biostudies-arrayexpressMetabolomicsUnknownTranscriptomicsGenomicsProteomicsNoriyuki Nishimuratranscription profiling by arrayArabidopsis thalianaArabidopsis thalianahttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-16613Transgenic plants overexpressing ABI1, AHG1, and AFP3 exhibit the phenotype implicating ABA response in seeds; however, it is not well known whether they have key roles in the ABA response under normal growth conditions. To elucidate whether ABI1, AHG1, and AFP3 affect and regulate downstream pathways, we performed microarray experiments using the plants of YFP-ABI1ox, YFP-AHG1ox, YFP-AFP3ox, and YFPox control lines that were grown for 3 weeks on normal MS plates.biostudies-arrayexpressHybridization - Cy3-labeled, linearly amplified cRNA was purified using the RNeasy Mini Kit (Qiagen), removing unincorporated labeled nucleotides (monomers). Purified cRNA (1.65 µg) was combined with Blocking Agent and Fragmentation Buffer and incubated at 60 °C for 30 min to fragment the RNA. After cooling, an equal volume of 2× GE Hybridization Buffer HI RPM was added gently. The hybridization mixture was applied to an Agilent SureHyb gasket slide and overlaid with an Agilent Arabidopsis Gene Expression Microarray (Agilent 021169, V4). The assembled chamber was incubated at 65 °C for 17 h in a hybridization oven equipped with a rotator set to 10 rpm (Agilent G2545A). After hybridization, the cover slip was removed while the slide was submerged in Wash Buffer 1. Slides were washed for 1 min in Wash Buffer 1 followed by 1 min in Wash Buffer 2. Slides were then lifted slowly from Wash Buffer 2 and gently blotted to remove excess liquid before scanning.Growth Protocol - Arabidopsis thaliana (ecotype Col-0) plants were grown on MS plates containing 1 x Murashige and Skoog salt mix, 1 % sucrose, 2.5 mM MES (pH 5.8), and 0.8 % agar at 22 °C under 16 h light/8 h dark cycles for three weeks.Sample Collection - Three-week-old seedlings grown on MS plates were harvested and immediately frozen in liquid nitrogen. All samples were stored at −80 °C until RNA extraction.Nucleic Acid Extraction - Total RNA was isolated with a RNeasy Plant Mini Kit (Qiagen) according to the manufacturer’s instructions. RNA concentration and purity were measured with a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific), and RNA integrity was verified by agarose gel electrophoresis.Labeling - The reaction mixtures, which included a total of 200 ng of diluted total RNA, Spike-In controls, and T7 promoter primers, were incubated at 65 °C for 10 min to denature RNA and primers , followed by cooling on ice for 5 min. cDNA synthesis was performed by adding cDNA Master Mix and incubating at 40 °C for 2 h, after which the reaction was heated at 70 °C for 15 min to inactivate the enzyme. After cooling for 5 min, Transcription Master Mix was added, and in vitro transcription with Cy3 labeling was carried out at 40 °C for 2 h to generate Cy3-labeled cRNA.Scaning - Hybridized slides were loaded into the Agilent G2565CA Microarray Scanner system using manufacturer-recommended one-color settings (AgilentHD_GX_1Color for 4×44K microarrays). The resulting TIFF images were loaded into Agilent Feature Extraction software, and feature extraction was performed using the standard one-color protocol. After extraction, quality control metrics were reviewed using the QC Report in the Summary Report tab, and grid placement was verified by examining spot finding results across the array, including the four corners.MIAME ScoreRaw DataOrganizationAssays and DataMAGE-TAB FilesArray DesignsNoriyuki NishimurafalseTranscriptional profiling by microarray analysis of control plants and Arabidopsis thaliana lines overexpressing YFP-AHG1, YFP-AFP3, and YFP-ABI1Transgenic plants overexpressing ABI1, AHG1, and AFP3 exhibit the phenotype implicating ABA response in seeds; however, it is not well known whether they have key roles in the ABA response under normal growth conditions. To elucidate whether ABI1, AHG1, and AFP3 affect and regulate downstream pathways, we performed microarray experiments using the plants of YFP-ABI1ox, YFP-AHG1ox, YFP-AFP3ox, and YFPox control lines that were grown for 3 weeks on normal MS plates.2026-07-28T00:00:00Z2026-07-28T09:33:04.826Z2026-01-29T13:39:12.65ZE-MTAB-16613EFO_0002768EFO_0002944EFO_0003814EFO_0003813EFO_0003789EFO_0005518EFO_0003815