Short CommunicationEvidence of systematic expressed sequence tag IMAGE clone cross-hybridization on cDNA microarrays
Section snippets
Results
We examined data from three cDNA microarray experiments: two experiments performed by separate investigators using commercially manufactured single-dye spotted cDNA arrays at the University of Pittsburgh (GF400, GeneFilters Microarrays; Research Genetics, Carlsbad, CA, USA; Peters et al., unpublished data, and Field et al., unpublished data) and one two-dye (Cy3/Cy5) experiment published on the publicly available Stanford Microarray Database Web site [6]. The first spotted cDNA microarray
Expressed sequence tags
ESTs are short (200–500 bp) sequences derived from directionally cloned plasmid cDNA libraries. Typically, total mRNA is isolated from cells in a particular type of tissue, stage of development, pathological state (e.g., normal versus tumor), or environmental/nutritional state (e.g., heat shock). The mRNA is reverse-transcribed using an oligo(dT) sequence primed with a restriction site. The resultant cDNA is then cloned into a plasmid vector, isolated, and one-pass sequenced, with the sequence
TZD-treated adipocytes experiment
The first spotted cDNA microarray experiment consisted of a time-sequenced sampling of differential mRNA expression from 3T3L1 cultured mouse adipocytes treated with the insulin-sensitizing agent TZD (Peters et al., unpublished data). Cells were harvested in 5 ml of Trizol (Invitrogen Corp., Carlsbad, CA, USA) and RNA was extracted according to the manufacturer's instructions. RNA integrity for each sample was confirmed on formaldehyde/formamide agarose gels prior to microarray analysis. cDNA
Acknowledgements
The authors thank Jay Kadane, Takis Benos, and Joe Ramsey for their helpful comments. This work was supported by NASA Grant NCC2-1227 and the Copeland Fund of the Pittsburgh Foundation Grant D200-0251.
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