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Figure 1 | Proteome Science

Figure 1

From: A systems approach to clinical oncology: Focus on breast cancer

Figure 1

How the DASL Assay works. A. The DASL Assay monitors gene expression with probe groups to query total RNA-generated cDNA target sequences. Total RNA, prepared from FFPE tumour tissue, is converted to cDNA using both biotinylated (designated as b) random nonamers and oligo dT. Primer sites, incorporated within each probe group, are used for PCR amplification. Also within the probe group is an address sequence complementary to one of the 1536 sequences on the Sentrix® Array Matrix. Three probe group per gene are used that allows for detection of small (1.3) fold-changes in expression between samples. Probe groups are annealed to the biotinylated cDNA, followed by selection of the duplexes on streptavidin beads to remove unhybridized oligos. Correctly annealed, assay specific oligos are extended and ligated to a locus-specific oligo. The locus-specific oligo incorporates an address sequence and primer site for the generation of amplifiable products. B. Address-containing templates, that are labeled with either cy3 or cy5 fluorescent dyes during PCR amplification, are subsequently hybridized to complementary address sequences immobilized to beads on the arrays. The array is then scanned to acquire intensity data.

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