A custom-built insulin resistance gene chip.
OBJECTIVE
Microarray (gene chip) technology offers a powerful new tool for analyzing the expression of large numbers of genes in many experimental samples. The aim of this study was to design, construct, and use a gene chip to measure the expression levels of key genes in metabolic pathways related to insulin resistance.


METHODS
We selected genes that were implicated in the development of insulin resistance, including genes involved in insulin signaling; glucose uptake, oxidation, and storage; fat uptake, oxidation, and storage; cytoskeletal components; and transcription factors. The key regulatory genes in the pathways were identified, along with other recently identified candidate genes such as calpain-10. A total of 242 selected genes (including 32 internal control elements) were sequence-verified, purified, and arrayed on aldehyde-coated slides.


RESULTS
Where more than 1 clone containing the gene of interest was available, we chose those containing the genes in the 5' orientation and an insert size of around 1.5 kb. Of the 262 clones purchased, 56 (21%) were found to contain sequences other than those expected. In addition, 2 (1%) did not grow under standard conditions and were assumed to be nonviable. In these cases, alternate clones containing the gene of interest were chosen as described above. The current version of the Insulin Resistance Gene Chip contains 210 genes of interest, plus 48 control elements. A full list of the genes is available at http://www.hbs.deakin.edu.au/mru/research/gene_chip_tech/genechip_three.htm/.


CONCLUSIONS
The human Insulin Resistance Gene Chip that we have constructed will be a very useful tool for investigating variation in the expression of genes relevant to insulin resistance under various experimental conditions. Initially, the gene chip will be used in studies such as exercise interventions, fasting, euglycemic-hyperinsulinemic clamps, and administration of antidiabetic agents.
Walder
Ken
K
Metabolic Research Unit, Deakin University, Geelong, VIC, Australia. walder@deakin.edu.au
Segal
David
D
Chehab
Sam
S
Augert
Guy
G
Cameron-Smith
David
D
Hargreaves
Mark
M
Collier
Greg R
GR
D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D004587:Electrophoresis, Agar Gel; D007333:Insulin Resistance / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis; D015203:Reproducibility of Results
D017931:DNA Primers