De novo yeast genome assemblies from MinION, PacBio and MiSeq platforms.

Long-read sequencing technologies such as Pacific Biosciences and Oxford Nanopore MinION are capable of producing long sequencing reads with average fragment lengths of over 10,000 base-pairs and maximum lengths reaching 100,000 base- pairs. Compared with short reads, the assemblies obtained from long-read sequencing platforms have much higher contig continuity and genome completeness as long fragments are able to extend paths into problematic or repetitive regions. Many successful assembly applications of the Pacific Biosciences technology have been reported ranging from small bacterial genomes to large plant and animal genomes. Recently, genome assemblies using Oxford Nanopore MinION data have attracted much attention due to the portability and low cost of this novel sequencing instrument. In this paper, we re-sequenced a well characterized genome, the Saccharomyces cerevisiae S288C strain using three different platforms: MinION, PacBio and MiSeq. We present a comprehensive metric comparison of assemblies generated by various pipelines and discuss how the platform associated data characteristics affect the assembly quality. With a given read depth of 31X, the assemblies from both Pacific Biosciences and Oxford Nanopore MinION show excellent continuity and completeness for the 16 nuclear chromosomes, but not for the mitochondrial genome, whose reconstruction still represents a significant challenge.

Understanding the Genomic Structure of Copy-Number Variation of the Low-Affinity Fcγ Receptor Region Allows Confirmation of the Association of FCGR3B Deletion with Rheumatoid Arthritis.

Fcγ receptors are a family of cell-surface receptors that are expressed by a host of different innate and adaptive immune cells, and mediate inflammatory responses by binding the Fc portion of immunoglobulin G. In humans, five low-affinity receptors are encoded by the genes FCGR2A, FCGR2B, FCGR2C, FCGR3A, and FCGR3B, which are located in an 82.5-kb segmental tandem duplication on chromosome 1q23.3, which shows extensive copy-number variation (CNV). Deletions of FCGR3B have been suggested to increase the risk of inflammatory diseases such as systemic lupus erythematosus and rheumatoid arthritis (RA). In this study, we identify the deletion breakpoints of FCGR3B deletion alleles in the UK population and endogamous native American population, and show that some but not all alleles are likely to be identical-by-descent. We also localize a duplication breakpoint, confirming that the mechanism of CNV generation is nonallelic homologous recombination, and identify several alleles with gene conversion events using fosmid sequencing data. We use information on the structure of the deletion alleles to distinguish FCGR3B deletions from FCGR3A deletions in whole-genome array comparative genomic hybridization (aCGH) data. Reanalysis of published aCGH data using this approach supports association of FCGR3B deletion with increased risk of RA in a large cohort of 1,982 cases and 3,271 controls (odds ratio 1.61, P = 2.9×10-3 ).

Development and validation of a comprehensive genomic diagnostic tool for myeloid malignancies.

The diagnosis of hematologic malignancies relies on multidisciplinary workflows involving morphology, flow cytometry, cytogenetic, and molecular genetic analyses. Advances in cancer genomics have identified numerous recurrent mutations with clear prognostic and/or therapeutic significance to different cancers. In myeloid malignancies, there is a clinical imperative to test for such mutations in mainstream diagnosis; however, progress toward this has been slow and piecemeal. Here we describe Karyogene, an integrated targeted resequencing/analytical platform that detects nucleotide substitutions, insertions/deletions, chromosomal translocations, copy number abnormalities, and zygosity changes in a single assay. We validate the approach against 62 acute myeloid leukemia, 50 myelodysplastic syndrome, and 40 blood DNA samples from individuals without evidence of clonal blood disorders. We demonstrate robust detection of sequence changes in 49 genes, including difficult-to-detect mutations such as FLT3 internal-tandem and mixed-lineage leukemia (MLL) partial-tandem duplications, and clinically significant chromosomal rearrangements including MLL translocations to known and unknown partners, identifying the novel fusion gene MLL-DIAPH2 in the process. Additionally, we identify most significant chromosomal gains and losses, and several copy neutral loss-of-heterozygosity mutations at a genome-wide level, including previously unreported changes such as homozygosity for DNMT3A R882 mutations. Karyogene represents a dependable genomic diagnosis platform for translational research and for the clinical management of myeloid malignancies, which can be readily adapted for use in other cancers.

GapMis: a tool for pairwise sequence alignment with a single gap.

MOTIVATION: Pairwise sequence alignment has received a new motivation due to the advent of recent patents in next-generation sequencing technologies, particularly so for the application of re-sequencing---the assembly of a genome directed by a reference sequence. After the fast alignment between a factor of the reference sequence and a high-quality fragment of a short read by a short-read alignment programme, an important problem is to find the alignment between a relatively short succeeding factor of the reference sequence and the remaining low-quality part of the read allowing a number of mismatches and the insertion of a single gap in the alignment. RESULTS: We present GapMis, a tool for pairwise sequence alignment with a single gap. It is based on a simple algorithm, which computes a different version of the traditional dynamic programming matrix. The presented experimental results demonstrate that GapMis is more suitable and efficient than most popular tools for this task.

Transcriptome map of mouse isochores in embryonic and neonatal cortex.

Several studies on adult tissues agree on the presence of a positive effect of the genomic and genic base composition on mammalian gene expression. Recent literature supports the idea that during developmental processes GC-poor genomic regions are preferentially implicated. We investigate the relationship between the compositional properties of the isochores and of the genes with their respective expression activity during developmental processes. Using RNA-seq data from two distinct developmental stages of the mouse cortex, embryonic day 18 (E18) and postnatal day 7 (P7), we established for the first time a developmental-related transcriptome map of the mouse isochores. Additionally, for each stage we estimated the correlation between isochores' GC level and their expression activity, and the genes' expression patterns for each isochore family. Our analyses add evidence supporting the idea that during development GC-poor isochores are preferentially implicated, and confirm the positive effect of genes' GC level on their expression activity.

Next-generation sequencing and large genome assemblies.

The next-generation sequencing (NGS) revolution has drastically reduced time and cost requirements for sequencing of large genomes, and also qualitatively changed the problem of assembly. This article reviews the state of the art in de novo genome assembly, paying particular attention to mammalian-sized genomes. The strengths and weaknesses of the main sequencing platforms are highlighted, leading to a discussion of assembly and the new challenges associated with NGS data. Current approaches to assembly are outlined and the various software packages available are introduced and compared. The question of whether quality assemblies can be produced using short-read NGS data alone, or whether it must be combined with more expensive sequencing techniques, is considered. Prospects for future assemblers and tests of assembly performance are also discussed.

Transcriptome map of mouse isochores.

BACKGROUND: The availability of fully sequenced genomes and the implementation of transcriptome technologies have increased the studies investigating the expression profiles for a variety of tissues, conditions, and species. In this study, using RNA-seq data for three distinct tissues (brain, liver, and muscle), we investigate how base composition affects mammalian gene expression, an issue of prime practical and evolutionary interest. RESULTS: We present the transcriptome map of the mouse isochores (DNA segments with a fairly homogeneous base composition) for the three different tissues and the effects of isochores' base composition on their expression activity. Our analyses also cover the relations between the genes' expression activity and their localization in the isochore families. CONCLUSIONS: This study is the first where next-generation sequencing data are used to associate the effects of both genomic and genic compositional properties to their corresponding expression activity. Our findings confirm previous results, and further support the existence of a relationship between isochores and gene expression. This relationship corroborates that isochores are primarily a product of evolutionary adaptation rather than a simple by-product of neutral evolutionary processes.

Integrating color constancy into JPEG2000.

The human visual system is able to perceive colors as approximately constant. This ability is known as color constancy. In contrast, the colors measured by a sensor vary with the type of illuminant used. Color constancy is very important for digital photography and automatic color-based object recognition. In digital photography, this ability is known under the name automatic white balance. A number of algorithms have been developed for color constancy. We review two well-known color constancy algorithms, the gray world assumption and the Retinex algorithm and show how a color constancy algorithm may be integrated into the JPEG2000 framework. Since computer images are usually stored in compressed form anyway, little overhead is required to add color constancy into the processing pipeline.