DNA microarray technology can be applied to carcinogen identification, toxicology, and drug safety. This technology is also a useful tool for investigating differentially-expressed genes in animals. Combining DNA microarray technology and genotyping, studies in our laboratory identified many novel genetic markers for animal selection. Microscopic DNA spots are printed using pins or needles controlled by a robotic arm and attached to a solid surface. The DNA spots contain a specific DNA sequence, known as probes. These can be oligonucleotides, cDNA or small fragments of PCR products that correspond to mRNAs. Probe-target hybridization is usually detected and quantified by detection of fluorophore- or chemiluminescence-labeled targets to determine relative levels of transcripts. A flow chart of fabrication and analysis of DNA microarray for identifying novel SNP markers. In order to identify differentially-expressed genes that are correlated with hatchability in the Tsaiya duck, a cDNA microarray-aided assay was performed. Total RNA samples from the magnum epithelium of 6 laying brown Tsaiya ducks with wide range variations in hatchability (44.00–81.82%) were collected and pooled for cDNA library construction. Polymerase chain reaction was performed to amplify the cDNA inserts. The PCR products were subsequently purified by isopropanol precipitation, washed with 70% ethanol, resuspended in 20 μl of 50% dimethyl sulfoxide (DMSO), and transferred to 384-well microplates as microarray sources for printing. Printing was performed under the conditions of 60% relative humidity and a 300-μm dot space. The 2912 amplicons were printed onto GAPS II-coated glass slides (Corning Life Sciences, Corning, NY, USA) using an OmniGrid AccentTM microarrayer (GeneMachine, Bethesda, MD, USA); each amplicon was spotted in triplicate, resulting in 8736 spots on each slide. The fluorophore-labeled targets hybridize to probes on the microarray and hybridization signal scanning was carried out. Differentially-expressed transcripts associated with hatchability were found.
Author(s) Details:
Hsiu-Lin Huang,
Department of Animal Science, National Chung Hsing University, 145 Xingda Road, Taichung 402, Taiwan.
I-Yen Huang,
Ugintech Co. Ltd., 5F-5, No. 282, Zhishi North 2nd Road, Taichung 407, Taiwan.
Chia-Yu Lin,
Johnson Chemical Pharmaceutical Works Co. Ltd., No. 77, Sec. 4, Sanhe Road, Sanchong Dist., New Taipei City 241, Taiwan.
Mu-Chiou Huang,
Department of Animal Science, National Chung Hsing University, 145 Xingda Road, Taichung 402, Taiwan.
