Microarray Gene Expression Analysis Protocol

Microarray analysis was performed as previously described^{56 (link)}. In brief, Alexa 555- or 647-labelled cDNA was produced from the RNA, using a Superscript direct cDNA labelling system (Invitrogen) and Alexa 555 and 647 dUTP label mix. The cDNA was then purified using an Invitrogen PureLink PCR Purification system. The cDNA was hybridized to the array using a Gene Expression Hybridization kit (Agilent). The array was an Agilent custom-designed array containing 60-mer oligonucleotides synthesized in situ on the array and contained 4 × 44,000 probes. Following hybridization for at least 17 h, the array was washed using a Gene Expression Wash Buffer kit (Agilent) and scanned in an Agilent Array Scanner. The microarray signal was extracted using GenePix.

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Pai C.C., Deegan R.S., Subramanian L., Gal C., Sarkar S., Blaikley E.J., Walker C., Hulme L., Bernhard E., Codlin S., Bähler J., Allshire R., Whitehall S, & Humphrey T.C. (2014). A histone H3K36 chromatin switch coordinates DNA double-strand break repair pathway choice. Nature communications, 5, 4091.

Publication 2014

Superscript direct cDNA labelling system PureLink PCR Purification system Gene Expression Hybridization kit custom-designed array Gene Expression Wash Buffer kit

Buffer Cdna Dutp Gene expression Hybridization Microarray Oligonucleotides

Corresponding Organization :

Other organizations : University of Oxford, University of Edinburgh, Newcastle University, University College London

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Gene expression levels

control variables

None explicitly mentioned

controls

Positive control: None mentioned
Negative control: None mentioned

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