Identification Of Structural Variants Research Articles

Structural variation analysis with strobe reads

Structural variation including deletions, duplications and rearrangements of DNA sequence are an important contributor to genome variation in many organisms. In human, many structural variants are found in complex and highly repetitive regions of the genome making their identification difficult. A new sequencing technology called strobe sequencing generates strobe reads containing multiple subreads from a single contiguous fragment of DNA. Strobe reads thus generalize the concept of paired reads, or mate pairs, that have been routinely used for structural variant detection. Strobe sequencing holds promise for unraveling complex variants that have been difficult to characterize with current sequencing technologies. We introduce an algorithm for identification of structural variants using strobe sequencing data. We consider strobe reads from a test genome that have multiple possible alignments to a reference genome due to sequencing errors and/or repetitive sequences in the reference. We formulate the combinatorial optimization problem of finding the minimum number of structural variants in the test genome that are consistent with these alignments. We solve this problem using an integer linear program. Using simulated strobe sequencing data, we show that our algorithm has better sensitivity and specificity than paired read approaches for structural variation identification. braphael@brown.edu

A geometric approach for classification and comparison of structural variants

Motivation: Structural variants, including duplications, insertions, deletions and inversions of large blocks of DNA sequence, are an important contributor to human genome variation. Measuring structural variants in a genome sequence is typically more challenging than measuring single nucleotide changes. Current approaches for structural variant identification, including paired-end DNA sequencing/mapping and array comparative genomic hybridization (aCGH), do not identify the boundaries of variants precisely. Consequently, most reported human structural variants are poorly defined and not readily compared across different studies and measurement techniques.Results: We introduce Geometric Analysis of Structural Variants (GASV), a geometric approach for identification, classification and comparison of structural variants. This approach represents the uncertainty in measurement of a structural variant as a polygon in the plane, and identifies measurements supporting the same variant by computing intersections of polygons. We derive a computational geometry algorithm to efficiently identify all such intersections. We apply GASV to sequencing data from nine individual human genomes and several cancer genomes. We obtain better localization of the boundaries of structural variants, distinguish genetic from putative somatic structural variants in cancer genomes, and integrate aCGH and paired-end sequencing measurements of structural variants. This work presents the first general framework for comparing structural variants across multiple samples and measurement techniques, and will be useful for studies of both genetic structural variants and somatic rearrangements in cancer.Availability: http://cs.brown.edu/people/braphael/software.htmlContact: braphael@brown.edu

The diploid genome sequence of an Asian individual

Here we present the first diploid genome sequence of an Asian individual. The genome was sequenced to 36-fold average coverage using massively parallel sequencing technology. We aligned the short reads onto the NCBI human reference genome to 99.97% coverage, and guided by the reference genome, we used uniquely mapped reads to assemble a high-quality consensus sequence for 92% of the Asian individual's genome. We identified approximately 3 million single-nucleotide polymorphisms (SNPs) inside this region, of which 13.6% were not in the dbSNP database. Genotyping analysis showed that SNP identification had high accuracy and consistency, indicating the high sequence quality of this assembly. We also carried out heterozygote phasing and haplotype prediction against HapMap CHB and JPT haplotypes (Chinese and Japanese, respectively), sequence comparison with the two available individual genomes (J. D. Watson and J. C. Venter), and structural variation identification. These variations were considered for their potential biological impact. Our sequence data and analyses demonstrate the potential usefulness of next-generation sequencing technologies for personal genomics.

A robust framework for detecting structural variations in a genome

Motivation: Recently, structural genomic variants have come to the forefront as a significant source of variation in the human population, but the identification of these variants in a large genome remains a challenge. The complete sequencing of a human individual is prohibitive at current costs, while current polymorphism detection technologies, such as SNP arrays, are not able to identify many of the large scale events. One of the most promising methods to detect such variants is the computational mapping of clone-end sequences to a reference genome.Results: Here, we present a probabilistic framework for the identification of structural variants using clone-end sequencing. Unlike previous methods, our approach does not rely on an a priori determined mapping of all reads to the reference. Instead, we build a framework for finding the most probable assignment of sequenced clones to potential structural variants based on the other clones. We compare our predictions with the structural variants identified in three previous studies. While there is a statistically significant correlation between the predictions, we also find a significant number of previously uncharacterized structural variants. Furthermore, we identify a number of putative cross-chromosomal events, primarily located proximally to the centromeres of the chromosomes.Availability: Our dataset, results and source code are available at http://compbio.cs.toronto.edu/structvar/Contact:seunghak@cs.toronto.edu,echeran@cs.toronto.edu,brudno@cs.toronto.edu

Multiplex PCR strategy for rapid identification of structural types and variants of the mec element in methicillin-resistant Staphylococcus aureus.

Full characterization of methicillin-resistant Staphylococcus aureus (MRSA) requires definition of not only the bacterial genetic background but also the structure of the complex and heterologous mec element these bacteria carry, which is associated with drug resistance determinant mecA. We report the development, validation, and application of a multiplex PCR strategy that allows quick presumptive characterization of the mec element types based on the structural features that were shown to be typical of mec elements carried by several MRSA clones. The strategy was validated by using a representative collection of pandemic MRSA clones in which the full structure of the associated mec elements was previously determined by hybridization and PCR screenings and also by DNA sequencing. The method was tested together with multilocus sequence typing and other typing methods for the characterization of 18 isolates representative of the MRSA clones recovered during a hospital outbreak in Barcelona, Spain. The multiplex PCR was shown to be rapid, robust, and capable in a single assay of identifying five structural types of the mec element among these strains, three major and two minor variants, each one of which has been already been seen among MRSA characterized earlier. This technique should be a useful addition to the armamentarium of molecular typing tools for the characterization of MRSA clonal types and for the rapid tentative identification of structural variants of the mec element.

Cell type-specific integrin variants with alternative alpha chain cytoplasmic domains.

The integrin heterodimers composed of the alpha 6 subunit with the beta 1 or beta 4 subunit (alpha 6 beta 1 and alpha 6 beta 4) are receptors for laminin and basement membrane components, respectively. The alpha 3 beta 1 integrin recognizes laminin, collagen, fibronectin, or epiligrin. We report the identification of structural variants (A and B) of the alpha 6 and alpha 3 subunits, containing distinct cytoplasmic domains. The expression of one cytoplasmic domain or the other, based probably on alternative exon usage, is cell-type dependent. Most transformed cell lines express both alpha 6A and alpha 6B isoforms, as determined by mRNA amplification or antibody immunoprecipitation. In contrast, embryonic fibroblasts express exclusively alpha 6A, and embryonic stem cells express exclusively alpha 6B. In most normal tissues, both alpha 6 isoforms are detectable, but several tissues express either alpha 6A or alpha 6B. The alpha 3B mRNA was amplified from heart and brain, while all other tissues and cell lines tested contained only alpha 3A mRNA. Alternative cytoplasmic domains may provide a means for varying the cellular responses to the ligands of alpha 6 and alpha 3 integrins according to the cell type.

10] Isolation and characterization of apolipoproteins A-I, A-II, and A-IV

A number of different analytical techniques are now available for the isolation of apoA-I, apoA-II, and apoA-IV. The choice of a particular technique is dependent on the instrumentation available, and the quantity of isolated apolipoprotein required. The isolation and characterization of the separate isoforms and the precursor isoproteins of the individual apolipoproteins are detailed, and methods for the evaluation of the purity of the separate apolipoproteins presented. A method for the evaluation of apolipoproteins in plasma is now available which permits the identification of structural variants of plasma apolipoproteins in patients with dyslipoproteinemias.