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A whole-genome array comparative genomic hybridization assay is used to identify and analyze segmental copy-number variations (CNVs) in the human genome that represent a major genetic component of the phenotypic diversity. The in-depth survey provides a valuable baseline for studies involving human genetics.

Abstract Background Recent advances in global genomic profiling methodologies have enabled multi-dimensional characterization of biological systems. Complete analysis of these genomic profiles require an in depth look at parallel profiles of segmental DNA copy number status, DNA methylation state, single nucleotide polymorphisms, as well as gene expression profiles. Due to the differences in data types it is difficult to conduct parallel analysis of multiple datasets from diverse platforms. Results To address this issue, we have developed an integrative genomic analysis platform MD-SeeGH, a software tool that allows users to rapidly and directly analyze genomic datasets spanning multiple genomic experiments. With MD-SeeGH, users have the flexibility to easily update datasets in accordance with new genomic builds, make a quality assessment of data using the filtering features, and identify genetic alterations within single or across multiple experiments. Multiple sample analysis in MD-SeeGH allows users to compare profiles from many experiments alongside tracks containing detailed localized gene information, microRNA, CpG islands, and copy number variations. Conclusion MD-SeeGH is a new platform for the integrative analysis of diverse microarray data, facilitating multiple profile analyses and group comparisons.

Abstract Background Array comparative genomic hybridization (CGH) is a technique which detects copy number differences in DNA segments. Complete sequencing of the human genome and the development of an array representing a tiling set of tens of thousands of DNA segments spanning the entire human genome has made high resolution copy number analysis throughout the genome possible. Since array CGH provides signal ratio for each DNA segment, visualization would require the reassembly of individual data points into chromosome profiles. Results We have developed a visualization tool for displaying whole genome array CGH data in the context of chromosomal location. SeeGH is an application that translates spot signal ratio data from array CGH experiments to displays of high resolution chromosome profiles. Data is imported from a simple tab delimited text file obtained from standard microarray image analysis software. SeeGH processes the signal ratio data and graphically displays it in a conventional CGH karyotype diagram with the added features of magnification and DNA segment annotation. In this process, SeeGH imports the data into a database, calculates the average ratio and standard deviation for each replicate spot, and links them to chromosome regions for graphical display. Once the data is displayed, users have the option of hiding or flagging DNA segments based on user defined criteria, and retrieve annotation information such as clone name, NCBI sequence accession number, ratio, base pair position on the chromosome, and standard deviation. Conclusions SeeGH represents a novel software tool used to view and analyze array CGH data. The software gives users the ability to view the data in an overall genomic view as well as magnify specific chromosomal regions facilitating the precise localization of genetic alterations. SeeGH is easily installed and runs on Microsoft Windows 2000 or later environments.

Abstract Background The recent development of array based comparative genomic hybridization (CGH) technology provides improved resolution for detection of genomic DNA copy number alterations. In array CGH, generating spotting solution is a multi-step process where bacterial artificial chromosome (BAC) clones are converted to replenishable PCR amplified fragments pools (AFP) for use as spotting solution in a microarray format on glass substrate. With completion of the human and mouse genome sequencing, large BAC clone sets providing complete genome coverage are available for construction of whole genome BAC arrays. Currently, Southern hybridization, fluorescent in-situ hybridization (FISH), and BAC end sequencing methods are commonly used to identify the initial BAC clone but not the end product used for spotting arrays. The AFP sequencing technique described in this study is a novel method designed to verify the identity of array spotting solution in a high throughput manner. Results We show here that Southern hybridization, FISH, and AFP sequencing can be used to verify the identity of final spotting solutions using less than 10% of the AFP product. Single pass AFP sequencing identified over half of the 960 AFPs analyzed. Moreover, using two vector primers approximately 90% of the AFP spotting solutions can be identified. Conclusions In this feasibility study we demonstrate that current methods for identifying initial BAC clones can be adapted to verify the identity of AFP spotting solutions used in printing arrays. Of these methods, AFP sequencing proves to be the most efficient for large scale identification of spotting solution in a high throughput manner.

Rituximab binds an epitope on the CD20 antigen, encompassed in exon 5 of the MS4A1 gene. We sequenced this region and correlated the presence of mutations with CD20 protein expression and response to R-CHOP in patients with diffuse large B-cell lymphoma: 264 diagnostic biopsies and 15 biopsies taken at the time of relapse were successfully sequenced. CD20 mutations involving the rituximab epitope were detected in only 1/264 (0.4%) and 1/15 (6%) of the biopsies taken at diagnosis and relapse, respectively. No polymorphic sequence variants were detected in this region. Three patients had malignant cells that were CD20 protein-positive at diagnosis but CD20-negative at relapse. Thus, CD20 mutations involving the rituximab epitope are rare in both de novo and relapsed diffuse large B-cell lymphoma, and do not represent a significant cause of R-CHOP resistance. CD20 protein-negative relapses occur after R-CHOP therapy but their clinical relevance is unknown.

Pheochromocytomas (PCC) are relatively rare neuroendocrine tumors, mainly of the adrenal medulla. They arise sporadically or occur secondary to inherited cancer syndromes, such as multiple endocrine neoplasia type II (MEN2), von Hippel-Lindau disease (VHL), or neurofibromatosis type I (NF1). Loss of 1p is the most frequently encountered genetic alteration, especially in MEN2-related and sporadic PCC. Previous studies have revealed three regions of common somatic loss on chromosome arm 1p, using chromosome-based comparative genomic hybridization (CGH) and LOH analysis. To investigate these chromosomal aberrations with a higher resolution and sensitivity, we performed microarray-based CGH with 13 sporadic and 11 syndrome-related (10 MEN2A-related and 1 NF1-related) tumors. The array consisted of 642 overlapping bacterial artificial chromosome (BAC) clones mapped to 1p11.2-p36.33. Chromosomal deletions on 1p were detected in 18 of 24 cases (75%). Among 9 tumors with partial 1p loss, the deleted region was restricted to 1cen-1p32.3 in six cases (25%), indicating a region of genetic instability. The consensus regions of deletion in this study involved 1cen-1p21.1, 1p21.3-1p31.3, and 1p34.3-1p36.33. In conclusion, these data strongly suggest that chromosome arm 1p is the site for multiple tumor suppressor genes, although the potential candidate genes CDKN2C and PTPRF/LAR are not included in these regions., (Copyright 2005 Wiley-Liss, Inc)