Microarray Analysis of Spleen Transcriptome

For microarray-based gene-expression analysis, we pooled equal shares of seven individual RNA samples isolated from the spleen in six separate specimens according to treatment and tank (A1, B1, C1; A2, B2, C2). These six RNA pools were converted to Cy3-labelled cRNA and hybridised with 8 × 60 K Agilent-049158 Salmon Oligo Microarrays (Agilent Technologies; GEO platform: GPL21057) following the Agilent 60-mer oligo microarray processing protocol, as described in our previous paper³⁶. The fluorescence signals of the hybridised Agilent microarrays were scanned with a G2505C Microarray Scanner System (Agilent Technologies) at a resolution of 3 µm.
For all hybridisations, two technical replicates were included representing exactly the same samples but applied to independent arrays. The reliability of the microarray-predicted data has previously been proven in various quantitative PCR studies of our group^{9 ,23 (link),69 (link),82 (link),83 (link)}.

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Rebl A., Korytář T., Borchel A., Bochert R., Strzelczyk J.E., Goldammer T, & Verleih M. (2020). The synergistic interaction of thermal stress coupled with overstocking strongly modulates the transcriptomic activity and immune capacity of rainbow trout (Oncorhynchus mykiss). Scientific Reports, 10, 14913.

Publication 2020

G2505C Microarray Scanner System

Crna Fluorescence Gene expression Hybridisations Microarray Oligo Salmon Specimens treatment Spleen

Corresponding Organization :

Other organizations : Research Institute for Farm Animal Biology (FBN), University of South Bohemia in České Budějovice, University of Bergen, Friedrich-Loeffler-Institut, University of Rostock

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Variable analysis

independent variables

Treatment

dependent variables

Gene expression

control variables

Equal shares of seven individual RNA samples isolated from the spleen in six separate specimens
Cy3-labelled cRNA hybridisation with 8 × 60 K Agilent-049158 Salmon Oligo Microarrays
Two technical replicates representing the same samples but applied to independent arrays

positive controls

Quantitative PCR studies from previous publications by the research group (references 9, 23, 69, 82, 83)

negative controls

Not explicitly mentioned

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