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DNA Services Overview

November 20, 2012

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Sequencing & Analysis Services at a glance

WHAT WE CAN DO . . .

      Illumina or Agilent library preparation Clustered flowcell generation Paired end and single end sequencing 2x100  Whole genome   Adapted libraries Prepared flowcells Target capture by hybridization  Agilent SureSelect exomes  Agilent SureSelect custom targeted Illumina TruSeq Amplicon-Cancer panel* Data Analysis *available January 2013

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Sequencing & Analysis Services at a glance

WHAT WE DO NOT CURRENTLY OFFER . . .

     In house DNA or RNA extraction - Rutgers In house RNA to cDNA conversion Specialized/custom sample prep – chip / methyl Long-read sequencing De novo assembly – will allign to provided reference

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Sequencing & Analysis Services at PKI

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Product Offering High Throughput Sample Prep

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Automated library prep for sequencing

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Automated target capture by hybridization

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Strict SOP-driven processes

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Extensive quality checking

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CT licensed clinical lab (No. CL-0685)

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CLIA registered ( ID# 07D2034530)

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Project consulting services

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Sequencing Apps Exome re-sequencing Targeted re-sequencing Microbial re-sequencing Prepared library re-sequencing Whole genome re-sequencing (on demand) RNA Clinical sequencing Operational Infrastructure

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4—6 week turnaround

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LIMS for sample tracking 3 HiSeqs + 1 MiSeq

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> 6000 samples successfully processed to date Secure private cloud in sequencing center; semi-public cloud hosted by PerkinElmer Informatics

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Current analysis capacity of 50-60 exomes per day—highly scaleable computing infrastructure

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Employing Proven Technologies & World-Class Automation…

PerkinElmer is focused on bringing best in class technologies

– we continually evaluate new technologies to ensure we are providing the best available systems & processes to our customers

CURRENT PLATFORMS SAMPLE & PROCESS TRACKING

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PerkinElmer Informatics LIMS: providing sample tracking and status updates AUTOMATION

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PKI JANUS Automated Workstations

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PKI EnSpire Plate Reader

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Caliper Labchip GXII TARGETED CAPTURE

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Agilent SureSelect SEQUENCING

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Illumina HiSeq 2000 system

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Illumina MiSeq ANALYSIS

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PerkinElmer Informatics Analysis Edition: private cloud implementation

…ensures faster turn-around-times and high quality control

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Workflow flexibility to accommodate customer needs

DNA Library preparation

Prepared by us

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Libraries

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Captures

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Flowcells

Target Enrichment [Optional] Automated Amplification (Clustering)

Prepared by customer

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Libraries

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Captures

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Flowcells

Automated Sequencing Automated Data Analysis

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Illumina HiSeq 2000-Overview

Variable read lengths (50-100bp) Sequence in one or two directions

Up to 2.8 x10 9 reads per flow cell (280 Gb)

Run time is variable    1 x 50bp =~ 2.5 days 1 x 100bp = ~6 days 2 x 100bp = ~12 days Output deliverables can vary depending on project requirements

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Sample Submission

Each sample submission needs to be accompanied by sample manifest   Required DNA concentration is ≥25 ng/µL Required concentration for ready-to-sequence libraries is ≥ 10nM  Required amount of SureSelect XT library for capture ≥ 700ng When submitting, send shipping notification to Seqproject @ perkinelmer.com along with electronic copy of shipping manifest and tracking number Samples should be shipped overnight via express shipping Follow instructions contained in “Sample Prep and Shipping Guidelines_v3.2” for sample purity, concentration, etc.

Not following instructions will delay the project Include hardcopy of sample manifest with samples

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Library Preparation Guidelines

We use either Illumina TruSeq or Agilent SureSelect for library prep  Currently evaluating Nextera, but not yet in production  We do not use NuGen kits Required for everything but sequencing of PCR products  Whole genome resequencing   Targeted resequencing RNA-Seq Minimal sample requirements (as per kit providers protocols)  1 µg purified DNA for TruSeq libraries   3 µg for purified DNA for SureSelect libraries  Lower input amounts are at sample owners risk All incoming DNA samples subjected to QC  Gel electrophoresis-qualitative assessment for degradation  Picogreen fluorometry -quantitation    Samples failing QC will not be processed without consent of owner Any unused sample can be returned to owner or destroyed  Return requires charges for packaging and shipping Any unused library can be returned to owner or destroyed  Return requires charges for packaging and shipping

Targeted Capture Guidelines

We currently use Agilent SureSelect products     Whole exome Custom Kinome Others Can/will sequence captured materials sent to us   TruSeq Exome Enrichment Nimblegen

Thank You!

For additional information, please contact: [email protected]

[email protected]

Phone: +1-877-737-5468