Transcriptome sequencing in an ecologically important tree species: assembly, annotation, and marker discovery

<p>Abstract</p> <p>Background</p> <p>Massively parallel sequencing of cDNA is now an efficient route for generating enormous sequence collections that represent expressed genes. This approach provides a valuable starting point for characterizing functional genetic variation in non-model organisms, especially where whole genome sequencing efforts are currently cost and time prohibitive. The large and complex genomes of pines (<it>Pinus </it>spp.) have hindered the development of genomic resources, despite the ecological and economical importance of the group. While most genomic studies have focused on a single species (<it>P. taeda</it>), genomic level resources for other pines are insufficiently developed to facilitate ecological genomic research. Lodgepole pine (<it>P. contorta</it>) is an ecologically important foundation species of montane forest ecosystems and exhibits substantial adaptive variation across its range in western North America. Here we describe a sequencing study of expressed genes from <it>P. contorta</it>, including their assembly and annotation, and their potential for molecular marker development to support population and association genetic studies.</p> <p>Results</p> <p>We obtained 586,732 sequencing reads from a 454 GS XLR70 Titanium pyrosequencer (mean length: 306 base pairs). A combination of reference-based and <it>de novo </it>assemblies yielded 63,657 contigs, with 239,793 reads remaining as singletons. Based on sequence similarity with known proteins, these sequences represent approximately 17,000 unique genes, many of which are well covered by contig sequences. This sequence collection also included a surprisingly large number of retrotransposon sequences, suggesting that they are highly transcriptionally active in the tissues we sampled. We located and characterized thousands of simple sequence repeats and single nucleotide polymorphisms as potential molecular markers in our assembled and annotated sequences. High quality PCR primers were designed for a substantial number of the SSR loci, and a large number of these were amplified successfully in initial screening.</p> <p>Conclusions</p> <p>This sequence collection represents a major genomic resource for <it>P. contorta</it>, and the large number of genetic markers characterized should contribute to future research in this and other pines. Our results illustrate the utility of next generation sequencing as a basis for marker development and population genomics in non-model species.</p

Toward a General Model for the Evolutionary Dynamics of Gene Duplicates

Gene duplication is an important process in the functional divergence of genes and genomes. Several processes have been described that lead to duplicate gene retention over different timescales after both smaller-scale events and whole-genome duplication, including neofunctionalization, subfunctionalization, and dosage balance. Two common modes of duplicate gene loss include nonfunctionalization and loss due to population dynamics (failed fixation). Previous work has characterized expectations of duplicate gene retention under the neofunctionalization and subfunctionalization models. Here, that work is extended to dosage balance using simulations. A general model for duplicate gene loss/retention is then presented that is capable of fitting expectations under the different models, is defined at t = 0, and decays to an orthologous asymptotic rate rather than zero, based upon a modified Weibull hazard function. The model in a maximum likelihood framework shows the property of identifiability, recovering the evolutionary mechanism and parameters of simulation. This model is also capable of recovering the evolutionary mechanism of simulation from data generated using an unrelated network population genetic model. Lastly, the general model is applied as part of a mixture model to recent gene duplicates from the Oikopleura dioica genome, suggesting that neofunctionalization may be an important process leading to duplicate gene retention in that organism

The Evolution of Protein Structures and Structural Ensembles Under Functional Constraint

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The interface of protein structure, protein biophysics, and molecular evolution

The interface of protein structural biology, protein biophysics, molecular evolution, and molecular population genetics forms the foundations for a mechanistic understanding of many aspects of protein biochemistry. Current efforts in interdisciplinary protein modeling are in their infancy and the state-of-the art of such models is described. Beyond the relationship between amino acid substitution and static protein structure, protein function, and corresponding organismal fitness, other considerations are also discussed. More complex mutational processes such as insertion and deletion and domain rearrangements and even circular permutations should be evaluated. The role of intrinsically disordered proteins is still controversial, but may be increasingly important to consider. Protein geometry and protein dynamics as a deviation from static considerations of protein structure are also important. Protein expression level is known to be a major determinant of evolutionary rate and several considerations including selection at the mRNA level and the role of interaction specificity are discussed. Lastly, the relationship between modeling and needed high-throughput experimental data as well as experimental examination of protein evolution using ancestral sequence resurrection and in vitro biochemistry are presented, towards an aim of ultimately generating better models for biological inference and prediction