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REM e System

REM e System. January, 2010. IMPORTANT NOTICE Intended Use. Unless explicitly stated otherwise, all Roche Applied Science and 454 Sequencing products and services referenced in this presentation / document are intended for the following use: For Life Science Research Only.

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REM e System

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  1. REM e System January, 2010

  2. IMPORTANT NOTICEIntended Use Unless explicitly stated otherwise, all Roche Applied Science and 454 Sequencing products and services referenced in this presentation / document are intended for the following use: For Life Science Research Only. Not for Use in Diagnostic Procedures. 2 of 37 For Internal Use Only

  3. Agenda • What is the REM e System? • Product Positioning • 454 Life Sciences and EAP Data • Customer Requirements • Liquid Handlers • Consumables • Technician Skill Sets • Installation and Service • Troubleshooting • Service 3 of 37 For Internal Use Only

  4. What is the REM e System? • REM e System automates bead enrichment & sequence primer annealing (all steps after breaking) • Mechanical functions: • Vortexing • Vacuum filtration (replaces centrifugation) • Magnetic bead capture • Heating (65 °C) and cooling • A deck accessory compatible with common liquid handling robots • Liquid handler delivers and transfers buffers (mostly non-critical volumes), and sends serial commands to the REM e Control Box 4 of 37 For Internal Use Only

  5. Where Does the REM e System Fit in ? 454 Sequencing Workflow Analysis Sequencing Library Prep emPCR Enrichment GS FLX Seq. Fragment Ligate Adaptors Single Strand Emul. Prep & Amplif. Breaking Seq Prep Analysis • REM e System • Enrichment • Collecting beads • Primer annealing 5 of 37 For Internal Use Only

  6. Simplifies Workflow Manual ProcessREM e Automated Process Manual Steps Enrich Separate null beads 30 xfers, 8 spins, 10 vortex, 28 washes 4 magnet concentrations Automated Steps Setup Robot Prepare reagents Start robot program Collect DNA beads Separate enrich beads 10 xfers, 10 spins, 4 vortex, 6 washes 4 magnet concentrations Liquid Handler & REM Enrich Collect DNA Beads Anneal Seq Primer Anneal Seq Primer anneal 6 xfers, 2 washes 1 incubate Setup: 1 installation, 1 programming Each Run:load robot deck, walk away Setup: none Each Run:50 transfer, 24 spins, 16 vortex, 48 washes, 6 of 37 For Internal Use Only

  7. Reduces Hands-On Time * Estimate is based on one (1) REM e System installed on a liquid handler with span-8 channel and 1mL disposable tips * * Initial setup and programming time for REM e System is not included 7 of 37 For Internal Use Only

  8. 454 Sequencing Center Data • Sample - E. coli K12: • Library prepared with GS FLX Titanium emPCR Kit • REM e System installed on Hamilton Starlet; Bulks were broken via 454 SOP (similar to standard user instruction). • Sequenced on a 4 pad PicoTiterPlate Device • Conclusion – data looks good! 8 of 37 For Internal Use Only

  9. 454 Sequencing Center DataCross Contamination Check • Four bulks of MID-labeled Bacillus libraries • “Tip swap” – Tip changes each time when it touches the beads • Cross contamination is represented by % of unexpected MID • Conclusion-REM e System does not introduce cross contamination • Conclusion- “tip swap” does not further prevent cross contamination 9 of 37 For Internal Use Only

  10. Customer Requirements: • One dedicated or semi-dedicated liquid handler system • Purchase reasonable amount consumables from third party • Possess working knowledge of robots 10 of 37 For Internal Use Only

  11. Liquid Handler Requirements • Dispensing Capability: • Minimum: 4 channel spanning dispense head with 1 mL disposable tips. • Ideally: 8 channel spanning dispense head with 1 mL disposable tips • Possible but with longer processing time: multichannel 200 uL • Communication Hardware & Software: • One dedicated RS-232 port on Liquid handler’s PC (for REM eSystem communication) • If serial port not available, user must install a serial port card, or a USB to RS-232 converter. • Liquid handler scripting must be capable of sending text commands through a PC RS-232 port to the REM e System (example: sending “VOR1100” “\r” to REM e System causes vortexer to orbit at 1100 rpm). If a special software module is required for this functionality, the customer must acquire and install it. • RS-232 port settings: 9600 baud, 1 stop bit, no flow control, no parity, 8 data bits. • Liquid Handler Deck Hardware: • 2 microtiter plate positions low enough for pipette tips to clear 125mm tall REM e System and 125mm tall loaded tube rack. • Recommend to place the REM System on a corner deck position to accommodate the rear overhang. • Three 200 mL volume reagent troughs 11 of 37 For Internal Use Only

  12. Liquid Handling System Examples: • Compatible - tested at 454 Life Sciences: • Hamilton STARlet, 8 dispensing channels, 1 mL disposable tips • Tecan Genesis RSP 100, 8 dispensing channels, 1 mL disposable tips • Compatible - tested by early access partners: • Biomek FX, 8 dispensing channels, 1 mL disposable tips • Hamilton Star 12 of 37 For Internal Use Only

  13. Liquid Handling System Examples: • Should be compatible based on specifications: • Beckman Biomek NX • Tecan Freedom EVO and older Genesis • PerkinElmer Janus and older Packard MultiPROBE • MultiPROBE – planned to be tested by Roche/Pleasanton and Broad • Not compatible unless with substantial engineering modifications: • Beckman Biomek 2000 and Biomek 3000 • Require machining ability to product a REM e System adaptor. Many labs do not have such capacity • Do not recommend, or recommend with great caution. • Example: Baylor were able to run the Biomek 2000 because the lab could design and machine an adaptor. But Penn State could not get it running due to the lack of machining or engineering capacity 13 of 37 For Internal Use Only

  14. Consumables Supplied by Users REM e Specific Disposables per 1 LVE Kit Vendor Part # Cost / LVE* 6   ULTRAFREE CL Filter Tubes @ $ 3 . 61 each Millipore UFC40SV25 $ 21.66 4  2 mL conical vials @ 12 cents ea Fisher Scientific UFC30SV00 $ 0.48 ~ 40 Tips (Robot dependent) @ ~12 cents ea Various Various $ 4.80 $ 26.94 *Not bulk pricing Total cost per LVE kit: REM e Specific Disposables per 8 SVEs Vendor Part # Cost / LVE* 16   ULTRAFREE MC Filter tubes @ $ 2 . 29 each Millipore UFC40SV25 $ 36.64 8  2 mL conical vials @ 12 cents ea Fisher Scientific UFC30SV00 $ 0.96 ~ 60 Tips (Robot dependent) @ ~12 cents ea Various Various $ 7.20 $ 44.80 *Not bulk pricing Total cost per 8 SVEs: 14 of 37 For Internal Use Only

  15. Skill Sets Required for Users: • Working knowledge of programming liquid handler protocols • Capable of implementing third party deck hardware onto liquid handlers • Teaching the x, y, z coordinates of the REM e System deck ware • Sending text commands from the liquid handler RS-232 port to the REM e System • Programming the liquid handler script based on provided generic 454 System protocols 15 of 37 For Internal Use Only

  16. Installation • REM e System Contains 4 Components: • Deck Module • Control Box • Pump • Waste Container • Installation by Customers • Connecting 4 components • Installing onto Liquid Handler • Fairly simple steps by following manual • Approximately. 30 minutes based on Early Access Programs Waste Pump Control Deck module 16 of 37 For Internal Use Only

  17. Quick Installation Guide Included In the Box 17 of 37 For Internal Use Only

  18. Service Or Repair • All subcomponents: • Set up as “accessories” • Can be ordered by customers and Roche personnel • No spare parts available due to high labor/material cost • No repair available for subcomponents or system • In the event of subcomponent failure: • Order subcomponents from Indy or Mannheim • Return defect subcomponents to 454 Life Sciences Customer Service for diagnose • System Facts File will be available at GSS home page 18 of 37 For Internal Use Only

  19. Availability and Part Numbers 19 of 37 For Internal Use Only

  20. Additional Info: Following info will be posted at www.454.com/my454: • List of Compatible Liquid Handlers • Software scripts from Tecan, Hamilton Starlet, Beckman Fx • Installation Guide • User Manual and Protocols • REM e Command Software 20 of 37 For Internal Use Only

  21. Images of the REM e 31 of 37 For Internal Use Only

  22. Images of REM eDispensing of Reagents 32 of 37 For Internal Use Only

  23. Images of REM e 33 of 37 For Internal Use Only

  24. Images of REM e 34 of 37 For Internal Use Only

  25. Images of REM e 35 of 37 For Internal Use Only

  26. Software Images 36 of 37 For Internal Use Only

  27. Software Images 37 of 37 For Internal Use Only

  28. Software Images 38 of 37 For Internal Use Only

  29. For life science research only. Not for use in diagnostic procedures. 454, 454 SEQUENCING, 454 LIFE SCIENCES, PICOTITERPLATE, EMPCR, GS FLX, GS FLX TITANIUM, GS JUNIOR, and REM are trademarks of Roche 39 of 37 For Internal Use Only

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