The microarray is manufactured by Agilent using its proprietary SurePrint inkjet technogy that synthesizes 60mer oligonucleotides onto specially-prepared glass slides. For detailed information see manufacturer's web site http://www.agilent.com/.
Description
The microarray was designed in collaboration with an Agilent-certified service provider (imaGenes, Berlin, Germany) that applied an in-house developed approach for selection of the best performing probes for each gene (http://www.imagenes-bio.de/services/microarray/pss/mechanism). Using the F. g. genome annotation v3.0 annotation (FG3), for each gene model, several 60mer oligonucleotides were selected by the software eArray (https://earray.chem.agilent.com/earray/) and were then filtered by proprietary software (imaGenes). In addition to the 13,332 genes of the F. g. genome annotation v3.0, available cDNA sequences of the TRI (= trichothecene biosynthesis) gene cluster from nivalenol (NIV) producing strains of F. graminearum were chosen for oligonucleotide selection. Several suitable oligonucleotides per gene were assembled on a pre-selection array (2x105K format) and hybridized against Cy3-labeled genomic DNA and cRNA (from a pooled RNA sample), respectively. Based on the RNA hybridization results, for each gene the relative ratio of probes was determined that yielded significant signal intensities. In case that this ratio was 60% higher than the background, a gene was considered expressed. For these genes, final oligonucleotide selection relied on the results from RNA hybridization, whereas for genes considered non-expressed hybridization results from genomic DNA were used. The final 15K array design comprises 15,744 features (15,208 F. g. - specific oligos + 536 internal controls). 11,476 genes and 10 TRI cluster ESTs were represented by one oligonucleotide, 1,854 genes and 7 TRI cluster ESTs two probes were chosen.
