High Impact Research Coordinator

PhD, MSc, LLM(Merit), BSc, LLB, CBiol, MIBiol

High Impact Research Building, Level 1

ISB (Genetics & Molecular Biology), Faculty of Science, University of Malaya

50603 Kuala Lumpur

Email: kokgan@um.edu.my

DR KOK GAN CHAN PhD, MSc, LLM(Merit), BSc, LLB, CBiol, MIBiol.

HIR Central Lab High End Instrument

HIR Central Labs

HIR Building Ground, 1st & 2nd floors

The rationale

• To equip UM-HIR grantees with state-of-the-art instrument

• To support high impact research funded by HIR

• To promote high quality research To enable researchers to obtain high quality data

• To promote UM-HIR to overseas collaborators

• To establish UM-HIR as nucleus for high impact research, regionally and internationally

HIR Priority Areas

1. Genomics

2. Proteomics

3. Analytical chemistry

4. Metabolomics

5. Bioinformatics

Genomics • HiSeq2500, G floor

• Whole-Genome Sequencing: sequence

a whole human genome in a day (2x100bp

reads). Or, in high output mode &

sequence multiple genomes in a single run (2x150bp reads) 176 Gb in ~40h including on board cluster generation and sequencing, with 90.2% of bases at or above Q30, high quality alignment and variant calling.

• Targeted Resequencing: individual custom panels to whole exomes

• Whole-Transcriptome Sequencing: gene expression profiling to deep transcriptional analysis, HiSeq offers unprecedented RNA analysis solutions. Measure abundance of individual transcripts and isoforms, discover new genes, and identify regulatory non-coding RNAs.

• De Novo Sequencing: de novo sequencing of any size genome; NGS of metagenomic analyses

• Epigenetic Regulation: Study DNA-protein interactions and gene regulation using ChIP-Seq, and quantify DNA methylation with single base resolution.

• PacBio SMRT, G floor The PacBio RS II sequencing technology resolves single molecules in real time, allowing observation of structural and cell type variation not accessible with other technologies. De Novo assembly: extra-long read lengths that simplify and improve genome assembly. • Highest N50 • Fewest contigs • Detect structural variation • 99.999% accuracy • Genome finishing at 1/10th the cost Targeted sequencing: common use of targeted sequencing is single nucleotide polymorphism (SNP) detection and validation.

SMRT directly measures individual molecules, using long reads to fully characterize genetic complexity, including rare SNPs, indels, structural variants, haplotypes and phasing. Single molecule resolution allows comprehensive characterization of heterogeneous samples and identification of variation invisible to multi-molecule sequencing technologies. • Benefits: • Compound Mutations and Haplotype Phasing - Multi-kilobase reads facilitate the study of linked mutations

hundreds, even thousands, of bases apart. • Repeat Expansions - Long reads and low bias allow accurate sequencing across repeat expansions, even in low

complexity regions. • Full-Length Transcripts and Splice Variants - Single-molecule resolution and long reads span entire cDNAs, allowing

full characterization of splicing in the transcriptome. • Minor Variants and Quasispecies - Single molecule sequencing simplifies the analysis of mixed populations of

sequences. Exquisitely sensitive and specific. • SNP Detection and Validation - Single molecule sequencing detects and validates SNPs with high accuracy by

avoiding mapping errors and systematic errors.

Base Modification Detection: DNA base modifications are important to the understanding of biological processes such as gene expression, host-pathogen interactions, DNA damage, and DNA repair. (SMRT) Analysis software works by measuring the rate of DNA base incorporation during sequencing. This information, automatically gathered during the sequencing process, is a unique feature of our SMRT technology. Resolve strand-specific modifications - unamplified double-stranded input DNA enables detection of strand-specific modifications such as hemimethylation Single-base resolution - visualize a wide variety of base modifications in high-resolution Integrated sequence and chemical modification detection - epigenetic and other base modification data are gathered in the sequencing process, as an integral part of the sequencing workflow Hypothesis-free detection - no prior knowledge of the modification is needed, allowing discovery of unknown or unexpected modification

Other genomics facility

• DNA Microarray • Nanostring • qPCR (Quant Studio ABI, BioRad) • digital droplet PCR • Pyrosequencer • Luminex • Oxford Nanopore: NGS of >50kb?!?! • More NGS in 2013 • Liquid handling robot, DNA and library prep workflow

Single cell analysis and NGS

• Microfludic single cell analysis enabling profile of multiple single cells for the expression of hundreds of genes in just a few hours, using standard reagents and easy-to-use analysis tools enable hundreds of individual cells to be tested for the expression of hundreds of genes in a few hours. Advantages: • High Sample Throughput—Screen up to 96 single cells across 96

assays in one array. • Produce More Results Faster—Produce 9,216 data points in just

four hours with one array. • Assay Flexibility—Practice tried-and-true techniques such as

TaqMan assays or other common assays.

Microfludic single cell analysis

• Enabling: Measure expression levels of genes, alleles and spliced variants

• Compare tissue or cell type expression levels

• Map transcription initiation sites

• Characterize alternate splicing patterns

• Evaluate post-transcriptional activity

• Discover new transcripts and gene fusions

Proteomics

• 1D & 2D gel, Western blot, ELIZA • FPLC • Typhoon system • Isothermal Titration Calorimeter (ITC): GE Healthcare’s MicroCal (ITC) allows characterization of biomolecular interactions by

providing direct, label-free measurements of binding affinity and thermodynamics. In a single ITC experiment, you can determine binding affinity (KD), stoichiometry or

number of binding sites, enthalpy (ΔH), and entropy (ΔS). Thermodynamic data, specifically ΔH and ΔS, reveal the forces that drive complex formation and mechanism of action. Thermodynamic measurements provide information on conformational changes, hydrogen bonding, hydrophobic interactions, and charge-charge interactions.

provide valuable information about binding interactions, molecular stability, protein folding, and enzyme kinetics.

• Surface Plasmon Resonance

Mass spectrometry instrument

• UHPLC • GC • GC-MIDI • GC-MS • GC-Q-TOF • LC-MSMS • LC-Q-TOF • Orbitrap mass spec (proteomics, lipidomics,

metabolomics) • LC-NMR-MS

Orbitrap mass spec • protein and metabolite identification,

characterization and quantitation.

Chromatography, Magnetic Resonance & Mass Spectrometry

• Unequivocal molecular structures

confirmation via exact mass of the analyzed

sample and access to the sum formula

• ideal for sensitive and instable compounds, fully automated analysis, or

provide a very

convenient ,

quick path from

LC separation to

NMR.

• UV-VIS Spectrometry: Agilent Cary 60

• Luminometer: fluorescence,

luminecence, visible light

• Atomic emission spectrometer

• Flow cytometer

• Chemostat fermenter

OmniLog

• Bacteria, Yeast & Fungi ID

• Microbial Community Analysis: Biolog EcoPlates: provide a sensitive and reliable index of

environmental change.

•Measure the metabolism of 31 carbon sources per assay •Each assay panel tests in triplicate •Simple colorimetric readout •Readable with any microplate reader

• Phenotype MicroArrays for Microbial Cells 1. make it possible to quantitatively measure thousands of cellular

phenotypes all at once. 2. Correlate genotypes with phenotypes 3. Determine a cell's metabolic and chemical sensitivity properties 4. Discover new targets for antimicrobial compounds. 5. Optimize cell lines and culture conditions in bioprocess development. 6. Characterize cell phenotypes for taxonomic or epidemiological studies Important application: • Testing Cell Lines Exposed to Drugs or Other Chemicals • Evaluating New Drug Candidates • Toxicological Testing • Testing Cell Lines with Genetic Differences • Determining Functions of Important Genes (Functional Genomics) • Improvement of Cell Lines • Cell Cultures for Phenotypic Stability • Direct Testing of Cell Lines

• Phenotype MicroArrays for Mammalian Cells (PMM)

• PMM assays are cell-based assays provided in five categories and used to determine up to 1,400 metabolic and chemical sensitivity phenotypes of mammalian cells.

• Monitor the stability of cell lines

used in research.

• Use PMs as a tool to

understand gene function.

• Perform other comparisons

of cell lines in basic research

and drug target studies.

• Use PMs as a tool to test drug leads

• Improve Bio Processes with PMs

• Perform cell proliferation and chemosensitivity assays

• Photon camera

• EM-CCD technology performs best in low light conditions. How low?

<10 photons per pixel

Luminescence imaging, single

fluorescent molecule in cells

expressing fusion protein, confocal

calcium ion imaging

• Capillary electrophoresis • Nanodrop • Bioanalyser • Qubit

Bioinformatics

• Central Bioinformatics lab

• High speed server and high performance cluster computers

• 11,000 Human Genomes

• 13.3 mil bacteria genomes

Use of HIR Central Instrument

• No human intervention: all booking online

• Nominal charges for rental esp for UM users

• Packages for high volume users

Why HIR Central Equipment?

Proof of Using HIR Central Instrument to increase High Impact Publication

Based on true story…

from myself

On This Day in History

University of Malaya The Premier University in Malaysia, >100 yrs

Instrument Before HIR

Dr Kok Gan CHAN

Thank you