The germline mutation rate determines the pace of genome evolution and is an evolving parameter itself1. However, little is known about what determines its evolution, as most studies of mutation rates have focused on single species with different methodologies2. Here we quantify germline mutation rates across vertebrates by sequencing and comparing the high-coverage genomes of 151 parent–offspring trios from 68 species of mammals, fishes, birds and reptiles. We show that the per-generation mutation rate varies among species by a factor of 40, with mutation rates being higher for males than for females in mammals and birds, but not in reptiles and fishes. The generation time, age at maturity and species-level fecundity are the key life-history traits affecting this variation among species. Furthermore, species with higher long-term effective population sizes tend to have lower mutation rates per generation, providing support for the drift barrier hypothesis3. The exceptionally high yearly mutation rates of domesticated animals, which have been continually selected on fecundity traits including shorter generation times, further support the importance of generation time in the evolution of mutation rates. Overall, our comparative analysis of pedigree-based mutation rates provides ecological insights on the mutation rate evolution in vertebrates.

Background Understanding the rate and pattern of germline mutations is of fundamental importance for understanding evolutionary processes. Results Here we analyzed 19 parent-offspring trios of rhesus macaques (Macaca mulatta) at high sequencing coverage of ∼76× per individual and estimated a mean rate of 0.77 × 10−8de novo mutations per site per generation (95% CI: 0.69 × 10−8 to 0.85 × 10−8). By phasing 50% of the mutations to parental origins, we found that the mutation rate is positively correlated with the paternal age. The paternal lineage contributed a mean of 81% of the de novo mutations, with a trend of an increasing male contribution for older fathers. Approximately 3.5% of de novo mutations were shared between siblings, with no parental bias, suggesting that they arose from early development (postzygotic) stages. Finally, the divergence times between closely related primates calculated on the basis of the yearly mutation rate of rhesus macaque generally reconcile with divergence estimated with molecular clock methods, except for the Cercopithecoidea/Hominoidea molecular divergence dated at 58 Mya using our new estimate of the yearly mutation rate. Conclusions When compared to the traditional molecular clock methods, new estimated rates from pedigree samples can provide insights into the evolution of well-studied groups such as primates.

In the past decade, several studies have estimated the human per-generation germline mutation rate using large pedigrees. More recently, estimates for various non-human species have been published. However, methodological differences among studies in detecting germline mutations and estimating mutation rates make direct comparisons difficult. Here, we describe the many different steps involved in estimating pedigree-based mutation rates, including sampling, sequencing, mapping, variant calling, filtering, and how to appropriately account for false-positive and false-negative rates. For each step, we review the different methods and parameter choices that have been used in the recent literature. Additionally, we present the results from a 'Mutationathon', a competition organized among five research labs to compare germline mutation rate estimates for a single pedigree of rhesus macaques. We report almost a two-fold variation in the final estimated rate among groups using different post-alignment processing, calling, and filtering criteria and provide details into the sources of variation across studies. Though the difference among estimates is not statistically significant, this discrepancy emphasizes the need for standardized methods in mutation rate estimations and the difficulty in comparing rates from different studies. Finally, this work aims to provide guidelines for computational and statistical benchmarks for future studies interested in identifying germline mutations from pedigrees.

The accurate and complete assembly of both haplotype sequences of a diploid organism is essential to understanding the role of variation in genome functions, phenotypes, and diseases1. Here, using a trio-binning approach, we present a high-quality, diploid reference genome, with both haplotypes assembled independently at the chromosome level, for the common marmoset (Callithrix jacchus), an important primate model system widely used in biomedical research2,3. The full heterozygosity spectrum between the two haplotypes involves 1.36% of the genome, much higher than the 0.13% indicated by the standard single nucleotide heterozygosity estimation alone. The de novo mutation rate is 0.43 × 10-8 per site per generation, where the paternal inherited genome acquired twice as many mutations as the maternal. Our diploid assembly enabled us to discover a recent expansion of the sex differentiated region and unique evolutionary changes in the marmoset Y chromosome. Additionally, we identified many genes with signatures of positive selection that might have contributed to the evolution of Callithrix biological features. Brain related genes were highly conserved between marmosets and humans, though several genes experienced lineage-specific copy number variations or diversifying selection, providing important implications for the application of marmosets as a model system.

32Understanding the rate and pattern of germline mutations is of fundamental importance for 33 understanding evolutionary processes. Here we analyzed 19 parent-offspring trios of rhesus macaques 34 (Macaca mulatta) at high sequencing coverage of ca. 76X per individual, and estimated an average 35 r a t e of 0.73 × 10 −8 de novo mutations per site per generation (95 % CI: 0.65 × 10 −8 -0.81 × 36 10 −8 ). By phasing 50 % of the mutations to parental origins, we found that the mutation rate is 37 positively correlated with the paternal age. The paternal lineage contributed an average of 80 % of 38 the de novo mutations, with a trend of an increasing male contribution for older fathers. About 1.9 39 % of de novo mutations were shared between siblings, with no parental bias, suggesting that they 40 arose from early development (postzygotic) stages. Finally, the divergence times between closely 41 related primates calculated based on the yearly mutation rate of rhesus macaque generally 42 reconcile with divergence estimated with molecular clock methods, except for the 43 Cercopithecidae/Hominoidea molecular divergence dated at 54 Mya using our new estimate of the 44 yearly mutation rate. 45

Background The Nile rat (Avicanthis niloticus) is an important animal model because of its robust diurnal rhythm, a cone-rich retina, and a propensity to develop diet-induced diabetes without chemical or genetic modifications. A closer similarity to humans in these aspects, compared to the widely used Mus musculus and Rattus norvegicus models, holds the promise of better translation of research findings to the clinic. Results We report a 2.5 Gb, chromosome-level reference genome assembly with fully resolved parental haplotypes, generated with the Vertebrate Genomes Project (VGP). The assembly is highly contiguous, with contig N50 of 11.1 Mb, scaffold N50 of 83 Mb, and 95.2% of the sequence assigned to chromosomes. We used a novel workflow to identify 3613 segmental duplications and quantify duplicated genes. Comparative analyses revealed unique genomic features of the Nile rat, including some that affect genes associated with type 2 diabetes and metabolic dysfunctions. We discuss 14 genes that are heterozygous in the Nile rat or highly diverged from the house mouse. Conclusions Our findings reflect the exceptional level of genomic resolution present in this assembly, which will greatly expand the potential of the Nile rat as a model organism.

In the past decade, several studies have estimated the human per-generation germline mutation rate using large pedigrees. More recently, estimates for various non-human species have been published. However, methodological differences among studies in detecting germline mutations and estimating mutation rates make direct comparisons difficult. Here, we describe the many different steps involved in estimating pedigree-based mutation rates, including sampling, sequencing, mapping, variant calling, filtering, and how to appropriately account for false-positive and false-negative rates. For each step, we review the different methods and parameter choices that have been used in the recent literature. Additionally, we present the results from a "Mutationathon", a competition organized among five research labs to compare germline mutation rate estimates for a single pedigree of rhesus macaques. We report almost a two-fold variation in the final estimated rate among groups using different post-alignment processing, calling, and filtering criteria and provide details into the sources of variation across studies. Though the difference among estimates is not statistically significant, this discrepancy emphasizes the need for standardized methods in mutation rate estimations and the difficulty in comparing rates from different studies. Finally, this work aims to provide guidelines for computational and statistical benchmarks for future studies interested in identifying germline mutations from pedigrees.

Lewontin's paradox, the observation that levels of genetic diversity (π) among animals do not scale linearly with variation in census population sizes (Nc), is an evolutionary conundrum, where the most extreme mismatches between π and Nc are found for highly abundant marine invertebrates. Yet, whether new mutations influence π relative to extrinsic processes remains unknown for most taxa. Here, we provide the first direct germline mutation rate (μ) estimate for a marine invertebrate, using high-coverage (60x) whole-genome sequencing of wild-caughtAcanthaster cf. solariscrown-of-thorns sea stars (Echinodermata). We also provide empirical estimates of adult Nc in Australia's Great Barrier Reef to jointly examine the determinants of π. Based on direct observations of 63 de novo mutations across 14 parent-offspring trios, theA. cf. solarismean μ was 9.13 x 10-09 mutations per-site per-generation (95% CI: 6.51 x 10-09 to 1.18 x 10-08). This value exceeds estimates for other invertebrates, showing greater concordance with reported vertebrate germline mutation rates. Lower-than-expected Ne (~70,000-180,000) and low Ne/Nc values (0.0047-0.048) indicated significant genetic drift and weak influences of contemporary population outbreaks on long-term π. Our findings of elevated μ and low Ne inA. cf. solarismay help explain high mutational loads and extreme polymorphism levels observed in some marine invertebrate taxa and are consistent with μ evolving in response to Ne (drift-barrier hypothesis). This study advances our understanding of the processes controlling levels of natural genetic variation and provides new data valuable for further testing hypotheses about mutation rate evolution across animal phyla.