Standard microarray protocols include DNA extraction, DNA amplification and labeling, hybridization, washing, scanning, and data analysis.
Of note, our microarray protocol included an RNA amplification step, which was excluded in the k-PCR analysis.
For microarray, RNA was isolated using the Qiagen RNeasy kit following the manufacturer's protocol, including the addition of DNase.
The technology use transcript specific color coded molecular probe pairs to capture and count individual transcripts and has several advantages in comparison to microarray platform including easy protocols, less hands-on time and higher sample throughput [ 17].
RNA isolation and microarray protocols were carried out as described in Hu et al [30].
All microarray protocols were carried out by Mogene LLC (St . Louis MO, USA), an Agilent Certified Service Provider.
Microarray hybridization and washing conditions were based on E. coli cDNA microarray protocols [ 66] with some modifications.
The protocol included 3×12 reps.
More detailed protocols regarding the microarray assay, including microarray preparation, labeling of the cDNA probe, sample hybridization and washing can be obtained from the dbZach website [ 50].
The optimization of the microarray workflow, including the hybridization step, is thus a primary target for the evolution of more efficient protocols.
