...genetic experiments involving multiple generations can be completed in only a few days. * Propagation is simple, as the standard sexual morph is the self-fertilising hermaphrodite. Because of this ...mode of reproduction, issues of inbreeding depression (where inbreeding results in lowered reproductive fitness of lines because of homozygous deleterious mutations) are largely absent. The rest of the "pseudocoelomates" are now placed in the Lophotrochozoa 75,76, a group that includes Mollusca (snails and clams), Annelida (ragworms and earthworms), and Platyhelminthes (flatworms), amongst others. ...the worm is only one nematode of many, and nematodes are only one sort of worm.
Roche 454 pyrosequencing has become a method of choice for generating transcriptome data from non-model organisms. Once the tens to hundreds of thousands of short (250-450 base) reads have been ...produced, it is important to correctly assemble these to estimate the sequence of all the transcripts. Most transcriptome assembly projects use only one program for assembling 454 pyrosequencing reads, but there is no evidence that the programs used to date are optimal. We have carried out a systematic comparison of five assemblers (CAP3, MIRA, Newbler, SeqMan and CLC) to establish best practices for transcriptome assemblies, using a new dataset from the parasitic nematode Litomosoides sigmodontis.
Although no single assembler performed best on all our criteria, Newbler 2.5 gave longer contigs, better alignments to some reference sequences, and was fast and easy to use. SeqMan assemblies performed best on the criterion of recapitulating known transcripts, and had more novel sequence than the other assemblers, but generated an excess of small, redundant contigs. The remaining assemblers all performed almost as well, with the exception of Newbler 2.3 (the version currently used by most assembly projects), which generated assemblies that had significantly lower total length. As different assemblers use different underlying algorithms to generate contigs, we also explored merging of assemblies and found that the merged datasets not only aligned better to reference sequences than individual assemblies, but were also more consistent in the number and size of contigs.
Transcriptome assemblies are smaller than genome assemblies and thus should be more computationally tractable, but are often harder because individual contigs can have highly variable read coverage. Comparing single assemblers, Newbler 2.5 performed best on our trial data set, but other assemblers were closely comparable. Combining differently optimal assemblies from different programs however gave a more credible final product, and this strategy is recommended.
Revealing the Dark Matter of the Genome Blaxter, Mark
Science (American Association for the Advancement of Science),
12/2010, Letnik:
330, Številka:
6012
Journal Article
Recenzirano
Odprti dostop
Integrated data sets from two animal model organisms provide insights into the organization, structure, and function of their genomes.
Animal embryos successfully transform the two-dimensional code ...of their genome into multidimensional organisms that are ready to meet the challenge of natural selection. In addition to the three dimensions of the body, animal genomes inform additional dimensions: of cells coordinating to form tissues, tissues functioning together as organs, and organs shaping the body's systems; and of individuals responding appropriately to the varied challenges of life and surviving to breed. Poisons in food are detoxified, pathogens are killed, parasites are eliminated, and predators avoided through the deft employment of responses encoded in the genome. It is not currently possible to compute an organism from its genome, performing the transformation so efficiently executed by embryos, but two articles in this issue, by Gerstein
et al.
on page 1775 (
1
) and the modENCODE Consortium on page 1787 (
2
), bring this goal closer.
RADSeq: next-generation population genetics Davey, John W; Davey, John L; Blaxter, Mark L ...
Briefings in functional genomics,
12/2010, Letnik:
9, Številka:
5-6
Journal Article
Odprti dostop
Next-generation sequencing technologies are making a substantial impact on many areas of biology, including the analysis of genetic diversity in populations. However, genome-scale population genetic ...studies have been accessible only to well-funded model systems. Restriction-site associated DNA sequencing, a method that samples at reduced complexity across target genomes, promises to deliver high resolution population genomic data-thousands of sequenced markers across many individuals-for any organism at reasonable costs. It has found application in wild populations and non-traditional study species, and promises to become an important technology for ecological population genomics.
The Darwin Tree of Life (DToL) project aims to sequence all described terrestrial and aquatic eukaryotic species found in Britain and Ireland. Reference genome sequences are generated from single ...individuals for each target species. In addition to the target genome, sequenced samples often contain genetic material from microbiomes, endosymbionts, parasites, and other cobionts. Wolbachia endosymbiotic bacteria are found in a diversity of terrestrial arthropods and nematodes, with supergroups A and B the most common in insects. We identified and assembled 110 complete Wolbachia genomes from 93 host species spanning 92 families by filtering data from 368 insect species generated by the DToL project. From 15 infected species, we assembled more than one Wolbachia genome, including cases where individuals carried simultaneous supergroup A and B infections. Different insect orders had distinct patterns of infection, with Lepidopteran hosts mostly infected with supergroup B, while infections in Diptera and Hymenoptera were dominated by A-type Wolbachia. Other than these large-scale order-level associations, host and Wolbachia phylogenies revealed no (or very limited) cophylogeny. This points to the occurrence of frequent host switching events, including between insect orders, in the evolutionary history of the Wolbachia pandemic. While supergroup A and B genomes had distinct GC% and GC skew, and B genomes had a larger core gene set and tended to be longer, it was the abundance of copies of bacteriophage WO who was a strong determinant of Wolbachia genome size. Mining raw genome data generated for reference genome assemblies is a robust way of identifying and analysing cobiont genomes and giving greater ecological context for their hosts.
We present a genome assembly from an individual
(the spotted kaleidoscope jellyfish; Cnidaria; Staurozoa; Stauromedusae; Haliclystidae). The genome sequence is 262 megabases in span. Most of the ...assembly (98.3%) is scaffolded into nine (9) chromosomal pseudomolecules. The mitochondrial genome was also assembled and is 18.3 kilobases in length.
Reconstruction of target genomes from sequence data produced by instruments that are agnostic as to the species-of-origin may be confounded by contaminant DNA. Whether introduced during sample ...processing or through co-extraction alongside the target DNA, if insufficient care is taken during the assembly process, the final assembled genome may be a mixture of data from several species. Such assemblies can confound sequence-based biological inference and, when deposited in public databases, may be included in downstream analyses by users unaware of underlying problems. We present BlobToolKit, a software suite to aid researchers in identifying and isolating non-target data in draft and publicly available genome assemblies. BlobToolKit can be used to process assembly, read and analysis files for fully reproducible interactive exploration in the browser-based Viewer. BlobToolKit can be used during assembly to filter non-target DNA, helping researchers produce assemblies with high biological credibility. We have been running an automated BlobToolKit pipeline on eukaryotic assemblies publicly available in the International Nucleotide Sequence Data Collaboration and are making the results available through a public instance of the Viewer at https://blobtoolkit.genomehubs.org/view We aim to complete analysis of all publicly available genomes and then maintain currency with the flow of new genomes. We have worked to embed these views into the presentation of genome assemblies at the European Nucleotide Archive, providing an indication of assembly quality alongside the public record with links out to allow full exploration in the Viewer.
The promise of a DNA taxonomy Blaxter, Mark L.; Blaxter, Mark L.
Philosophical transactions of the Royal Society of London. Series B. Biological sciences,
04/2004, Letnik:
359, Številka:
1444
Journal Article
Recenzirano
Odprti dostop
Not only is the number of described species a very small proportion of the estimated extant number of taxa, but it also appears that all concepts of the extent and boundaries of 'species' fail in ...many cases. Using conserved molecular sequences it is possible to define and diagnose molecular operational taxonomic units (MOTU) that have a similar extent to traditional 'species'. Use of a MOTU system not only allows the rapid and effective identification of most taxa, including those not encountered before, but also allows investigation of the evolution of patterns of diversity. A MOTU approach is not without problems, particularly in the area of deciding what level of molecular difference defines a biologically relevant taxon, but has many benefits. Molecular data are extremely well suited to re-analysis and meta-analysis, and data from multiple independent studies can be readily collated and investigated by using new parameters and assumptions. Previous molecular taxonomic efforts have focused narrowly. Advances in high-throughput sequencing methodologies, however, place the idea of a universal, multi-locus molecular barcoding system in the realm of the possible.
Restriction-site associated DNA (RAD) sequencing is a powerful new method for targeted sequencing across the genomes of many individuals. This approach has broad potential for genetic analysis of ...non-model organisms including genotype-phenotype association mapping, phylogeography, population genetics and scaffolding genome assemblies through linkage mapping. We constructed a RAD library using genomic DNA from a Plutella xylostella (diamondback moth) backcross that segregated for resistance to the insecticide spinosad. Sequencing of 24 individuals was performed on a single Illumina GAIIx lane (51 base paired-end reads). Taking advantage of the lack of crossing over in homologous chromosomes in female Lepidoptera, 3,177 maternally inherited RAD alleles were assigned to the 31 chromosomes, enabling identification of the spinosad resistance and W/Z sex chromosomes. Paired-end reads for each RAD allele were assembled into contigs and compared to the genome of Bombyx mori (n = 28) using BLAST, revealing 28 homologous matches plus 3 expected fusion/breakage events which account for the difference in chromosome number. A genome-wide linkage map (1292 cM) was inferred with 2,878 segregating RAD alleles inherited from the backcross father, producing chromosome and location specific sequenced RAD markers. Here we have used RAD sequencing to construct a genetic linkage map de novo for an organism that has no previous genome data. Comparative analysis of P. xyloxtella linkage groups with B. mori chromosomes shows for the first time, genetic synteny appears common beyond the Macrolepidoptera. RAD sequencing is a powerful system capable of rapidly generating chromosome specific data for non-model organisms.
The goals of the Earth Biogenome Project-to sequence the genomes of all eukaryotic life on earth-are as daunting as they are ambitious. The Darwin Tree of Life Project was founded to demonstrate the ...credibility of these goals and to deliver at-scale genome sequences of unprecedented quality for a biogeographic region: the archipelago of islands that constitute Britain and Ireland. The Darwin Tree of Life Project is a collaboration between biodiversity organizations (museums, botanical gardens, and biodiversity institutes) and genomics institutes. Together, we have built a workflow that collects specimens from the field, robustly identifies them, performs sequencing, generates high-quality, curated assemblies, and releases these openly for the global community to use to build future science and conservation efforts.
PDF
