SureSelect Target Enrichment for the Ion Proton TM Next Generation Sequencing System

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1 SureSelect Target Enrichment for the Ion Proton TM Next Generation Sequencing System Demonstrated performance you can count on Christina Chiu Product Manager, SureSelect Kyeong Jeong Ph.D. R&D Scientist

2 SureSelect for Ion Proton Outline SureSelect for Ion Proton SureSelect Key Benefits Workflow Performance Variant Calling

3 SureSelect for Ion Proton Outline SureSelect for Ion Proton SureSelect Key Benefits Workflow Performance Variant Calling

4 SureSelect The Pioneer in Target Enrichment

5 SureSelect Target Enrichment System Workflow

6 SureSelect Sequencing Platforms

7 SureSelect Available for Ion Proton

8 SureSelect Target Enrichment Reagent Kit Ion Proton SureSelect TE Reagent Kit, Ion Proton compatible with SureSelect XT baits Human Exome All Exon 50Mb All Exon V4 All Exon V5 All Exon V5 + UTRs All Exon V5 + lncrna Other DNA Kinome Mouse All Exon Custom 1kb-24Mb

9 SureSelect for Ion Proton Outline SureSelect TE Reagent Kit, Ion Proton SureSelect Key Benefits Workflow Performance Variant Calling

10 SureSelect Key Benefits PROVEN Technology High PERFORMANCE FLEXIBLE & COMPLETE Solution

11 SureSelect Key Benefits PROVEN Technology High PERFORMANCE FLEXIBLE & COMPLETE Solution

12 SureSelect Most Published Target Enrichment Platform >350 Publications and many more to come

13 SureSelect Key Benefits PROVEN Technology High PERFORMANCE FLEXIBLE & COMPLETE Solution

14 SureSelect High Performance Core Technology Benefits Ultra-Long RNA Baits (120-mer) Binding strength RNA:DNA > DNA:DNA Better Sensitivity Detect more SNP, InDels, CNV, fusions Better Allelic Balance Equal representation of both alleles

15 SureSelect Key Benefits PROVEN Technology High PERFORMANCE FLEXIBLE & COMPLETE Solution

16 SureSelect Flexible & Complete Solution Exomes or custom (up to 24Mb) captures SureDesign enables easy custom designs SureSelect baits are compatible with all major sequencing platforms Library QC with BioAnalyzer and TapeStation Data Analysis with GeneSpring

17 SureSelect for Ion Proton Outline SureSelect TE Reagent Kit, Ion Proton SureSelect Key Benefits Workflow SureSelect Target Enrichment Performance Variant Calling

18 SureSelect for Ion Proton Workflow

19 SureSelect for Ion Proton Workflow Shearing / SPRI Size Selection Ligation /Nick Repair PCR Day 1 Hybridization (16~24hr) Capture/ Wash PCR Pooling / Size Selection Sequencing Day 2 barcode A Insert B

20 SureSelect for Ion Proton Enzymatic Shearing Fragment sizes with enzymatic shearing may not be consistent Fresh-frozen tumor tissue Cultured cells FFPE From Ion TargetSeq Exome Enrichment Protocol

21 SureSelect for Ion Proton Enzymatic Shearing Fragment sizes with enzymatic shearing may not be consistent Use double SPRI to get consistent sizes

22 SureSelect for Ion Proton Final DNA Libraries Reproducible fragment sizes Consistent sequencing results Easy index pooling after post-capture amplification 1 2 3

23 SureSelect for Ion Proton Double SPRI size selection allows for easy index pooling Reproducible fragment sizes Consistent sequencing results Easy index pooling after post-capture amplification Post-capture PCR products Pooled library + E-gel Samples can be pooled first before E-gel since they have consistent sizes. 16 E-gel lanes in 4 E-gels for 16 samples 1 E-gel lane

24 Index proportions SureSelect for Ion Proton Double SPRI size selection allows for easy index pooling 10.0% 9.0% Pooling --> E-gel 8.0% 7.0% 6.0% 5.0% 4.0% 3.0% 2.0% 1.0% 0.0% Number of index

25 SureSelect for Ion Proton Performance Evaluation NA12878 HapMap DNA Captures Evaluated Exon V5 DNA Kinome (3.2Mb) Custom design (1Mb)

26 SureSelect for Ion Proton Software Run Report

27 Coverage (%) SureSelect for Ion Proton Capture Performance Exome V5 and Kinome x 10x 20x Exon V5 Exon V5 Exon V5 Kinome Kinome Kinome Kinome Kinome Kinome Kinome Kinome Analyzed data: 4 Gb (Exon V5) and 0.3 Gb (Kinome)

28 Coverage (%) SureSelect for Ion Proton Capture Performance 1Mb Custom Design x 10x 20x Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb 1Mb Analyzed data: 0.1 Gb

29 SureSelect for Ion Proton SNP Comparison Exome V5 Hom/Het for dbsnp135 (SureSelect) HiSeq Proton n_eval_sites n_overlapping_sites n_concordant n_novel_sites concordance_rate

30 SureSelect for Ion Proton Insertions/Deletions Indels for dbsnp HiSeq Proton n_eval_sites n_overlapping_sites n_concordant n_novel_sites concordance_rate homopolymer-associated indel errors

31 BAF: B-Allele Frequency VARW: VARiation of the Width of gaps and inserts

32 SureSelect for Ion Proton Improving indel detection (Kinome) Coefficient of Variation B-Allele Frequency Total Number of Variants Percentage of reported Indels Percentage of true positive no filter no filter BAF: B-Allele Frequency VARW: VARiation of the Width of gaps and inserts

33 SureSelect for Ion Proton Summary Workflow for SureSelect for Ion Proton was simplified and improved SureSelect for Ion Proton shows similar capture perfomance as compared with SureSelect for Illumina SNP calling is comparable between Proton and HiSeq False calls for Indels can be removed by incorporating the appropriate filters

34 SureSelect for Ion Proton Summary PROVEN Technology High PERFORMANCE FLEXIBLE & COMPLETE Solution

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