GeneChip microarrays

Primary PCR products are subjected to labeling PCR as described by Affymetrix. The 24 labeled PCR samples corresponding to each genomic DNA sample are then pooled, diluted in hybridization buffer, heated to 95°C, cooled to 4°C, then hybridized with Affymetrix HuSNP GeneChip microarrays at 45°C for 18 h. 

There are many protocols and different types of platforms available, e.g. GeneChips from Affymetrix, Illumina Bead Arrays, Arrays from Agilent, Applied Biosystems, GE Healthcare, customized spotted cDNA microarrays etc. the basic procedure for a large-scale measurement of gene expression involves the preparation of total or mRNA from the biological sample(s) under investigation (e.g. candidate tissue) and the hybridization of copied labelled RNA or cDNA to the DNA elements on the array surface (Fig. 1). 

The commercial success of the spotted microarray, like that of Affymetrix s GeneChip, is content-driven. In order to provide customers with comprehensive gene expression microarray products, manufacturers must obtain gene-specific annotated sequences covering genomes of major interest to the scienhfic community (e.g., genomes of humans, yeasts, and mice). 

The proteomics field is currently wide open. Only a few protein microarray products have been commercialized. Unlike DNA microarrays that are commonly constructed on essentially two platforms (GeneChip or slide array) employing oligonucleotides or cDNA probes, the protein microarray appears on a variety of platforms and with different types of probes. 

The Affymetrix U95A GeneChip containing 12,500 known human genes was used for this purpose. Two adjacent breast tumors from an 87-year old woman were removed and used in the study along with an additional five breast tumors obtained from different individuals. Thus, seemingly related tumors from one individual could be compared with tumors from different sources to validate the microarray-based prognosis. In addition to validation of the microarray for this purpose, the researchers examined a number of... 

Wen Z et al (2010) Evaluation of gene expression data generated firom expired Affymettix GeneChip(R) microarrays using MAQC reference BNA samples. BMC Bioinformatics ll(Suppl 6) S10. doi 1471-2105-ll-S6-S10 (pii)10.1186/1471-2105-ll-S6-S10... 

Woo Y et al (2004) A comparison of cDNA, oligonucleotide, and Affymetrix GeneChip gene expression microarray platforms. J Biomol Tech 15 276-284. doi 15/4/276 (pii)... 

Alternative microarray approaches are available that involve the 3 tailing of the miRNA molecule with poly(A), followed by labeling and hybridization. The NCode miRNA microarray platform (Life Technologies) involves the addition of the poly(A) tail to the miRNA prior to ligation of fluorescent dye molecules and subsequent hybridization to the antisense miRNA probes on the microarray chip. In the case of Affymetrix GeneChip microRNA arrays, the poly(A)-tailed miRNAs are labeled with a biotinylated signal molecule. A limitation of hybridization-based microarrays is that because of the small size of miRNAs, the Tm of the probes has a wide range that may diminish specificity and/or sensitivity for miRNAs with a low GC content (10,56). 

Material collection —> mRNA microarray (GeneChip or CDNA) — Data mining tools <-> Drug Discovery... 

The transcriptome is defined as the whole set of transcripts present in a cell in a given developmental stage and under a specific physiological condition. As we have been describing, the transcriptome is composed by mRNAs, rRNAs, tRNAs, and a set of ncRNAs (22,102). Over the past decade, the transcriptome has been analyzed by hybridization-based approaches (microarrays). Typically, the RNA is previously converted into cDNA, fluorescently labeled, and hybridized onto a specific microarray (such as Affymetrix GeneChips). They have been widely used over the past 15 years and have given quite precise information about the way the transcriptome is regulated... 

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