Application of DNA microarrays in structural and functional genomics.

DNA microarrays started their career in the last decade of XX century. From that time a wide spectrum of microarray-based methods have been developed, contributing to the fast progress of structural and functional genomics. In a typical microarray experiment, DNA probes (short and long oligonucleotides, cDNA clones), complementary to the target sequences, are deposited on a solid surface . Target samples (DNA or RNA extracted from cells or tissues) are amplified (if necessary), fluorescently labeled, and hybridized with a microarray. In a two-color assay control and investigated samples are labeled with two different dyes, mixed and hybridized with the same array . Post hybridization scanning of a microarray with one/two lasers excite the fluorescent dyes that emit light. A digital microarray image generated by the scanner is then processed to convert signal intensities into quantitative data organized in tab-delimited files . These raw data files are submitted to normalization and analysis, including statistical testing and clustering , to retrieve biologically relevant information.

There are two basic applications of DNA microarrays: studying genome structure and gene expression analysis. In genome structure studies the following microarray-based methods are applied:

-          CGH (comparative genomic hybridization) arrays, at the beginning consisted of long chromosomal fragments (e.g. BAC clones, cDNA clones), sufficient for detection of large changes – deletions, insertions and copy number variation, today much more sensitive due to oligonucleotide probes application 

-          SNP arrays, useful in genotyping, designed to identify not only SNP (single nucleotide polymorphism), but also LOH (loss-of-heterozygosity), CNV (copy number variation), AI (allelic imbalance), UPD (uniparental disomy); at present DNA chips for identification millions of SNP and copy number variants in human genome are available from Affymetrix  or Illumina 

-          tiling arrays with oligonucleotide probes covering the whole genome sequence (except for repetitive sequences), applied for CGH, chromatin immunoprecipitation (ChIP-on-chip, based on analysis of DNA-proteins interactions ), methyl-DNA immunoprecipitation (MeDIP-chip, epigentetic studies), chromatin hypersensitive sites localization (DNase-chip), gene annotation and mapping 

-          multiple-tiling arrays, e.g. with double-tiled probes spanning the entire yeast genome 

-          TIP-chips (Transposon Insertion site Profiling chips), used for detection of transposons in yeast .

Although sequencing by hybridization did not gain popularity, array-like structures are common in DNA sequencing technologies. DNA microarrays are also used for studying relationships between organisms and microbial strain identification .

Isoenergetic microarrays, built of short oligonucleotide probes, help to predict secondary structure of RNA molecules. More often, RNA molecules in microarray experiment serve as targets for functional studies - differential expression of mRNAs and microRNAs . Such analysis determines which genes are up- and down-regulated comparing to control samples. Gene expression profiles (signatures) are specific for particular samples and conditions, e.g. developmental stage, disease type, stress . Here, the following arrays are applied:

-          Short oligonucleotide microarrays with probes synthesized in situ (DNA chips, multiple probes per gene) 

-          Microarrays consisted of long oligonucleotide probes (usually one per gene), synthesized in situ or pre-synthesized and deposited on the solid surface

-          cDNA (complementary DNA) arrays, where the probes, cDNA clones, are spotted onto a glass slide using printing robots

-          Tiling arrays, covering the whole genome sequence, applied for discovering new transcriptionally active regions and splice variants 

-          Exon arrays with separate probes designed to detect each exon, useful in detection of splicing isoforms .

In case of organisms with not sequenced genomes, cross-species hybridization (CSH, with microarray designed for a relative species), is a solution to follow gene expression . 

There is no doubt that microarrays are powerful tools in biology and medicine, applicable for genotyping, biomarker selection, medical diagnosis and prognosis, screening of potential therapeutics (pharmacogenomics), etc.. However, at the large-scale research field, sequencing technologies have more advantages, being more precise, accurate and less dependent on technical and statistical errors . The contemporary science started also to benefit from the integrated approaches, including sequencing as well as gene expression profiling with microarrays, as it is applied in cancer research area . 

The following materials and learning resources are available in this lecture:
[1] DNA microarray definition and description


[2] Film: “DNA microarrays” 


[3] Film: “Microarrays” 


[4] Film: “Microarray Gene Expression” 


[5] DNA microarrays and chips – description from “Genomes 2” 


[6] Article “The incredible shrinking world of DNA microarrays” 


[7] Article “DNA Microarrays: a Powerful Genomic Tool for Biomedical and Clinical Research” 


[8] Lecture: Introduction to array CGH analysis 


[9] Article “Identification of disease genes by whole genome CGH arrays” 


[10] SNP array


[11] Film: “Microarray Method for Genetic Testing” 


[12] Affymetrix SNP arrays 


[13] Illumina Whole Genome Genotyping arrays 


[14] ChIP-on-chip technology 


[15] Tiling array 


[16] Sequencing by hybridization 


[17] Article “Application of DNA microarray technology for detection, identification, and characterization of food-borne pathogens” 


[18] Article abstract “Binding of short oligonucleotides to RNA…” 


[19] Cross-species hybridization (CSH)


[20] Film: “Diagnostic Micoarrays” 


[21] Article “Progress in the Application of DNA Microarrays” 


[22] DNA Microarrays in Medicine 


[23] Article "The next generation of microarray research: applications in evolutionary and ecological genomics