IMGM Laboratories GmbH Dr. Jennifer K. Kuhn Sales Manager
About IMGM Laboratories IMGM Laboratories was founded in 2001 IMGM operates as professional provider of advanced genomic services from research projects to clinical studies Our customers come from pharma, CROs, biotech, diagnostics, and academia We have formed a strategic partnership with the Center for Human Genetics and Laboratory Medicine, Dr. Klein, Dr. Rost and Colleagues, one of Germany s leading diagnostic laboratories IMGM is accredited according to DIN EN ISO/IEC 17025 including an accreditation for expression analysis based on whole genome microarrays and quantitative real-time PCR IMGM is Certified Service Provider (CSP) for Agilent and Affymetrix microarray analysis, and for Life Technologies Ion Torrent PGM sequencing Slide 2
IMGM s Service Portfolio Biomarker Discovery RNA Services Service Portfolio f Biomarker Discovery DNA Services Metagenomics Services Pharmacogenetics Services Slide 3
Biomarker Discovery RNA Services Service Portfolio Biomarker Discovery DNA Services IMGM s Service Portfolio Metagenomics Services Pharmacogenetics Services RNA Services Small RNA / RNA isolation from diverse samples (e.g. FFPE) Small RNA / RNA quality control (Agilent Bioanalyzer Nanodrop) Quantitative real-time PCR (ViiA7, TaqMan assays + arrays, 96- and 384-well blocks) Microarrays (Agilent and Affymetrix platform) Next Generation Sequencing (Roche 454 GS FLX+, GS Junior Illumina MiSeq, GA IIx Ion Torrent PGM) Slide 4
mrna gene expression profiling The general workflow at IMGM Experimental setup IMGM/Customer Agilent Bioanalyzer Catalog/Custom Arrays Project Consulting Sample Preparation Quality Control Microarrays Interpretation of Results Hit Validation with qpcr Report Generation Bioinformatic Data Analysis TaqMan Technology Regulated Genes GeneOntology/Pathways Slide 5
Characteristics of Agilent and Affymetrix Microarrays Array Agilent Affymetrix Characteristics Technology HP inkjet Photolithography, high density Flexibility different formats, e.g. 8x60K Single arrays Catalog Arrays 30 species 22 species Customization No minimum order quantity at Minimum order quantity regular price, earray Design (40 100 arrays) Resolution ~1 probe per gene ~26 40 probes per gene Probe Length 60 25 Composition Slide 6 RefSeq Build 50, Ensemble Release 52 (Dec 2008), Unigene Build 216, GenBank (April 2009), Broad Institute Human lincrna catalog and TUCP transcripts catalog (Nov 2011) RefSeq Build 51, Ensembl Release 65 (Dec 2011), lncrna db (2011), Broad Institute Human lincrna catalog and TUCP transcripts catalog (Nov 2011) Linear dynamic range > 5 orders of magnitude < 5 logs orders of magnitude
Bioinformatic Data Analysis Normalization Correlation analysis Identification of differentially expressed genes Statistical significance Biomarker Discovery Gene signatures Functional analysis of deregulated genes PANTHER ontology (including Gene Ontology) Biological processes Molecular functions Cellular components Protein classes Pathways Slide 7
Report Generation Slide 8
Quantitative real-time PCR TaqMan technology At IMGM, we use TaqMan assays/arrays y (Applied Biosystems) by default: Highest specificity, greatest sensitivity, and largest dynamic range of any gene expression technology Wide selection of assays for numerous species: >1.2 million predesigned gene expression assays for 19 species Possibility of custom assay/array design and ordering Applicable for single samples/biomarkers up to high-throughput studies Slide 9
Further applications Determination of biodistribution, e.g. Small RNA molecules DNA plasmids Human cells injected in rodents Absolute quantification Assay design and validation Determination of quality parameters based on OECD guidelines Slide 10
Biomarker Discovery RNA Services Service Portfolio Biomarker Discovery DNA Services IMGM s Service Portfolio Metagenomics Services Pharmacogenetics Services DNA Services DNA isolation and quality control (e.g. FFPE) CNV analysis (TaqMan Copy Number Assays Agilent acgh arrays) SNP genotyping (Pyrosequencing TaqMan assays Affymetrix SNP arrays) Sanger sequencing (AB 3730XL 96 capillaries) Next Generation Sequencing (Roche 454 FLX(+) + Junior Illumina MiSeq + GA IIx Ion Torrent PGM) High-resolution HLA typing Slide 11
NGS Applications De novo sequencing (viruses, bacteria, fungi, higher h eukaryotes, plants, humans etc.) Targeted resequencing (single genes, gene panels or chromosomal regions) Whole exome resequencing Whole genome resequencing Metagenome Genome Gene expression profiling Small RNA analysis Whole transcriptome analysis Transcriptome Epigenome Methylation analysis ChIP-Seq (Chromatin immunoprecipitation seq.) Slide 12
NGS Platforms Roche 454 GS Junior Roche 454 GS FLX(+) Titanium IonTorrent PGM Illumina MiSeq Slide 13
NGS Platforms Roche 454 GS Junior Roche 454 GS FLX 10 hrs/run 10 hrs/run ~ 1x10 5 reads/run ~ 1x10 6 reads/run 400 Q20 bases Roche 400 Q20 bases Roche 35 MB/run 454 GS 400 MB/run 70 MB/day (20hr) 454 GS 800 MB/day (20hrs) IonTorrent PGM (318) 2 hrs/run ~ 5x10 6 reads/run 200 Q20 bases 1,000 MB/run 10,000 MB/day (20hr) Junior IonTorrent PGM FLX(+) Titanium Illumina MiSeq Roche 454 GS FLX+ 23 hrs/run ~ 1x10 6 reads/run 600 Q20 bases 600 MB/run Illumina MiSeq 27 hrs/run ~ 3.3x10 6 reads/run 2x150 Q30 bases 1,000 MB/run 1,000 MB/day (20hr) Slide 14
NGS Workflow Assay Development DNA Preparation Library Preparation Sequence Generation Primary Data Analysis Secondary Data Analysis Primer Design with IMGM pipeline for Roche, Illumina and LifeTech Primer Design with Illumina DesignStudio Primer Design with IonTorrent AmpliSeq Primer Validation Qiagen Macherey&Nagel etc. Target Enrichment Shotgun Library (mechanical or enzymatical fragmentation) Amplicon Library (e.g. single amplicons, AccessArray, TruSeq, AmpliSeq) Sample Indexing Roche GS Junior Roche GS FLX Roche GS FLX+ Illumina MiSeq Ion Torrent PGM Image Processing Signal Processing Contig generation Contig assembly Read Mapping SNP Calling Structural Variation Annotation ti Slide 15
RNA Services DNA Services Service Portfolio IMGM s Service Portfolio Metagenomics Services Pharmacogenetics Services Metagenomics Services Identification of bacterial communities in environmental and host associated samples such as water, soil, mucus, feces, etc. In-depth consultation for experimental setup (variable regions, coverage, sample and library preparation, etc.) Set of universal IMGM primers available Analysis of e.g. up to 6 consecutive variable regions of the 16S rrna gene in a single read possible with the Roche 454 GS FLX+ sequencing platform enabling indepth species detection Pearson Education, Inc. Slide 16
16S rrna Metagenomics Example Short reads 3 single reads per DNA molecule for 3 variant regions (V1, V2, V3) 16s variant in V1 16s variant in V2 16s variant in V3 A A A B B B C Long reads 1 combined read per DNA molecule for 3 variant regions (V1, V2, V3) A AA B A A B A B C B A 16s variant in V1 V3 Slide 17
IMGM Data Slide 18
IMGM Data The distribution of reads for the 780bp amplicon run (> Q30) Slide 19
IMGM Data The distribution of reads for the 1,050bp amplicon run (> Q30) Slide 20
RNA Services DNA Services Service Portfolio IMGM s Service Portfolio Metagenomics Services Pharmacogenetics Services Pharmacogenetics Services Using our whole spectrum of small- to high-throughput technologies, including Pharma specific applications such as DMET Plus and U133 Arrays by Affymetrix Further parameters needed for clinical studies available through one of Europe s leading Diagnostic Centers within the same building and under common leadership as IMGM In-depth consultation including gene and polymorphism selection, clinical protocol, informed consent, appropriate technical approach, result interpretation, etc. Analytical protocol and report (GCP compliant, e.g. customizable data formats) Rigorous QA program Slide 21
Summary IMGM - YOUR flexible and reliable partner Longstanding experience in biomarker discovery Expert in unique metagenomics NGS sequencing Applying our knowledge to pharmacogenetic services Slide 22
Summary IMGM - YOUR flexible and reliable partner Personal project manager, direct communication In-depth project consulting Broad range of up-to-date methods and technologies Delivery of detailed and comprehensive reports fitting your needs Professional quality management system (DAkkS) Working according to GxP Guidelines Professional business relationship (e.g. NDAs, FSAs, TATs) Slide 23
IMGM Team Dr. Hanns-Georg Klein CEO Dr. Ralph Oehlmann Director Business Development & Marketing Dr. Jennifer Kuhn Sales Manager Stefan Kotschote Head of Laboratory Dr. Michael Haberl Head of Quality Assurance??? Anja Weber??? Marketing Assistant Sales Assistant Dr. Dirk Haase Project Manager IT Sabine Brauer Deputy Head of Laboratory Dr. Marion Deuter Head of New Technologies and Product Development Dr. Cornelia Graf Project Manager Dr. Ovidiu Rücker Project Manager Dr. Alexandra Dangel Project Manager Melanie Sohns Head Lab Assistant Andrea Matu Laboratory Assistant Johannes Tank Laboratory Assistant Simone Hardel Assistant New Technologies Slide 24