Total RNA for the microarray was isolated with Direct-Zol RNA Mincludingit (Zymo Research, Irvine, CA) including on-column DNAse treatment.
Total RNA for analysis by microarray was isolated in triplicate from 30 ml of cultures of infected and uninfected cells and cells treated with LPS for 4 and 18 h.
The RNA, to be used for the microarrays, was isolated using Trizol reagent (Invitrogen, USA).
Of the 29 putative Pat substrates identified by the microarray experiments, 22 were isolated using the ASKA collection [23].
For the creation of the cDNA microarrays, midgut tissues were isolated from neonates of both species that had fed for 24 h on WT plants elicited with 250 µg methyljasmonate (MeJA) in 20 µl of lanolin, three days prior to larval feeding.
For the fibroblast ECM microarray analysis, cardiac fibroblasts were isolated from adult C57BL/6J mice and stimulated with or without 10 ng/ml TGF-β1 for 24 hours[ 61].
Importantly, total RNA pools used in microarray and qRT-PCR analyses were isolated in independent experiments.
To identify the primary target, DEGs were isolated after microarray analysis and classified according to their temporal expression pattern; GO terms of early changed DEGs were analyzed using BinGO software.
Then, cells were harvested after 60 h, RNAs were isolated, and microarrays were performed using Human Microarray (OneArray, PhalanxBio, Inc., San Diego, CA, USA).
