A Decade of Microarrays ACGT Microarray Facility Nicky

A Decade of Microarrays ACGT Microarray Facility Nicky Olivier 1

In the beginning ● ● ● Institutions involved: ● UP & CSIR – Shared start-up costs Physical location: Agriculture Building ● Special installation required Department Plant Science and FABI Equipment - June 2001 ● Setting up and validation Core service: July 2002 2

People • Director – Prof Dave Berger • Scientific Officers – Danie Theron (2001 -2006) – Nicky Olivier (2006 -present) • Statisticians – Loveness Dzikiti – Nanette Coetzer 3

Advisors Prof Jane Morris (ACGT) Prof Henk Huismans (UP) • Equipment and hardware acquisitions • Approval of projects • Approval Annual Financial Statements • Future plans for Facility 4

Start-up Instrumentation 2001 Molecular Dynamics Gen. III Spotter Genepix 4000 B Scanner 532 nm / 635 nm 5

Instrumentation 2008 • Agilent Hybridization oven and chambers Multiplexing slides 1 x 244 K, 2 x 105 K, 4 x 44 K, 8 x 15 K 6

Instrumentation 2009 • Bio-Rad Automated Electrophoresis system Quantification of RNA quality/concentration RQI score for consistency 7

BIOPAD project (2005 -2007) • Microarray Capacity Building project – UP, Inqaba Biotec, WITS • Focus areas – Training • Workshops • Inqaba intern project – Standard Operating Procedures – External support for projects 8

Workshops - BIOPAD • Lab training (2005 & 2006) Region Attendees ACGT 22 Gauteng 10 SA 9 9

Workshops – Starter kit • Scaled down experiment (3 days) Region Attendees ACGT 9 Gauteng 4 SA 3 10

Workshops • ICTTD & Veterinary Science (2003) Region Attendees ACGT 12 Gauteng 3 SA 3 Outside SA 6 11

Workshops - Annual • Data analysis – Genetics & Bioinformatics Hons – Interested researchers/students • NBN – Data analysis for Bioinformatics students 12

Services – Project planning • Experimental design – Statistical relevance • Choice of slides/platform • Oligonucleotide design 13

Services – Sample QC • RNA/DNA isolation • Experion – RNA analyses – microarray, q. PCR, RNAseq – DNA analyses – HTS • Evaluation of external Bioanalyzer results • Evaluation of RNA gels 14

Services – Hybridization • Lab space and tech support • Hyb-up chambers in waterbath – c. DNA, oligo, g. DNA • Agilent hyb oven 15

Services – Scanning • Spotted arrays – resolution 10 µm • Agilent arrays – resolution 5 µm • Single channel or dual channel scans • Data backup at Facility • Data capture assistance 16

Services – Data analysis • Results file generation and QC • Data normalization • Statistical analysis + output • Downstream protocols • Annotation assistance • GEO/Array. Express submission assistance 17

Software • Spotverify – Verification of differential gene expression • SSHscreen – SSH library quality evaluation • SSH-DB – Moderated database for sequences, BLAST • MADIBA – Annotation, biological function etc. 18

Communication • Lectures to UP undergraduate students • Lectures to UP postgraduate students • Lectures to external undergraduate students • Lectures to external postgraduate students • Discussions and demos for visitors 19

Operating philosophy • Non-profit • No hybridisation service offered – Student/Researcher training – Increase capacity in techniques – Make microarray technology accessible 20

Costing model • Not full-cost – Scientific Officer salary from UP • Departmental responsibilities • Reagents at cost price • Data analysis with free software – Scripts given to researchers for further use 21

Costing Spotting Scanning (per slide) Data analysis ACGT R 450 - R 550 Free Regional R 450 - R 550 R 180 Free External R 600 – R 750 R 350/h Region 22

Financial Administration • Funds in UP cost point • Yearly financial statements to advisors • UP users: – Internal fund transfers • External users – Invoice issued by UP – Payments administered by UP Finance • Excess funds for instrument maintenance 23

Projects by region Region Past Current Future ACGT 17 4 8 Gauteng 4 1 5 SA 4 1 0 Africa 1 0 2 24

Slides to date Number Slides printed 1970 Slides scanned 2261 25

Student training Degree Number Hons 3 MSc 11 Ph. D 17 Post-Doctoral 6 Principal Investigators 7 26

Publications • 14 Publications in peer review journals • 1 Book chapter • 3 Hons reports • 8 MSc theses • 11 Ph. D theses 27

Challenges / Opportunities • • • Spotter printing pins Spotted arrays becoming obsolete Scanner laser service Scanner resolution RNAseq • Agilent slides becoming more accessible – Partnership with supplier – greater awareness 28

The Future • New Plant Science building – Dedicated labs • RNA isolation, Hybridization, Scanning • Equipment needed – High resolution Agilent scanner • 2 µm scanning resolution – high density • 1 x 1 M, 2 x 400 K, 4 x 180 K, 8 x 60 K 29

Thank you 30