1. The History of DNA Sequencing Technology

2. ARC Biotechnology Platform
Dr Jasper Rees

3. Dr Jasper Rees ReesJ@arc.agric.za
ARC Biotechnology Platform In 2010, the ARC set out to create a new single integrated platform for advanced high throughput, next generation technologies
Dr Jasper Rees

4. Vision
To create a world class biotechnology platform to lead research in agriculture in Africa

5. Mission
To provide excellent biotechnology research and service platforms to enhance food security and promote agriculture in Africa
To provide biotechnology tools in support of all areas of research and development within the mission of the ARC
To create a world class research and innovation environment to attract the best researchers and students
To train the next generation of specialist researchers in cutting edge biotechnologies

6. The Objectives
To create Service, Development and Research structures in support of and in collaboration with all ARC Institutes and external partners
To address the needs of commercial and emerging agriculture nationally and continentally
To train new researchers and postgraduates at the highest level
To compete at the top level internationally
To have the “best in class” resources for modern biotechnology
To raise funds external funding, nationally and internationally

7. Service and Research Facilities Model
Core Services
Development group
Research teams within platform
Unit
Research teams at Institutes
Technology Focus of Unit
Key technologies being performed with the sub-platform as services to research groups and pipelines with the ARC and to outside clients
Each sub-platform to have a development group responsible for introducing new technology and applications into the core services
Internal Research teams, lead by SR, SpR, PR level scientists. With MSc, PhD and PD level students as the major part of each team. Primarily externally funded with competitive grants
External Research teams, led by SR, SpR, PR level scientists. MSc, PhD and PD level students are the major part of each team. Primarily externally funded with competitive grants

8. Biotech Platform Phase I – 2010/11
DNA Isolation
Next Gen Sequencing
Genotyping
Genetic Mapping
Development group
Research teams within platform
Genomics
Sample Preparation
2D PAGE
MALDI-MS
LC/GC-MS-MS
Proteomics
Databases
Annotation
Assembly
Expression Analysis
Bioinformatics
Tissue Culture
Construct development
Transformation
Regeneration
Plant Transformation

9. Biotech Platform Phase II/III: 2012-2015
Marker Assisted Breeding
Plant Phenotyping
Genome Engineering
Functional Genomics
Robotic platform
High throughput imaging and sensing
GC MS/MS
Informatics
Reverse Genetics
Gene identification
Vector Development
Construct development
Robotic platform
High throughput DNA Isolation
Genotyping
Informatics
Mutation Breeding
Gene Silencing
Zinc Finger Nucleases
Gene targeting
Male Sterile Technology
Targeted Mutagenesis

10. Platforms and Programs
Drought Tolerant Sorghum
Apple Breeding
Vaccine Development
Animal Reproductive Biotechnologies
Diagnostics
Genomics 1 2
Proteomics
Bioinformatics
Plant Transformation 3
Functional Genomics
Genome Engineering 4
Imaging
NanoBiotechnology
Protein Expression

11. ARC Biotech Platform Implementation 2010/11
Staff recruitment – 6 researchers team leaders in genomics, proteomics, bioinformatics; 3 more posts in 2011, more in the future.
>25 postgraduate and postdoctoral researchers already
Capital equipment procurement: >R10m to date
Sites in Pretoria and Stellenbosch
Building renovations in progress
Operational start: Jan 2011

12. ARC Biotech Platform Implementation 2010/11
Staff recruitment – 6 researchers team leaders in genomics, proteomics, bioinformatics; 3 more posts in 2011
Jasper Rees: Genomics and Bioinformatics: Fruit
Bongani Ndimba: Proteomics: Cereals and Fruit
Damaris Odeny; Genetics, Genomics: Vegetables
Joseph Mafofo: Genomics, plant pathogens: Fruit
Dirk Swanevelder: Genomics: Plant Pests, Cereals
Farai Muchadeyi: Genotyping: Animal Genetics

13. Genomics Applications
Sequencing applications
SNP applications
Genomes,
De novo and re-sequencing
SNPs and CNV calling
Transcriptomes,
SNPs and splicing variation
Expression profiling
Small RNA
discovery and expression analysis
GWAS
Association Genetics
SNP validation
Candidate Genes
Diversity studies

14. Platform Synergy for Agrigenomics Research
Discovery
Focused Research/Validation
Screening
>109
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106
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Whole Genome
Single Marker
Discover
Interrogate
Validate/Screen
Platform
HiSeq 2000
iScan/HiScan SQ
BeadXpress ►
De novo Genome Sequencing
mRNA Sequencing & Gene Expression
SNP Discovery
DNA Methylation, Targeted Resequencing ►
Genome-wide marker / phenotype association
Focused marker / phenotype association
Genetic Identity Confirmation
QTL Mapping ►
Genetic Purity Screening
Illumina is a market leading company providing tools for genetic analysis –
keen to ensure develop and provide correct tools to support Agrigenomics research
Genome wide discovery studies supported by the the GA,
genome wide validation and association studies can be carried out on the iScan where more than one million markers can be assessed per sample in a single assay. The iScan is also suited to conduct more focussed studies and screening applications in lower plex assays.
The BX platform is geared to screening studies in which high sample throughput rates are required with lower assay multiplexing.
Thus Illumina products cover a wide range of genotyping expt designs right from whole genome sequencing through to low multiplex genotyping. ►
Parentage / Lineage
Applications ►
Marker Assisted Selection ►
Disease Screening

15. Illumina HiScanSQ Platform
Dual function system
SNPs and Sequencing
High throughput SNPs
Infinium and GoldenGate technology
10k to 2.5M SNP chips
Next Generation Sequencing
Linear scanning optics
100 Gb of DNA sequence per run
Paired end sequencing
400 M filtered clusters per flow cell
8 lanes per flow cell
Sample Multiplexing

16. Illumina Sequencing Technology5’ 3’ G T C A
DNA ( ug) T G C A G T
Sample preparation
Cluster growth
Single molecule array
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Avitage Item ID: {{C2A4BBD5-93E1-4B59-A5D0-E38C126BE098}}
Sequencing 1 2 3 7 8 9 4 5 6
Base calling
T G C T A C G A T …
Image acquisition

17. What if, in one sequencing run you could…
Sequence one cancer & one normal genome
At 30x coverage
SIMULTANEOUSLY
Run multiple applications requiring different read lengths
Unravel 20 whole transcriptomes
In four days
Analyze two human methylomes
In one week
Whole genome sequencing
Profile 200 gene expression samples
In less than two days
One Sequencing Run
Targeted resequencing
Gene expression
Methylation
The goal of this slide is to capture the power and scope of research that will be made possible with HiSeq 2000
De novo
Metagenomics
ChIP-seq
Whole transcriptome

18. Focused Genotyping Infinium BeadChips HumanCVD BeadChip
BovineSNP50 BeadChip
CanineSNP20 BeadChip
CanineHD BeadChip
EquineSNP50 BeadChip
OvineSNP50 BeadChip
PorcineSNP60 BeadChip
MaizeSNP50 BeadChip
iSelect Custom Panel

19. Custom Genotyping Infinium iSelect HD
Interrogate virtually any SNP for any species
24-sample format (3,000 to 68,000 attempted bead types)
12-sample format (68,001 to 200,000 attempted bead types)
Custom GoldenGate Genotyping Panels
Create assays tailored directly to their specific needs for targeted region genotyping or fine-mapping of candidate disease association regions.
Interrogate 96, or from 384 to 1,536, SNP loci simultaneously
GoldenGate Indexing v1 Kit
Screen up to 16 times as many samples per reaction as the standard GoldenGate Assay
Increase throughput from 288 samples per day to greater than 2000, while decreasing total reagent consumption.
Current plexity ranges for GoldenGate Indexing include 96-plex, 192-plex, and 384-plex.
The GoldenGate Indexing Assay is conveniently processed on the Universal-32 BeadChip

20. The Rosaceae – Fruits, Nuts and Flowers

21. Rosaceae BeadChips Roscaeae BeadChip
SA and NZ working in collaboration with RosBREED
Using around 9,000 SNPs per species
Apple, Peach, Cherry, Strawberry
SNPs are from founders of major breeding programs and novel cultivars
Optimised for cultivated populations as opposed to wild crosses
Available March 2011
Maize is $195 per sample

22. Customize HiScanSQ To Meet Your Needs – Now or Later
SPLs: Samples HumanOmniExpress 12 sample BeadChips. Throughput varies by BeadChip
For illustration purposes only – consult your rep to discuss your expected sample throughput.
Assumptions:
6 Gb/day sequencing output.
Manual sample processing: 1 HiScan, 0 Autoloader2.x, 0 Tecan liquid handling robots, 5 day work week; 2 FTEs. Automated sample processing: 5 day work week; 24 hour scanning powered by 1 AutoLoader2.x, 1 Tecan liquid handling robot.

23. Predictions
Cost, Complexity and Complementary Solutions
Cost per data point will continue dramatic reduction for both Arrays and Sequencing
Sequencing output will scale quickly
Array content and complexity will increase significantly
Sample numbers for projects will grow
Sequencing and array solutions will continue to co-exist and complement

24. ARC Biotech Platform Implementation The First Genomes
First genome sequencing: March 2011
Apple, pear, aphid, tick, amaranth, heartwater, avian paramyxovirus
(15 apple genomes resequenced in 2010)
First major genotyping experiments, from May 2011
Apple (RosBREED - USA)
Peach (ARC)
Cattle (ARC)
Sheep (US, ARC)

25. Agric-Biotech Smart Tools
Rapid characterization of novel genomes (plants, animals, pathogens)
Gene expression and genotyping technologies
Molecular Breeding for rapid crop and animal improvement:
Genome-wide selection
Marker assisted selection
Tissue Culture and micro-propagation of plants
Cis-genesis and Genome Engineering
Reproductive biotechnologies
Vaccines, therapeutic proteins, diagnostics

26. Impact on Agricultural Research and Development
Faster, better, more selective and more sensitive:
Plant Breeding
Animal Breeding
Cultivar identification (PBR)
Pathogen identification
Diagnostics
Vaccine development
Vaccine quality control and safety
Bringing better crops and animals to farmer at all scales
Bringing novel, cash generating, sustainable products to the market
Increasing food and income security

27. ARC: BP – the 5 year horizon
By 2015, The ARC Biotech Centre will be a vibrant Agri-Biotech institution – a Centre of Excellence in research, training and services with National and International standing
Regional integration of services and research
Training of new generation of researchers and postgraduates
Implementation of high throughput systems for applied and basic research
Increased funding and collaborative opportunities

28. Dr Jasper Rees ReesJ@arc.agric.za
Thank you
Dr Jasper Rees