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Expression array design concepts

Transcript Level Expression Profiling from Predicted and Transcribed Sequences with a 5 µ m, PM-only Tomato Array. WT with every predicted Exon (Exon Array strategy) Select equal number of probes from every exon. Expression array design concepts. 3’ IVT Probe-set (Traditional strategy)

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Expression array design concepts

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  1. Transcript Level Expression Profiling from Predicted and Transcribed Sequences with a 5 µm, PM-only Tomato Array

  2. WT with every predicted Exon (Exon Array strategy) Select equal number of probes from every exon Expression array design concepts 3’ IVT Probe-set (Traditional strategy) Single probe-set per gene (if possible) Complicated by alternative polyadenylation and splicing WT for Expressed Sequence or Gene Model (Gene Level strategy) Select equal number of probes from every transcript or gene coding sequence (20-30 PM probes per gene)

  3. Tomato transcript level expression arrayBenefits • Enhanced detection of ESTs, gene fragments, and SNPs • Increased genome coverage to >35,000 transcripts through smaller feature size • Reduced array price through smaller format size • Leverage Affymetrix expertise in array and assay development

  4. BenefitsEnhanced Detection • Enhanced detection of non-3’ anchored ESTs and gene fragments with Whole Transcript Assay (WTA) • Random-primed amplification rather than 3’ anchored (oligo-dT priming) • Advantages • Greater number of probes per transcript (≥ 30) • Enables detection along length of transcript, not just 3’ end • Good sensitivity pairing WTA with gene level array • Potential for improvement with rRNA depletion

  5. BenefitsIncreased content • TOM1 • 12,000 spotted cDNAs • TOM2 • 11,000 spotted oligos • Affymetrix GeneChip Tomato Genome array • 222,000 probes representing 9,254 transcripts • 11 probe pair 3’ IVT strategy • Proposed GeneChip transcript level design • 2.6 million probes representing more than 35,000 transcripts • Full Unigene content • 30 or more probes per transcript • Facilitates SFP detection • 5 µm feature size • Additional space to enable other applications • Transcript discovery • ChIP-ChIP

  6. BenefitsReduced price • No Design Fee and reduced array price • First 500 arrays at €200 per array • Second 500 arrays at €150 per array • All remaining orders at €125 per array • Initial higher per array price to cover design and investment costs • Proposed GeneChip transcript level design • 2.6 million probes representing more than 35,000 transcripts

  7. Logistics • Tomato community designate to provide sequence files of transcripts to be detected on array • Recommend sequences be annotated prior to submission • Affymetrix QCs sequences, selects array probes, and creates array design • Design review by community designate • No manufacturing until all agree on final design • Timeline for array delivery is 8 weeks once design is complete

  8. Model Organism / Consortia DesignsExperience making arrays

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