Every step was performed in the HS 4800 hybridization station (Tecan, Mannedorf, Switzerland). Slide pretreatment included (a) stripping: 2.5 mmol/L Na2HPO4, 0.1% SDS at 76°C for 1 min; (b) slide blocking: 3% bovine serum albumin (fraction V; Sigma-Aldrich), 5× SSC, 0.2% SDS at 42°C for 20 min; and (c) prehybridization: 400 μg of salmon testes DNA and 67.5 μg of COT human DNA in 200 μL of hybridization solution incubated 1 h at 37°C. Hybridization was performed during 18 h at 37°C under low agitation.Four posthybridization washes were performed: 50% formamide, 2× SSC, 0.1% SDS at 45°C for 37 sec, four times; 2× SSC, 0.1% SDS at 45°C for 37 sec, four times; 4× SSC, 0.1 Tween 20 at 45°C for 37 sec, four times; slides were finally nitrogen-dried.
Scan protocol
ScanArray 4000 (Packard Bioscience, Meriden, CT). Images were quantified using the system's Imagene 5.1 software as described previously [18].
Description
none
Data processing
Images were quantified using the system's Imagene 5.1 software. Background substraction, replicates averaging and normalization to the median of sub-telomeric clones.
