RNA: 15 ug of Total RNA was labeled with amino-allyl dUTP during reverse transcriptase and subsequently coupled with the succinimidyl ester fluorescent dye AlexaFluor 546 as previously described (Hovel-Miner, G 2009. JB, 191:2461). Genomic DNA: 5 ug of genomic DNA was labeled with amino-allyl dUTP with klenow fragment and random hexamer as previously described (Porwollik, S. 2001. Mutat Res 483:1). Subsequently, the resulting DNA was coupled to the succinimidyl ester fluorescent dye AlexaFluor 647.
RNA: 15 ug of Total RNA was labeled with amino-allyl dUTP during reverse transcriptase and subsequently coupled with the succinimidyl ester fluorescent dye AlexaFluor 546 as previously described (Hovel-Miner, G 2009. JB, 191:2461). Genomic DNA: 5 ug of genomic DNA was labeled with amino-allyl dUTP with klenow fragment and random hexamer as previously described (Porwollik, S. 2001. Mutat Res 483:1). Subsequently, the resulting DNA was coupled to the succinimidyl ester fluorescent dye AlexaFluor 647.
Hybridization protocol
Hybridization was performed as previously described (Hovel-Miner, G 2009. JB, 191:2461).
Scan protocol
Scanning and image acquisition was performed on a InnoScan 710 using the mapix software. Spot intensity an local background was measured.
Description
Fraq_6h_rep1
Data processing
Data were processed with the mapix software. Values from each channel were background substracted and the % contribution of each sopts to the total fluorsecence in each channel was calculated. Then the ratio Ch2/Ch1 was calculated. We used gDNA for data normalization accross slides, as described in Talaat, 2002, NAR.
