High Cross-Platform Genotyping Concordance of Axiom High-Density Microarrays and Eureka Low-Density Targeted NGS Assays
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1 High Cross-Platform Genotyping Concordance of Axiom High-Density Microarrays and Eureka Low-Density Targeted NGS Assays Ali Pirani and Mohini A Patil ISAG July 2017 The world leader in serving science
2 Overview of Thermo Fisher Agrigenomic Genotyping Solutions Number of markers Gene and marker discovery Marker verification, trait mapping and gene expression Genomic selection Production & QC Parentage Millions M 10,000-5,000 <100 <100 Technology NGS Microarrays Genotyping by Sequencing STR analysis on capillary electrophoresis (CE) qpcr Products Ion Torrent technologies Axiom genotyping arrays Eureka & AgriSeq targeted GBS Taqman Assays and master mixes Animal genotype kits on genetic analyzers 2
3 Axiom Genotyping Solution overview SNPs Indels CNVs Pre-verified content from Axiom Genomic Database or other commercial products Axiom 24, 96 (~850,000 SNPs), or 384 (~50,000 SNPs) Customer s content from SNP discovery sequencing Axiom Reagent Kit Robust and reliable assay Target prep Automated and manual protocols GeneTitan MC Instrument Automated, hands-free array processing Axiom TM Analysis Suite Automated genotyping 3
4 Eureka Genotyping Workflow Day 1 Day 2 4 X 96 Samples Denature Quantify and QC libraries Eureka Genotyping Affordable targeted solution Ligation Flexible SNP panels Amplification Scalable throughput Analysis pipeline, QC, and visualization Probe hybridization Pool, clean, and concentrate amplicons Sequence libraries Genotypes ready 4
5 Automated Analysis Pipeline Automated Target Preparation Hands-free Automated Array Processing Automated data Transfer to server Axiom Analysis Suite Results Export using AxLE or CDCB Axiom Analysis Suite Windows GUI Export to Text (Forward strand) VCF PLINK (Forward strand) Array Power Tools 32/64 bit Command line Windows, Linux, MacOS-X AxLE Tool Export tool for long format CDCB Export tool Array Power Tools (APT) Streamlined Command line control. Output is text files Multiple computer cluster parallel job processing Scripting control to schedule jobs 5
6 Benefits of Axiom Long Format Export Tool Reduce the need to change existing bioinformatics pipelines Call for standardization by International Society of Animal Genetics (ISAG) Standardizing on top strand (versus bottom strand) format as the international nomenclature for ISAG SNP parentage markers Launch directly from Applied Biosystems Axiom Analysis Suite software 6
7 Automated Software Powerful and simple Simplified workflow Advanced visualization features Intuitive and easy to use user interface Simple, Easy, Integrated Load CEL data Get genotype results Load data 7
8 Experimental Design and Results Axiom Array and Eureka Panel Axiom 15K Bovine Array Eureka Bovine Parentage Panel 117 markers in common between platforms Samples 92 Bovine samples Results Average sample call rate for Axiom array = 99.4% Average sample call rate for Eureka NGS panel = 99.9% Relative concordance = 99.8% 9
9 Cluster Plots Same SNPs across platforms (1/2) Axiom Data Eureka Data 10
10 Cluster Plots Same SNPs across platforms (2/2) Axiom Data Eureka Data 11
11 Applied Biosystems Axiom Bovine Genotyping Array 50K Fast, economical, standardized genotyping tool for bovine breeders to screen for valuable genetic traits and perform parentage verification >50,000 markers PN format PN Mini 96 format 12
12 Axiom Bovine 384 array Single array with 50,000 markers covering Core markers VIP trait markers Parentage markers Y-Chromosomal markers Standardized genotyping platform across breeders Compatible with legacy bioinformatics pipelines Compatible with Council on Dairy Cattle Breeding submissions Available: Low /$20s per sample (inclusive of array processing) 13
13 Axiom Bovine Array Chromosomal Coverage 51,987 markers evenly spaced across the bovine genome 1213 markers on Y chromosome, 1063 markers on X chromosome # Markers MT X Y Chromosome 14
14 Automated Analysis of Y-Chromosome Simple Fertility Trait ID 15
15 Conclusions We have developed a method that generates highly concordant genotypes across both technologies Axiom and Eureka Researchers can screen a large number of variants on an array, and then select a small set of informative variants for routine application as a panel Researchers can seamlessly compare genotype results when studying multi-breed genomic selection on a microarray and parentage on a NGS assay 16
16 Thank you! For Research Use Only. Not for use in diagnostic procedures. 17
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