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1. Expressed exome capture sequencing: A method for cost‐effective exome sequencing for all organisms

   https://doi.org/10.1111/1755-0998.12905  

   Puritz, Jonathan B. ; Lotterhos, Katie E. ( June 2018 , Molecular Ecology Resources)

   Exome capture is an effective tool for surveying the genome for loci under selection. However, traditional methods require annotated genomic resources. Here, we present a method for creatingcDNAprobes from expressedmRNA, which are then used to enrich and capture genomicDNAfor exon regions. This approach, called “EecSeq,” eliminates the need for costly probe design and synthesis. We tested EecSeq in the eastern oyster,Crassostrea virginica, using a controlled exposure experiment. Four adult oysters were heat shocked at 36°C for 1 hr along with four control oysters kept at 14°C. StrandedmRNAlibraries were prepared for two individuals from each treatment and pooled. Half of the combined library was used for probe synthesis, and half was sequenced to evaluate capture efficiency. GenomicDNAwas extracted from all individuals, enriched via captured probes, and sequenced directly. We found that EecSeq had an average capture sensitivity of 86.8% across all known exons and had over 99.4% sensitivity for exons with detectable levels of expression in themRNAlibrary. For all mapped reads, over 47.9% mapped to exons and 37.0% mapped to expressed targets, which is similar to previously published exon capture studies. EecSeq displayed relatively even coverage within exons (i.e., minor “edge effects”) and even coverage across exonGCcontent. We discovered 5,951SNPs with a minimum average coverage of 80×, with 3,508SNPs appearing in exonic regions. We show that EecSeq provides comparable, if not superior, specificity and capture efficiency compared to costly, traditional methods.

    

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2. A bioinformatic pipeline for identifying informative SNP panels for parentage assignment from RAD seq data

   https://doi.org/10.1111/1755-0998.12910  

   Andrews, Kimberly R. ; Adams, Jennifer R. ; Cassirer, E. Frances ; Plowright, Raina K. ; Gardner, Colby ; Dwire, Maggie ; Hohenlohe, Paul A. ; Waits, Lisette P. ( July 2018 , Molecular Ecology Resources)

   The development of high‐throughput sequencing technologies is dramatically increasing the use of single nucleotide polymorphisms (SNPs) across the field of genetics, but most parentage studies of wild populations still rely on microsatellites. We developed a bioinformatic pipeline for identifyingSNPpanels that are informative for parentage analysis from restriction site‐associatedDNAsequencing (RADseq) data. This pipeline includes options for analysis with or without a reference genome, and provides methods to maximize genotyping accuracy and select sets of unlinked loci that have high statistical power. We test this pipeline on small populations of Mexican gray wolf and bighorn sheep, for which parentage analyses are expected to be challenging due to low genetic diversity and the presence of many closely related individuals. We compare the results of parentage analysis acrossSNPpanels generated with or without the use of a reference genome, and betweenSNPs and microsatellites. For Mexican gray wolf, we conducted parentage analyses for 30 pups from a single cohort where samples were available from 64% of possible mothers and 53% of possible fathers, and the accuracy of parentage assignments could be estimated because true identities of parents were known a priori based on field data. For bighorn sheep, we conducted maternity analyses for 39 lambs from five cohorts where 77% of possible mothers were sampled, but true identities of parents were unknown. Analyses with and without a reference genome producedSNPpanels with ≥95% parentage assignment accuracy for Mexican gray wolf, outperforming microsatellites at 78% accuracy. Maternity assignments were completely consistent across allSNPpanels for the bighorn sheep, and were 74.4% consistent with assignments from microsatellites. Accuracy and consistency of parentage analysis were not reduced when using as few as 284SNPs for Mexican gray wolf and 142SNPs for bighorn sheep, indicating our pipeline can be used to developSNPgenotyping assays for parentage analysis with relatively small numbers of loci.

    

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3. RAD cap: sequence capture of dual‐digest RAD seq libraries with identifiable duplicates and reduced missing data

   https://doi.org/10.1111/1755-0998.12566  

   Hoffberg, Sandra L. ; Kieran, Troy J. ; Catchen, Julian M. ; Devault, Alison ; Faircloth, Brant C. ; Mauricio, Rodney ; Glenn, Travis C. ( August 2016 , Molecular Ecology Resources)

   Molecular ecologists seek to genotype hundreds to thousands of loci from hundreds to thousands of individuals at minimal cost per sample. Current methods, such as restriction‐site‐associatedDNAsequencing (RADseq) and sequence capture, are constrained by costs associated with inefficient use of sequencing data and sample preparation. Here, we introduceRADcap, an approach that combines the major benefits ofRADseq (low cost with specific start positions) with those of sequence capture (repeatable sequencing of specific loci) to significantly increase efficiency and reduce costs relative to current approaches.RADcap uses a new version of dual‐digestRADseq (3RAD) to identify candidateSNPloci for capture bait design and subsequently uses custom sequence capture baits to consistently enrich candidateSNPloci across many individuals. We combined this approach with a new library preparation method for identifying and removingPCRduplicates from 3RADlibraries, which allows researchers to processRADseq data using traditional pipelines, and we tested theRADcap method by genotyping sets of 96–384Wisteriaplants. Our results demonstrate that ourRADcap method: (i) methodologically reduces (to <5%) and allows computational removal ofPCRduplicate reads from data, (ii) achieves 80–90% reads on target in 11 of 12 enrichments, (iii) returns consistent coverage (≥4×) across >90% of individuals at up to 99.8% of the targeted loci, (iv) produces consistently high occupancy matrices of genotypes across hundreds of individuals and (v) costs significantly less than current approaches.

    

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4. Signatures of positive selection and local adaptation to urbanization in white‐footed mice ( Peromyscus leucopus )

   https://doi.org/10.1111/mec.14369  

   Harris, Stephen E. ; Munshi‐South, Jason ( October 2017 , Molecular Ecology)

   Urbanization significantly alters natural ecosystems and has accelerated globally. Urban wildlife populations are often highly fragmented by human infrastructure, and isolated populations may adapt in response to local urban pressures. However, relatively few studies have identified genomic signatures of adaptation in urban animals. We used a landscape genomic approach to examine signatures of selection in urban populations of white‐footed mice (Peromyscus leucopus) in New York City. We analysed 154,770SNPs identified from transcriptome data from 48P. leucopusindividuals from three urban and three rural populations and used outlier tests to identify evidence of urban adaptation. We accounted for demography by simulating a neutralSNPdata set under an inferred demographic history as a null model for outlier analysis. We also tested whether candidate genes were associated with environmental variables related to urbanization. In total, we detected 381 outlier loci and after stringent filtering, identified and annotated 19 candidate loci. Many of the candidate genes were involved in metabolic processes and have well‐established roles in metabolizing lipids and carbohydrates. Our results indicate that white‐footed mice in New York City are adapting at the biomolecular level to local selective pressures in urban habitats. Annotation of outlier loci suggests selection is acting on metabolic pathways in urban populations, likely related to novel diets in cities that differ from diets in less disturbed areas.

    

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5. Temperature gradient affects differentiation of gene expression and SNP allele frequencies in the dominant Lake Baikal zooplankton species

   https://doi.org/10.1111/mec.14704  

   Bowman, Larry L. ; Kondrateva, Elizaveta S. ; Timofeyev, Maxim A. ; Yampolsky, Lev Y. ( May 2018 , Molecular Ecology)

   Local adaptation and phenotypic plasticity are main mechanisms of organisms’ resilience in changing environments. Both are affected by gene flow and are expected to be weak in zooplankton populations inhabiting large continuous water bodies and strongly affected by currents. Lake Baikal, the deepest and one of the coldest lakes on Earth, experienced epilimnion temperature increase during the last 100 years, exposing Baikal's zooplankton to novel selective pressures. We obtained a partial transcriptome ofEpischura baikalensis(Copepoda: Calanoida), the dominant component of Baikal's zooplankton, and estimatedSNPallele frequencies and transcript abundances in samples from regions of Baikal that differ in multiyear average surface temperatures. The strongest signal in bothSNPand transcript abundance differentiation is theSW‐NEgradient along the 600+ km long axis of the lake, suggesting isolation by distance.SNPdifferentiation is stronger for nonsynonymous than synonymousSNPs and is paralleled by differential survival during a laboratory exposure to increased temperature, indicating directional selection operating on the temperature gradient. Transcript abundance, generally collinear with theSNPdifferentiation, shows samples from the warmest, less deep location clustering together with the southernmost samples. Differential expression is more frequent among transcripts orthologous to candidate thermal response genes previously identified in model arthropods, including genes encoding cytoskeleton proteins, heat‐shock proteins, proteases, enzymes of central energy metabolism, lipid and antioxidant pathways. We conclude that the pivotal endemic zooplankton species in Lake Baikal exists under temperature‐mediated selection and possesses both genetic variation and plasticity to respond to novel temperature‐related environmental pressures.

    

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