This manual details the Affymetrix GeneChip Whole Transcript (WT) Sense Target Labeling Assay for generating amplified, biotinylated sense-strand DNA…
The Affymetrix GeneChip® Whole Transcript (WT) Sense Target Labeling Assay is designed to generate amplified and biotinylated sense-strand DNA targets from the entire expressed genome without bias. This assay is optimized specifically for use with GeneChip® ST Arrays, where probes are distributed throughout the entire length of each transcript.
The manual describes two configurations: the 1 µg Total RNA Labeling Protocol, which includes an rRNA reduction step recommended for high-sensitivity analysis on Exon ST Arrays, and the 100 ng Total RNA Labeling Protocol, which omits this step and is acceptable for Gene ST Arrays. Both protocols merge after rRNA reduction to synthesize double-stranded cDNA, amplify it into antisense cRNA, convert it back to single-stranded DNA in a sense orientation, fragment it, label it with biotin, and prepare it for hybridization.
| Compatible Arrays | GeneChip® ST Arrays (Sense Target) |
|---|---|
| Protocol 1 Input Requirement | 1 µg Total RNA |
| Protocol 2 Input Requirement | 100 ng Total RNA |
No, the WT Assay is not compatible with GeneChip arrays designed to focus on the 3’ ends of the transcripts.
GCOS will fail to generate a .cel file, and failed subgrids are visualized with red borders and an “x” in the center.