TBRT Meeting April 2018 Scott Weigel Sales Director

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1 TBRT Meeting April 2018 Scott Weigel Sales Director

2 About Us AgriPlex Genomics was formed in 2014 with the goal of creating a platform for targeted sequencing and genotyping in large numbers of samples. We currently occupy laboratories in the BioEnterprise Building in Cleveland, Ohio. We specialize in focused, simultaneous analysis of limited numbers of loci ( ) in any number of DNA samples in support of animal and plant breeders. The multiplexed Next-Gen PlexSeq platform will greatly improve throughput and efficiency while significantly reducing costs. AgriPlex Genomics partners with plant and animal breeders and researchers to identify desirable traits, leading to healthier and more productive crops and livestock.

3 PlexSeq our core technology - Focuced amplicon based sequencing method - Multiplex 10 to 3000 SNP s together - Barcoding of samples 20k+ samples - PlexCall software automates allele based frequency genotyping

4 Our Role in the Market WHERE WE COMPETE: PlexSeq technology provides much higher sequencing coverage for SNP s of interest in a large number of samples rapidly and inexpensively. Alternative to current Single-Plex technologies in large number of samples. No ligation No restriction digestion - No complicated library preparation No individual normalization Sequence PCR products directly. WHERE WE DON T: PlexSeq technology does not compete with whole genome genotyping by sequencing approaches such as Restriction Enzyme assisted GBS. PlexSeq technology does not compete with very large number SNP chip genotyping platforms.

5 Low Cost PlexSeq Features Flexible run any number of SNP s with any number of samples Accurate NGS data vs. fluorescence based assays Small amounts of DNA needed 4ng Works with crude extractions (NaOH, hot-shot ) High Assay design success rate 97% Works in polyploid species

6 Process, Reagents and Reaction Setup

7 PlexSeq Workflow 1.5 ul Assay Specific 2X PCR mix 1.5 ul 384 Amplicon MIX reactions 384 DNA Samples PCR

8 PlexSeq Workflow 1x BARCODING MIX (384 individual mixes) 3 ul Barcoding Reaction MIX PCR 100 nl automated transfer (disposable pin tool) Mix All Wells Purify and Quantitate 384 Amplicon MIX PCR Products NGS

9 PlexSeq Workflow Primer and Multiplex design reaction setup PCR Barcoding Reaction setup PlexCall Automated LIMS Compatible Report Illumina Sequencing Run Pool, Purify, Quantify Barcoding PCR

10 Project Specific PlexCall Analysis Software

11 PlexCall : Quantitative Analysis Personalized to your project and specific SNP s Automated allele frequency based genotyping calling directly from sequencer output Quantitative analysis using PlexCall provides exact allele ratios without cluster plot analysis. No more Cluster Plots

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14 Allele Ratio Output

15 Genotype Call Output

16 PlexSeq Assay Chicken Project Assay Statistics PlexSeq Assay SNP Performance Chicken Project SAMPLES Submitted: SNP s Submitted: SNPs Successfully Validated Number of Genotypes Average Success Rate per SNP , % Failed Validation 0% Failures 0-1% Failures 1-5% Failures 5-10% Failures >10% Failures

17 AgriPlex Data from Chicken Project

18 AgriPlex Throughput CAPACITY 100 SNP s in 250,000 Samples 25 Million Genotypes One Week including DNA Purification

19 Automation Capacity of 100,000 samples per day 2.5 days for project Purification

20 5 Hydrocyclers (3x32, 2x16) 128x384 plates - 4 times per day 200,000 PCR reactions per day 100,000 samples per day (primary and barcoding) 10 million genotypes per day 2.5 days for project PCR

21 HiSeq bp Single Read High Output 2 chips 4 billion reads 150x coverage 25 million genotypes 40 hours 2 days for project Sequencing

22 PlexCall 1 day for project Analysis

23 Plasticware Savings Plates Tips

24 Experience Plant Corn Soybean Tomato Squash Cotton Wheat Sunflower Grape Cowpea Chrysanthemum Eucalyptus Pine Flax Sesame Animal/Fish Chicken Pig Bovine Deer Salmon

25 Summary The flexibility of PlexSeq allows you to pick targets and supply as many samples as you need. The multiplex capabilities of PlexSeq provides data at a much lower cost and time than current platforms. Eliminate Cluster Plots and Analysis Bottlenecks using PlexCall Phone info@agriplexgenomics.com