The proprietary CHO-specific DNA microarray (Affymetrix) used in this study consisted of 61,223 probe sets, which targeted approximately 26,227 Chinese hamster unique gene IDs, and 14,657 unique Ensembl mouse genes. Before being hybridised to the microarray, all of the mRNA samples were diluted to the same concentration (50 ng/μl). Biotinylated cRNA was prepared according to the protocol described in the Affymetrix technical manual. The subsequent hybridisation was performed in a GeneChip Hybridisation oven 640 (Affymetrix), and the processing was carried out using a GeneChip Fluidics station 450 (Affymetrix).
Genechip hybridisation oven 640
Manufactured by Thermo Fisher Scientific
The GeneChip Hybridisation oven 640 is a laboratory instrument designed for the incubation and hybridization of microarray samples. It provides a controlled environment for the hybridization process, ensuring consistent temperature and agitation for optimal results.
Automatically generated - may contain errors
Lab products found in correlation
Genechip fluidics station 450, by Thermo Fisher Scientific (1 mentions)
Genechip human exon 1.0 st array, by Thermo Fisher Scientific (1 mentions)
Genechip human genome u133 plus 2.0 array, by Thermo Fisher Scientific (1 mentions)
Nugen ovation rna amplification system v2, by Tecan (1 mentions)
Fl ovation cdna biotin module v2, by Tecan (1 mentions)
Genechip scanner 3000, by Thermo Fisher Scientific (1 mentions)
Trizol reagent, by Thermo Fisher Scientific (1 mentions)
3 protocols using genechip hybridisation oven 640
1
CHO Cell Transcriptome Analysis via Affymetrix Microarray
2
Affymetrix GeneChip Human Exon 1.0 ST Array
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Patient samples and samples from functional studies on BL2 cells were analysed for mRNA expression using the Affymetrix GeneChip Human Exon 1.0 ST Array (Affymetrix, Santa Clara, CA, USA). The experiment was conducted according to the manufacturer’s protocol. In brief, 250 ng RNA per sample were amplified, transcribed to cDNA, fragmented and subsequently labelled with Biotin. Array hybridisation was performed at 45 °C for 16–18 h in the Affymetrix GeneChip Hybridisation Oven 640, arrays were subsequently washed in the Fluidics Station 450 and scanned on the GeneChip scanner 3000 7G. Complete GEP data sets are available at Gene Expression Omnibus (GEO) with accession number GSE98529.
3
RNA Extraction and Microarray Analysis
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RNA was extracted from explants cultured to time-point 120 h according to Chomczynski method [8] with TRIZOL reagent (Invitrogen, UK) and quality assessed. Details of the methods for RNA extraction, quality and integrity assessments are previously published [1] . Total RNA was processed into labelled cDNA with NuGEN™ Ovation™ RNA Amplification System V2 and FL-Ovation™ cDNA Biotin Module V2 (Nugen). The resultant fragmented and labelled cDNA was added to the hybridisation cocktail in accordance with the NuGEN™ guidelines for microarray hybridisation onto Affymetrix GeneChip® Human Genome U133 Plus 2.0 arrays (sample per array) in Affymetrix GeneChip® Hybridisation Oven 640 for 18 hours at 45°C. Features that retained bound labelled cRNA after washing were visualized using the GeneChip® Scanner 3000 (Affymetrix). The microarray data was published in the Gene Expression Omnibus (GEO) repository with accession number GEO: GSE74446.
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