequenced. We...

Drosophila melanogaster has proven an effective system with which to understand the evolutionary genetics and molecular mechanisms of insecticide resistance. Insecticide use has left signatures of selection in the fly genome, and both functional and quantitative genetics studies in the system have identified genes and variants associated with resis...

Drosophila melanogaster has proven an effective system with which to understand the evolutionary genetics and molecular mechanisms of insecticide resistance. Insecticide use has left signatures of selection in the fly genome, and both functional and quantitative genetics studies in the system have identified genes and variants associated with resis...

Deermice of the genus Peromyscus are well suited for addressing several questions of biologist interest, including the genetic bases of longevity, behavior, physiology, adaptation, and their ability to serve as disease vectors. Here we explore a diversity outbred approach for dissecting complex traits in Peromyscus leucopus, a non-traditional genet...

Despite the value of Recombinant Inbred Lines (RILs) for the dissection of complex traits, large panels can be difficult to maintain, distribute, and phenotype. An attractive alternative to RILs for many traits leverages selecting phenotypically extreme individuals from a segregating population, and subjecting pools of selected and control individu...

Little is known about the genetic architecture of antifungal immunity in natural populations. Using two population genetic approaches, Quantitative Trait Locus (QTL) Mapping and Evolve and Resequence (E&R), we explored D. melanogaster immune defense against infection with the fungus Beauveria bassiana. Immune defense was highly variable both in the...

Despite the value of Recombinant Inbred Lines (RILs) for the dissection of complex traits, large panels can be difficult to maintain, distribute, and phenotype. An attractive alternative to RILs for many traits leverages selecting phenotypically-extreme individuals from a segregating population, and subjecting pools of selected and control individu...

We describe the results of a 200 generation Evolve and Resequence (E&R) study initiated from an outbred dipliod recombined synthetic base population derived from 18 genetically diverse founders. Replicate populations were maintained at large effective population sizes (>10 ⁵ individuals), exposed to several different chemical challenges over 12 wee...

Although Peromyscus leucopus (deermouse) is not considered a genetic model system, its genus is well suited for addressing several questions of biologist interest, including the genetic bases of longevity, behavior, physiology, adaptation, and its ability to serve as a disease vector. Here we explore a diversity outbred approach for dissecting comp...

Background Despite marked recent improvements in long-read sequencing technology, the assembly of diploid genomes remains a difficult task. A major obstacle is distinguishing between alternative contigs that represent highly heterozygous regions. If primary and secondary contigs are not properly identified, the primary assembly will overrepresent b...

“Evolve and resequence” (E&R) studies combine experimental evolution and whole‐genome sequencing to interrogate the genetics underlying adaptation. Due to ease of handling, E&R work with asexual organisms such as bacteria can employ optimized experimental design, with large experiments and many generations of selection. By contrast, E&R experiments...

Background Despite marked recent improvements in long-read sequencing technology, the assembly of diploid genomes remains a difficult task. A major obstacle is distinguishing between alternative contigs that represent highly heterozygous regions. If primary and secondary contigs are not properly identified, the primary assembly will overrepresent b...

Advanced generation multi-parent populations (MPPs) are a valuable tool for dissecting complex traits, having more power than GWAS to detect rare variants, and higher resolution than F 2 linkage mapping. To extend the advantages of MPPs in budding yeast, we describe the creation and characterization of two outbred MPPs derived from eighteen genetic...

Advanced generation multi-parent populations (MPPs) are a valuable tool for dissecting complex traits, having more power than GWAS to detect rare variants, and higher resolution than F 2 linkage mapping. To extend the advantages of MPPs in budding yeast, we describe the creation and characterization of two outbred MPPs derived from eighteen genetic...

The cricetine rodents Peromyscus leucopus and P. maniculatus are key reservoirs for several zoonotic diseases in North America. We determined the complete circular mitochondrial genome sequences of representatives of 3 different stock colonies of P. leucopus, one stock colony of P. maniculatus and two wild populations of P. leucopus. The genomes we...

Using two advanced sequencing approaches, Illumina and PacBio, we derive the entire Dscam gene from an M2 assembly of the complete Penaeus monodon genome. The P. monodon Dscam (PmDscam) gene is ~266 kbp, with a total of 44 exons, 5 of which are subject to alternative splicing. PmDscam has a conserved architectural structure consisting of an extrace...

It has been hypothesized that individually-rare hidden structural variants (SVs) could account for a significant fraction of variation in complex traits. Here we identified more than 20,000 euchromatic SVs from 14 Drosophila melanogaster genome assemblies, of which ~40% are invisible to high specificity short-read genotyping approaches. SVs are com...

The rodent Peromyscus leucopus is the natural reservoir of several tick-borne infections, including Lyme disease. To expand the knowledge base for this key species in life cycles of several pathogens, we assembled and scaffolded the P. leucopus genome. The resulting assembly was 2.45 Gb in total length, with 24 chromosome-length scaffolds harboring...

Background In pancrustaceans, the Down syndrome cell adhesion molecule (Dscam) is an extraordinarily complex, single-locus gene, with the potential for generating thousands of isoforms by combining alternative splicing exons. In the present study, we used two advanced sequencing approaches, Illumina and PacBio, with hybrid assembly to analyze the e...

Pre-existing and de novo genetic variants can both drive adaptation to environmental changes, but their relative contributions and interplay remain poorly understood. Here we investigated the evolutionary dynamics in drug-treated yeast populations with different levels of pre-existing variation by experimental evolution coupled with time-resolved s...

Despite extensive effort to reveal the genetic basis of complex phenotypic variation, studies typically explain only a fraction of trait heritability. It has been hypothesized that individually rare hidden structural variants (SVs) could account for a significant fraction of variation in complex traits. To investigate this hypothesis, we assembled...

Vernal pool clam shrimp (Eulimnadia texana) are a promising model system due to their ease of lab culture, short generation time, modest sized genome, a somewhat rare stable androdioecious sex determination system, and a requirement to reproduce via desiccated diapaused eggs. We generated a highly contiguous genome assembly using 46X of PacBio long...

Standing and de novo genetic variants can both drive adaptation to environmental changes, but their relative contributions and interplay remain poorly understood. Here we investigated the dynamics of drug adaptation in yeast populations with different levels of standing variation by experimental evolution coupled with time-resolved sequencing and p...

Vernal pool clam shrimp ( Eulimnadia texana ) are a promising model system due to their ease of lab culture, short generation time, modest sized genome, a somewhat rare stable androdioecious sex determination system, and a requirement to reproduce via desiccated diapaused eggs. We generated a highly contiguous genome assembly using 46X of PacBio lo...

Vernal pool clam shrimp ( Eulimnadia texana ) are a promising model due to ease of culturing, short generation time, modest genome size, and obligate desiccated diapaused eggs. We collected Illumina data (Poolseq) from eleven pooled wild vernal pool clam shrimp populations. We hypothesized that restricted gene flow between vernal pools, separated b...

Fixation probability, the probability that the frequency of a newly arising mutation in a population will eventually reach unity, is a fundamental quantity in evolutionary genetics. Here we use a number of models (several versions of the Moran model and the haploid Wright-Fisher model) to examine fixation probabilities for a constant size populatio...

A major goal in the analysis of complex traits is to partition the observed genetic variation in a trait into components due to individual loci and perhaps variants within those loci. However, in both QTL mapping and genetic association studies, the estimated percent variation attributable to a QTL is upwardly biased conditional on it being discove...

The genetic component of complex disease risk in humans remains largely unexplained. A corollary is that the allelic spectrum of genetic variants contributing to complex disease risk is unknown. Theoretical models that relate population genetic processes to the maintenance of genetic variation for quantitative traits may suggest profitable avenues...

Supplemental note. (PDF)

Site frequency spectrum of neutral variants. For a sample n = 100 individuals, the relative site frequency spectrum is calculated as the proportion (y-axis) of all polymorphic sites which belong to each frequency class (x-axis). Sites with frequency was above 18 were grouped into one category to improve visualization. The data are grouped by λ, the...

Kurtosis of gamete properties. The kurtosis A) the number of mutations per gamete and B) the genetic value (sum of mutational effects) of a gamete over λ. The data are calculated for all risk mutations segregating in the simulated populations. Moments were calculated using the boost C++ statistical accumulators library. Data are plotted as the mean...

Genetic variance and load over time under the Tennessen et al.[40] model for European demography. The left column of panels shows how VG changes over time under this model. The right column shows how the mean number of deleterious mutations per individual changes. The models shown are: a = additive, g = GBR, and m = multiplicative with varying degr...

The burden ratio over time. The burden ratio [91] is calculated as the ratio of genetic load between simulations with only ancient growth and those with an additional recent bottleneck and growth. Here load is calculated as the average deviation from optimum fitness due to (left) fixed mutations, (middle) segregating mutations and (right) all mutat...

The probability density function of s. The probability of a new mutation with s = x on a log scale for various values of λ. The analytical result is an approximation obtained by assuming there is only a single deleterious mutation. For λ of 0.25 and 0.5 there is a large mass of lethals near s = 1. (TIFF)

Broad sense heritability. Broad-sense heritability, H2 = (VG)/(VP), as a function of λ: the mean effect size of a new deleterious mutation, as calculated explicitly from our simulated populations. Data are plotted as the mean across model replicates ± the standard error of the mean. The solid black horizontal line shows the predicted H2 under the r...

Relative heritability under rapid population growth. The y-axis is the ratio of mean broad-sense heritability under recent rapid growth to mean broad sense heritability for a constant-sized population, e.g. Mean[H2]growth/Mean[H2]constant. This ratio is plotted as a function of the mean effect size of causative mutations (λ). For co-dominant models...

Variance explained by additive and dominance effects at various frequencies. The percent of cumulative genetic variance explained by additive and dominance effects of variants with frequency less than or equal to a series of frequency values over λ. Shown here are the gene-based (GBR), additive co-dominant (AC), incomplete multiplicative recessive...

Frequency and effect sizes of risk variants. A) The mean frequency and B) mean effect size of a segregating risk variant over λ. Note they log10 y-axis scale in A. The mean effect size is the value pulled from the exponential distribution with mean λ, not the fitness effect or the quantitative genetic effect size. The data are calculated for all ri...

Average properties of haplotypes. A) The mean number of deleterious mutations per gamete in the population as a function of λ: the mean effect size of new causative mutation. The data plotted as mean over simulation replicates ±se. The data are calculated for the entire simulated population. B) The mean genetic value of a gamete, i.e. the average s...

Site frequency spectrum of risk and neutral variants. For a sample n = 100 individuals, the relative site frequency spectrum is calculated as the proportion (y-axis) of all polymorphic sites which belong to each frequency class (x-axis). Neutral variants are in orange and risk variants are shown in green. Y-axis is on a square-root scale and X-axis...

Distribution of significant hits under site based recessive models with incomplete dominance. Horizontal violin plots depict the distribution of minor allele frequencies (MAF) of the most strongly associated single marker in a GWAS. Individual hits are plotted as translucent points and jittered to provide a sense of the total number and density of...

Dominance for fitness effects versus dominance for trait effects. The dominance of fitness effects, shetshom, as a function of the dominance for trait effects, h. Values are based on idealized fitness effects of a mutation on a previously unaffected genetic background. The the relationship between fitness and trait dominance is influenced by the tr...

Empirical distribution of GWAS hits. Histogram GWAS hits (n = 1208) obtained from the NHGRI-EBI GWAS database for disease discussed in [26]. Data are described in S1 Table. (TIFF)

Additive genetic variance explained over allele frequency under the Tennessen et al. [40] model for European demography. (A) Population size change over time. Colored numbers represent population sizes at different times where we estimated the cumulative additive genetic variance (VA) as a function of allele frequency using regression (see Material...

The burden ratio based on mutations over time. The burden ratio [91] is calculated as the ratio of genetic load between simulations with only ancient growth and those with an additional recent bottleneck and growth. Here load is calculated as the average number of (left) fixed mutations, (middle) segregating mutations and (right) all mutations. The...

Site frequency spectrum of risk variants. For a sample n = 100 individuals, the relative site frequency spectrum is calculated as the proportion (y-axis) of all polymorphic sites which belong to each frequency class (x-axis). Sites with frequency was above 18 were grouped, into one category to improve visualization. The data are grouped by λ, the m...

The cumulative distribution of 2Ns. The probability of a new mutation with 2Ns ≤ x on a log scale for various values of λ. The dashed lines show the analytical result and the solid curves are empirical cumulative distribution functions based on a sample of 500 mutation effects from an exponential distribution. The analytical result is an approximat...

Mean Genetic Load. Genetic load(burden), L=wopt-w¯wopt, as a function of λ: the mean effect size of a new deleterious mutation. Data are plotted as the mean across model replicates ± the standard error of the mean. Solid curves show values for constant sized population simulations and dashed curves show values for rapid population expansion simulat...

Skew of gamete properties. The skewness A) the number of mutations per gamete and B) the genetic value (sum of mutational effects) of a gamete over λ. The data are calculated for all risk mutations segregating in the simulated populations. Moments were calculated using the boost C++ statistical accumulators library. Data are plotted as the mean acr...

Properties of Sample Heritability. Broad-sense heritability in a population sample of 6000 as a proportion of population wide broad-sense heritability. Data are grouped by demographic scenario, model and λ. The arbitrary dominance coefficient is parameterized such that h = 0 is complete recessivity, h = 1 would be exact co-dominance and h = 2 would...

Regression Based Estimates of Genetic Variance. (A) Regression estimates of variance explained by markers versus the classical formula 2pqα2. (B) Cumulative percent of variance explained across the risk allele frequency, based on regression estimates and classical formula. 1000 unlinked markers were simulated with effects drawn from an exponential...

Non-parametric comparison between empirical and simulated GWAS hits. A non-parametric comparison between distribution of allele frequencies between simulated and empirical GWAS hits. Shown are the -log10(p) values from the two-sample Kolmogorov-Smirnov test between the simulated and empirical allele frequencies. The lower and upper horizontal lines...

Details of Empirical GWAS Data. This table contains information specifying the empirical studies used in the manuscript. The data were obtained from the NHGRI-EBI catalog of published genome-wide association studies. All information fields contained in the download of the GWAS catalog are presented here. (XLSX)

Experimental evolutionary genomics now allows biologists to test fundamental theories concerning the genetic basis of adaptation. We have conducted one of the longest laboratory evolution experiments with any sexually-reproducing metazoan, Drosophila melanogaster. We used next-generation resequencing data from this experiment to examine genome-wide...

The genetic component of complex disease risk in humans remains largely unexplained. A corollary is that the allelic spectrum of genetic variants contributing to complex disease risk is unknown. Theoretical models that relate population genetic processes to the maintenance of genetic variation for quantitative traits may suggest profitable avenues...

Genome assemblies that are accurate, complete and contiguous are essential for identifying important structural and functional elements of genomes and for identifying genetic variation. Nevertheless, most recent genome assemblies remain incomplete and fragmented. While long molecule sequencing promises to deliver more complete genome assemblies wit...

Genome assemblies that are accurate, complete, and contiguous are essential for identifying important structural and functional elements of genomes and for identifying genetic variation. Nevertheless, most recent genome assemblies remain incomplete and fragmented. While long molecule sequencing promises to deliver more complete genome assemblies wi...

Genome-wide association studies (GWAS) have associated many single variants with complex disease, yet the better part heritable complex disease risk remains unexplained. Analytical tools designed to work under specific population genetic models are needed. Rare variants are increasingly shown to be important in human complex disease, but most existi...

Natural populations exhibit a great deal of interindividual genetic variation in the response to toxins, exemplified by the variable clinical efficacy of pharmaceutical drugs in humans, and the evolution of pesticide resistant insects. Such variation can result from several phenomena, including variable metabolic detoxification of the xenobiotic, a...

Variation in phenotype among DSPR RILs. Means (filled circles) and 1 SDs (vertical lines) for caffeine resistance. (PDF)

Founder haplotype means and 1-SDs for QTL mapped in just one DSPR population. The number of RILs for which we assign a founder genotype (probability > 0.95) is listed at the bottom of each bar. Only founder means associated with at least 5 observations are presented. (PDF)

Variation in phenotype among DGRP inbred lines. Means (filled circles) and 1 SDs (vertical lines) for caffeine resistance. (PDF)

Protein-coding genes implicated by mapped QTL. The "QTL" column indicates the QTL interval the gene is within. The "MappingPopulation" column indicates which population (pA and pB) the QTL was mapped in. The "ProteinCodingGene" column gives the symbol for the gene, and "GenePosition" gives the position of the gene in Release 5 of the Drosophila mel...

Results of GWAS for caffeine resistance loci using the DGRP. The data is filtered directly from the "gwas.all.assoc" file returned by the Mackay lab DGRP2 analysis pipeline (dgrp2.gnets.ncsu.edu; Accessed June 3, 2014). The "Chromosome" and "Position" columns give information on the variant under test, and the "MAF" column gives the minor allele fr...

Effect of CNV on Cyp12d1 expression in female heads. King et al. [36] generated array-based, genomewide expression data from female head tissue for 600 genotypes. Each genotype was the result of a cross between an independent pair of DSPR RILs. Using the normalized expression data for Cyp12d1, which represents a composite measure of expression from...

Copy number variation (CNV) genotype calls for the Cyp12d1 gene region in the DSPR and DGRP mapping panels. (PDF)

LOD scores for genomewide QTL scans in the DSPR. The "Chromosome" column indicates the chromosome arm of the position under test (X, 2L, 2R, 3L, and 3R). The "PhysicalPosition" and "GeneticPosition" columns indicate the physical and genetic positions of the site under test, respectively. The "LODscore" columns present the LOD scores at each positio...

Effect of copy number variation at Cyp12d1 on caffeine resistance in the DSPR and DGRP mapping panels. The plots show the mean phenotype (± 1-SD) of those lines with one copy of Cyp12d1 ("single") or two copies ("Dup"), with the number of lines in each class shown at the bottom of each bar. There is a significant effect of the duplication on caffei...

Expression levels of the five P450 genes under mapped QTL. Data is taken directly from the "gene_exp.diff" Cuffdiff output file. (PDF)

Vienna Drosophila Resource Center (VDRC) UAS-RNAi hairpin sequences for Cyp12d1-d and Cyp12d1-p genes. All data is taken directly from VDRC website (stockcenter.vdrc.at; Accessed January 8, 2015). Hairpin sequences designed to target one member of the gene pair have strong similarity with sequence from the other gene. Bases in hairpin sequences tha...

RNAseq-based expression data for all genes surviving a genomewide FDR threshold of 5% in at least one contrast. The data is taken directly from the "gene_exp.diff" Cuffdiff output file, and simply trimmed to remove data for those genes failing to reach genomewide significance (q < 0.05) in at least one contrast. The "Chromosome", "GeneStart", and "...

Raw caffeine resistance phenotypes measured in the DSPR and DGRP mapping panels. The "MappingPanel" column indicates which population the experimental genotype is derived from (the DSPR or DGRP). For DSPR genotypes, columns "Popn" and "PopnRep" indicate which population (pA or pB), and which subpopulation (pA1, pA2, pB1, pB2) the genotype is derive...

Inferred Cyp12d1 duplication status for all DSPR RILs. The "RIL" column provides the line code for all 1,715 genotypes phenotyped as part of this study. The "DuplicationStatus" column codes the copy number present at the Cyp12d1 gene; 0 = single copy, 1 = duplicated, NA = unknown. Only those RILs where the founder genotype at the Cyp12d1 locus is k...

Genome scan for caffeine resistance QTL after controlling for Cyp12d1 CNV status. The format of this plot is identical to that of Fig 1 in the main text, but with genomewide 5% permutation thresholds of 8.0 LOD (pA) and 7.2 LOD (pB). In the original genome scan we identified 10 QTL, three in both populations (Q1, Q2, Q3), one in pA only (Q9), and s...

Code used for RNAseq analysis. (PDF)

Evolve and resequence (E&R) experiments use experimental evolution to adapt populations to a novel environment, then next-generation sequencing to analyse genetic changes. They enable molecular evolution to be monitored in real time on a genome-wide scale. Here, we review the field of E&R experiments across diverse systems, ranging from simple non-...

For most complex traits we have a poor understanding of the positions, phenotypic effects, and population frequencies of the underlying genetic variants contributing to their variation. Recently, several groups have developed multi-parent advanced intercross mapping panels in different model organisms in an attempt to improve our ability to charact...

Transposable elements are a common source of genetic variation that may play a substantial role in contributing to gene expression variation. However, the contribution of transposable elements to expression variation thus far consists of a handful of examples. We used previously published gene expression data from 37 inbred Drosophila melanogaster...

Animals in nature are frequently challenged by toxic compounds, from those that occur naturally in plants as a defense against herbivory, to pesticides used to protect crops. On exposure to such xenobiotic substances, animals mount a transcriptional response, generating detoxification enzymes and transporters that metabolize and remove the toxin. G...

The severity of the toxic side effects of chemotherapy shows a great deal of interindividual variability, and much of this variation is likely genetically based. Simple DNA tests predictive of toxic side effects could revolutionize the way chemotherapy is carried out. Due to the challenges in identifying polymorphisms that affect toxicity in humans...

In "evolve-and-resequence" (E&R) experiments, whole-genome sequence data from laboratory-evolved populations can potentially uncover mechanisms of adaptive change. E&R experiments with initially isogenic, asexually-reproducing microbes have repeatedly shown that beneficial de novo mutations drive adaptation, and these mutations are not shared among...

Modern genetic mapping is plagued by the "missing heritability" problem, which refers to the discordance between the estimated heritabilities of quantitative traits and the variance accounted for by mapped causative variants. One major potential explanation for the missing heritability is allelic heterogeneity, in which there are multiple causative...

A novel approach for dissecting complex traits is to experimentally evolve laboratory populations under a controlled environment shift, resequence the resulting populations, and identify SNPs and/or genomic regions highly diverged in allele frequency. To better understand the power and localization ability of such an evolve and resequence approach,...

Human genome-wide association studies (GWAS) of longevity attempt to identify alleles at different frequencies in the extremely old, relative to a younger control sample. Here, we apply a GWAS approach to “synthetic” populations of Drosophila melanogaster derived from a small number of inbred founders. We used next-generation DNA sequencing to esti...

Here we present computational machinery to efficiently and accurately identify transposable element (TE) insertions in 146 next-generation sequenced inbred strains of Drosophila melanogaster. The panel of lines we use in our study is composed of strains from a pair of genetic mapping resources; the Drosophila Genetic Reference Panel (DGRP) and the...

The severity of the toxic side effects of chemotherapy varies among patients and much of this variation is likely genetically based. Here, we use the model system Drosophila melanogaster to genetically dissect toxicity of methotrexate (MTX), a drug used primarily to treat childhood acute lymphoblastic leukemia and rheumatoid arthritis. We utilize t...

Broad-sense heritability in different parts of the parameter space. (a) The deleterious mutation rate has an approximately linear effect on broad-sense heritability at large mean effect sizes of causative mutations (λ). All model parameters except the deleterious mutation rate (μd) are the same as in Figure 1d (see Methods). (b) The mean broad-sens...