Direct Hybridization Assay
The Direct Hybridization Assay system uses a standard Eberwine protocol
assay, in which gene-specific probes are used to detect labeled cRNAs. Each
bead in the array contains a 50-mer, sequence-specific oligo probe
synthesized in-house using Illumina’s proprietary oligo manufacturing
technology. Probe design incorporates a number of selection criteria
including:
` Sequence context evaluation identifying regions of a transcript
compatible with probe design, based on items such as splice-isoformspecific
regions, sequence complexity, and uniqueness.
` Self-complementarity for hairpin structure prediction
` Melting temperature (Tm)
` Distance from the 3' end of the transcript
A series of controls is incorporated into the assay, accomplishing these
important tasks:
` Assessment of success of the major steps in the experiment
` Information on background and noise levels
Direct Hybridization AssayThe Direct Hybridization Assay system uses a standard Eberwine protocolassay, in which gene-specific probes are used to detect labeled cRNAs. Eachbead in the array contains a 50-mer, sequence-specific oligo probesynthesized in-house using Illumina’s proprietary oligo manufacturingtechnology. Probe design incorporates a number of selection criteriaincluding:` Sequence context evaluation identifying regions of a transcriptcompatible with probe design, based on items such as splice-isoformspecificregions, sequence complexity, and uniqueness.` Self-complementarity for hairpin structure prediction` Melting temperature (Tm)` Distance from the 3' end of the transcriptA series of controls is incorporated into the assay, accomplishing theseimportant tasks:` Assessment of success of the major steps in the experiment` Information on background and noise levels
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