"Illumina ™ TotalPrep ™ RNA Amplification Kit + Bio-16-UTP (10 mM)

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	Check the cDNA Binding Buffer for precipitation before using it. If a precipitate is visible, redissolve it by warming the solution to 37°C for up to 10 min and vortexing vigorously. Cool to room temperature before use

Protocol tips
Check the cDNA Binding Buffer for precipitation before using it. If a precipitate is visible, redissolve it by warming the solution to 37°C for up to 10 min and vortexing vigorously. Cool to room temperature before use

Publication protocol

Gene array hybridization
Each of the nine RNA samples was amplified using the Ambion Illumina RNA amplification kit with biotin UTP labeling. The Ambion Illumina RNA amplification kit uses T7 oligo(dT) primer to generate single stranded cDNA followed by a second strand synthesis to generate double-stranded cDNA, which is then column purified. In vitro transcription was done to synthesize biotin-labeled cRNA using T7 RNA polymerase. The cRNA was then column purified and a total of 750 ng was hybridized for each array using standard Illumina protocols with streptavidin-Cy3 used for detection. The MouseRef-8 v2.0 Illumina beadchip targets ~25,600 annotated RefSeq transcripts, over 19,000 unique genes, and enables the interrogation of eight samples in parallel. For practical purposes we will refer to the transcriptome-wide scope of data from this assay, although technically it is limited because some transcripts are not represented and some probe sets will perform better than others. Slides were scanned on an Illumina Beadstation and analyzed using GenomeStudio (Illumina, Inc.).

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Manufacturer protocol

Download the product protocol from Thermo Fisher Scientific for "Illumina ™ TotalPrep ™ RNA Amplification Kit + Bio-16-UTP (10 mM) below.

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