Project description:Array-based comparative genomic hybridisation is a high-resolution method for measuring chromosomal copy number changes. Here we present a validated protocol using in-house spotted oligonucleotide libraries for array CGH. This oligo array CGH platform yields reproducible results and is capable of detecting single copy gains, multi-copy amplifications as well as homozygous and heterozygous deletions as small as 100 kb with high resolution. A human oligonucleotide library was printed on amine binding slides. Arrays were hybridised using a hybstation and analysed using BleuFuse feature extraction software, with over 95% of spots passing quality control. The protocol allows as little as 300 ng of input DNA without the need for amplification or target reduction and a 90% reduction of Cot1-DNA without compromising quality. High quality results have also been obtained with DNA from archival tissue. Finally, in addition to human oligo arrays, we have applied the protocol successfully to mouse oligo arrays. We believe that this oligo-based platform using “off-the-shelf” oligo-libraries provides an easy accessible alternative to BAC arrays for CGH, which is cost-effective, available at high resolution and easily implemented for any sequenced organism without compromising the quality of the results. Keywords: comparative genomic hybridization, oligonucleotide,
Project description:In this report, we have developed a rapid oligonucleotide microarray detection technique to identify the most common ten Legionella spp.. The sensitivity of the detection was at 1.0 ng with genomic DNA or 13 CFU/100 mL with Legionella cultures. The microarray detected seven air conditioner-condensed water samples with 100% accuracy, validating the technique as a promising method for applications in basic microbiology, clinical diagnosis, food safety, and epidemiological surveillance. The phylogenetic study based on the ITS has also revealed interestingly that the non-pathogenic L. fairfieldensis is the closest to L. pneumophila than the nine other pathogenic Legionella spp..
Project description:Both spotted long oligonucleotide arrays and Affymetrix GeneChips were used to measure differential gene expression in two RNA samples (K562 erythroleukemia RNA and the Stratagene Universal Human Reference RNA). For Affymetrix technology, the two RNAs were analyzed separately. There are two replicates for K562 RNA (GSM4843 and GSM4844) and three for the Stratagene Universal reference (GSM4845-GSM4847). Two types of spotted long oligonucleotide arrays were used. These arrays were used to do two-color hybridizations to directly compare K562 and Universal reference RNAs. One array (GPL273) was made with probes from the Operon Human Genome Oligo Set Version 1. These arrays were used with unamplified cDNA probes (6 replicates, GSM4848-GSM4853), with aRNA probes produced by one round of T7 RNA polymerase-based amplification (6 replicates, GSM4854-GSM4859), and with aRNA probes produced by two rounds of amplification (2 replicates, GSM4860-GSM4861). Keywords: parallel sample
Project description:Using high-resolution oligonucleotide arrayCGH, FISH, and RT-PCR we have performed a comprehensive analysis of genomic imbalances, and CRTC1-MAML2 gene fusion status in a series of 28 well characterized mucoepidermoid carcinomas (MECs) with the aims to identify distinct differences in genomic profiles and CRTC1-MAML2 gene fusion status between low- and high-grade MECs.
Project description:We used array-based comparative genomic hybridization (arrayCGH) of 76 hepatocellular carcinomas (HCCs) to search for genetically disrupted genes.
Project description:Microarrays offer a powerful tool for diverse applications plant biology and crop improvement. Recently, a global assembly of cotton ESTs was constructed based on three Gossypium. Using that assembly as a template, we now describe the design and creation and of a publicly available oligonucleotide array for cotton, useful for all four of the cultivated species. Synthetic oligonucleotide probes were generated from exemplar sequences of a global assembly of more than 150,000 cotton ESTs derived from 30 different cDNA libraries representing many different tissue types and tissue treatments. A total of 13,158 oligonucleotide probes are included on the arrays, optimized to target the diversity of the transcriptome but also including previously studied cotton genes, duplicated gene pairs derived from a paleoduplication event, transcription factors, and homology to protein coding genes in Arabidopsis. About 10% of the oligonucleotides target unidentified protein coding sequences, thereby providing an element of gene discovery. Because many oligonucleotides were based on ESTs from fiber-specific cDNA libraries, the array has direct application for analysis of the fiber transcriptome. To illustrate the utility of the array, we hybridized labeled bud and leaf cDNAs from G. hirsutum and demonstrate technical consistency of results. The cotton microarray provides a reproducible platform for transcription profiling in cotton, and is made publicly available through http://cottonevolution.info. Keywords: self vs. self; platform testing
Project description:Recently, long oligonucleotide (60-70mer) microarrays for two-color experiments have been developed and are gaining widespread use. In addition, when there is limited availability of mRNA from tissue sources, RNA amplification can and is being used to produce sufficient quantities of cRNA for microarray hybridization. Taking advantage of the selective degradation of RNA under alkaline conditions, we have developed a method to “strip” glass-based oligonucleotide microarrays that use fluorescent RNA in the hybridization, while leaving the DNA oligonucleotide probes intact and usable for a second experiment. Replicate microarray experiments conducted using stripped arrays showed high reproducibility, however, we found that arrays could only be stripped and reused once without compromising data quality. The intraclass correlation (ICC) between a virgin array and a stripped array hybridized with the same sample showed a range of 0.90-0.98, which is comparable to the ICC of two virgin arrays hybridized with the same sample. Using this method, once stripped oligonucleotide microarrays are usable, reliable, and should help to reduce costs. Keywords = Agilent microarray Keywords = Stripped arrays Keywords = Replicate reproducibility Keywords: other