Sentrix human ht 12 v3 expression beadchips

Transcriptional profiling was determined using Illumina Sentrix BeadChips. Total RNA was used to generate biotin-labeled cRNA using the Illumina TotalPrep RNA Amplification Kit. In short, 0.5ug of total RNA was first converted into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated incorporating biotin-16-UTP. A total of 0.75ug of biotin-labeled cRNA was hybridized at 58°C for 16 hours to Illumina’s Sentrix Human HT-12 v3 Expression BeadChips (Illumina, San Diego, CA). Each BeadChip has around 48,000 transcripts with approximately 15-fold redundancy. The arrays were washed, blocked and the labeled cRNA was detected by staining with streptavidin-Cy3. Hybridized arrays were scanned using an Illumina BeadStation 500X Genetic Analysis Systems scanner and the image data extracted using the Illumina GenomeStudio software, version 1.1.1). Data are available in the GEO database, under accession #GSE50004.

Transcriptional profiling was determined using Illumina Sentrix BeadChips. Total RNA was used to generate biotin-labeled cRNA using the Illumina TotalPrep RNA Amplification Kit. In short, 0.5 µg of total RNA was first converted into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated incorporating biotin-16-UTP. A total of 0.75 µg of biotin-labeled cRNA was hybridized at 58°C for 16 h to Illumina's Sentrix Human HT-12 v3 Expression BeadChips (Illumina, San Diego, CA). Each BeadChip has around 48,000 transcripts with approximately 15-fold redundancy. The arrays were washed, blocked and the labeled cRNA was detected by staining with streptavidin-Cy3. Hybridized arrays were scanned using an Illumina BeadStation 500× Genetic Analysis Systems scanner and the image data extracted using the Illumina GenomeStudio software, version 1.1.1). Data are available in the GEO database (accession # GSE57445).