349 research outputs found

    Enhancing Estimates of Breakpoints in Genome Copy Number Alteration using Confidence Masks

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    Chromosomal structural changes in human body known as copy number alteration (CNA) are often associated with diseases, such as various forms of cancer. Therefore, accurate estimation of breakpoints of the CNAs is important to understand the genetic basis of many diseases. The high‐resolution comparative genomic hybridization (HR‐CGH) and single‐nucleotide polymorphism (SNP) technologies enable cost‐efficient and high‐throughput CNA detection. However, probing provided using these profiles gives data highly contaminated by intensive Gaussian noise having white properties. We observe the probabilistic properties of CNA in HR‐CGH and SNP measurements and show that jitter in the breakpoints can statistically be described with either the discrete skew Laplace distribution when the segmental signal‐to‐noise ratio (SNR) exceeds unity or modified Bessel function‐based approximation when SNR is <1. Based upon these approaches, the confidence masks can be developed and used to enhance the estimates of the CNAs for the given confidence probability by removing some unlikely existing breakpoints

    Gene duplications during experimental evolution of Caenorhabditis elegans : duplication rates and evolutionary responses

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    Copy-number variants (CNVs) are a ubiquitous form of genetic variation. How often this form of variation arises and its adaptive significance are active areas of contemporary research. This work presents evidence regarding both of these subjects. First, it demonstrates that gene duplications occur at a frequency two orders of magnitude greater than point mutations. Specifically, the gene duplication rate is estimated to be 1.2 x 10-7/gene/generation, compared to a point mutation rate on the order of ~10-9/site/ generation. Second, it was found that populations in a low state of fitness due to mutation accumulation could recover some or all of their fitness over short spans of generations concurrent with an increase in frequency of duplications and deletions that arose during the recovery process. The pattern of frequency increase among CNVs over generations during recovery was consistent with the signature of positive selection. The median size of duplications that were identified after selection for ~200 generations were significantly larger (191.5 kb) than both duplications that occurred spontaneously (2 kb) in the absence of selection and deletions identified after selection for ~200 generations (12.5 kb). The median number of genes contained in the duplications during recovery was 38, evincing the ability of these events to increase the genetic information available for selection to act on. These results clearly demonstrate that gene duplication and deletion processes contribute significantly to the adaptability of populations

    The Baker's Yeast Diploid Genome Is Remarkably Stable in Vegetative Growth and Meiosis

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    Accurate estimates of mutation rates provide critical information to analyze genome evolution and organism fitness. We used whole-genome DNA sequencing, pulse-field gel electrophoresis, and comparative genome hybridization to determine mutation rates in diploid vegetative and meiotic mutation accumulation lines of Saccharomyces cerevisiae. The vegetative lines underwent only mitotic divisions while the meiotic lines underwent a meiotic cycle every ∌20 vegetative divisions. Similar base substitution rates were estimated for both lines. Given our experimental design, these measures indicated that the meiotic mutation rate is within the range of being equal to zero to being 55-fold higher than the vegetative rate. Mutations detected in vegetative lines were all heterozygous while those in meiotic lines were homozygous. A quantitative analysis of intra-tetrad mating events in the meiotic lines showed that inter-spore mating is primarily responsible for rapidly fixing mutations to homozygosity as well as for removing mutations. We did not observe 1–2 nt insertion/deletion (in-del) mutations in any of the sequenced lines and only one structural variant in a non-telomeric location was found. However, a large number of structural variations in subtelomeric sequences were seen in both vegetative and meiotic lines that did not affect viability. Our results indicate that the diploid yeast nuclear genome is remarkably stable during the vegetative and meiotic cell cycles and support the hypothesis that peripheral regions of chromosomes are more dynamic than gene-rich central sections where structural rearrangements could be deleterious. This work also provides an improved estimate for the mutational load carried by diploid organisms

    Genetic analysis of squamous cell carcinoma of the head and neck

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    Head and neck squamous cell carcinoma (HNSCC) is the sixth commonest cancer worldwide with an increasing incidence in developing countries. Despite numerous advances in surgery, radiotherapy and chemotherapy over the past few decades the overall survival rate for patients with HNSCC has changed little. Currently, the management of HNSCC patients is based on the assessment of a variety of clinical and pathological parameters. However, in many instances, these factors fail to accurately predict the clinical behaviour of an individual patients tumour. HNSCC therefore, is a tumour entity that would benefit from a greater insight into the chromosomal alterations underlying the disease. Knowledge of such alterations would be expected to provide many benefits to the HNSCC clinician in terms of diagnostic and prognostic markers and may eventually identify novel molecular targets for therapeutic intervention.This thesis was aimed at characterising the chromosomal abnormalities involved in the tumourigenesis of HNSCC, principally using the powerful molecular cytogenetic technique of comparative genomic hybridisation (CGH), and the clinical applications of such data. Firstly, the technique was optimised and initially applied to specimens of primary HNSCC and surrounding uninvolved mucosa from 19 patients in order to investigate the phenomenon of 'field cancerization'. Specimens of primary HNSCC and histologically normal mucosa taken from 1cm and 5cm distant to the primary site were analysed from each patient in order to characterise the chromosomal abnormalities associated with malignant tissue and attempt to identify aberrations underlying the `field change'. CGH of the primary tumour specimens revealed numerous chromosomal aberrations with a relatively consistent pattern. Frequent deletions of DNA were identified on chromosome 3p, 4p, 8p, 9p, 11 q, 13q and 18q and frequent gains on chromosomes 2q, 3q, 5p, 7q, 8q, 9q and 11q. The histologically normal mucosa did not show chromosomal abnormalities within the cells analysed. Therefore, if molecular abnormalities were present in the mucosa surrounding a primary HNSCC they would be below the resolution of CGH, such as subtle point mutations, or only present in a minority of cells.In order to investigate the genetic relationship between primary HNSCC and lymph node metastases, matched pairs of primary and metastatic tumours were obtained from 18 patients and analysed by CGH. Whilst the overall frequency of genetic alterations was similar between primary and metastatic tumours, a surprising degree of discordance was identified between each individual's matched pair of tumours. At least one common aberration was identified in all cases studied, however the percentage of aberrations detected in the lymph node metastases that were shared with the primary tumour varied greatly, ranging from 100% - 8.3%. Several chromosomal regions were found to be altered at similar frequencies in both the primary and metastatic tumours. Most interestingly, regions of the genome found to be altered at a higher frequency in the population of metastatic HNSCC included deletion of 4p15.3-pter and 17q22-qter and gain of 6gcen-q15 and 13q21-22. In addition, both gains and deletions of material from chromosome 22 were found at a higher frequency in the metastatic tumours. These chromosomal regions may contain genes important in the process of metastasis in HNSCC. The level of discordance identified between matched pairs of tumours also suggests that a linear progression model may not satisfactorily explain the progression to metastases in all HNSCC.This thesis also addressed the important clinical question of resistance to radiotherapy demonstrated by a significant fraction of laryngeal tumours. No markers that reliable predict the response of an individual tumour to radiotherapy are currently available. The expression of a panel of markers involved in DNA damage recognition, cell cycle arrest, DNA repair and apoptosis were evaluated in 23 glottic laryngeal tumours (8 radio-resistant and 13 radio-sensitive). Of these, the expression of bcl-2, an anti-apoptotic marker, was specifically associated with the resistant phenotype. This statistically significant association provides preliminary evidence for the dysregulation of apoptosis as a mechanism by which resistant tumours can evade radiotherapy induced tumour regression.Overall, CGH analysis of primary HNSCC identified a relatively consistent pattern of DNA alterations with several distinct regions of DNA deletion and gain identified. Frequent deletions of DNA were identified on chromosomes 1p, 2q, 3p, 4p, 4q, 5q, 7q, 8p, 9q, 10q, 11p, 11q, 13q, 17p, 18q, 19 and 21 and frequent gains of DNA on chromosomes 1q, 2q, 3q, 4q, 5p, 6q, 7p, 7q, 8q, llq, 12p, 13q, 18p and 18q. Chromosome 3 was the most frequent site of both deletions and gains. Follow up data was obtained for all patients analysed by CGH and Kaplan-Meier survival analysis demonstrated a significant correlation between gain of DNA on 3q25-27 and reduced overall survival. This finding highlights the necessity for further, high resolution, characterisation of this region in order that thespecific genetic marker can be identified. This thesis demonstrates that molecular analysis of tumours is able to offer new, and valuable information for the understanding of HNSCC carcinogenesis and that these data can be used to compliment existing methodology. Further work is required to isolate specific genes and to understand their interactions
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