1. Internal Training Slides qBasePLUS V2.1 Undiscovered possibility

2.  Laborious techniques were used to examine specific gene expression levels;  Northern Blots  Southern Blots  RNAse Protection Assays  Competitive PCR  Etc… etc…  These methods were Simi-quantitative at best. 10 fold! I don’t see this as 10 fold. It looks more like three fold to me! Historically… before qPCR

3. After the arrival of qPCR  Assay development was much easier*.  Sample required was much smaller.  Results were achieved much faster.  Data sets grew larger.  New instruments generated more numerical outputs.

4.  Results presented in many scientific publications are flawed... The Problem

5. A new beginning.

6. What we provide Bio-Rad CFX Manager Software v. 2.1 We have to provide the most scientific and professional solution for qPCR Experiments User friendly Conforms to MIQE Guidelines Built in analysis modules Multi-plate experiments And more...

7. Bio-Rad and Biogazelle Team UP We provide license of QbasePLUS free of charge with a newly sold CFX 96 Touch or CFX 384 Touch

8. Bio-Rad and Biogazelle Team UP  Combine the unsurpassed performance and flexibility of a CFX96 or CFX384 system with the impressive analysis tools in qbasePLUS software  This powerful combination will speed up your real-time PCR and accelerate discovery Return to Topics

9. Bio-Rad and Biogazelle Team UP  It is a great additional value for customers  And a sales tool for you when the customer:  -wants to expand the throughput of the analysis  -has to use biostatistical calculations  -want to be sure about the quality of the experiment  wants to analyse results from different istruments, different research group

10. qbase PLUS Software  Based on proven, peer-reviewed technology  qbase universal quantification model  More than 5000 downloads worldwide  Includes geNorm normalization, more than 1000 citations

11. qbase PLUS Software  Based on proven, peer-reviewed technology  qbase universal quantification model  More than 5000 downloads worldwide  Includes geNorm normalization, more than 1000 citations

12. Why qbase PLUS ?  Provides additional sophisticated options for analyzing and validating real-time PCR and gene expression data

13. Benefits qbase PLUS Software  Benefits of using qbase PLUS software include  Trust your results - based on proven solutions for quality control, normalization, and inter-run calibration  Save time - automated, fast calculations with the improved experiment accuracy  Handle large data sets - direct and batch import of data from multiple experiments, integrated data organization  Flexible - intuitive for small experiments, powerful for big experiments  Biostatistics tools - for biological replicates

14. Why qbase PLUS ? FeatureBenefit Compatible with all computer operating systems, including Microsoft Windows, Mac and Linux Customer can use their favorite operating system for analysis Import and compare data from any real-time PCR systems, Bio-Rad or other companies Reduced barrier to switching from a competitive platform. Facilitates inter-lab collaboration Advanced QC controls, normalization and inter-run calibration Improved data reliability means more meaningful and repeatable conclusions RDML and MIQE compliantMIQE (Minimum Information for publication of Quantitative real-time PCR Experiments) are annotations that allow editors/other researchers to evaluate the quality of real-time PCR data. MIQE is becoming a requirement for journal manuscript acceptance.

15. Why qbase PLUS ? FeatureBenefit geNorm evaluation of reference gene stability. Unlimited number of reference genes Reliable and repeatable gene expression relies on the selection of stable reference genes. Using 100s of reference genes is the latest approach for miRNA gene expression analysis normalization Analyze unlimited number of runs (200-300 384-well plates) Extended analysis capabilities for high throughput labs or large collaborative studies. Analyze SYBR Green/EvaGreen and probe-based experiments in a single experiment. Greater flexibility in post-run analysis reduces the need to repeat runs to answer new experimental questions. Saves raw data and analysis settings. Change analysis/QC settings and view the effects in real-time, on a single screen. For large or small data sets raw data is easily accessible and the analysis settings transparent. Removes the “black box” of data analysis.

16. 1. Run experiment(s) on CFX96 or CFX384 2. Set baseline, threshold, regression Workflow: Export data from CFX Manager Software to qbase PLUS 3. Copy and paste data into Microsoft Excel or export to RDML file 4.Import.xls or RDML run file(s) to qbase PLUS Perform QC and analysis 5.Export data/charts/RDML files as required

17. New Project Annotations An annotation box allows for the detailed description of the project. As a “Project” may have multiple experiments and a large number of combined assays, annotating becomes an important feature.

18. Experiment Settings One default amplification efficiency for all targets implies that the ΔΔCq (ΔΔc(t)) method is to be used and 100% efficiendy is assumed. Target specific amplification efficiencies implies that for a given gene, a single average efficiency will be used. Target and run specific amplification efficiencies implies that efficiencies will be determined on a “per run” basis for a specific target

19. Experiment Targets Target Properties should be defined for easier identification and future reference. Target name Official gene symbol Entrez Gene ID Primer & Probe sequences Amplicon sequence If sequence is unavailable, where primers and probes were acquired What type of Target; of Interest or Reference These annotations comply with some of the new MIQE (Minimum Information for Quantitative Experiments) parameters.

20. Experiment Samples The sample can be given a specific identifier and identified as an Unknown, Positive Control, Negative Control or Standard. A optional sample description box allows for a more detailed description of the sample.

21. Quality Control Quality Controls function displays information on three sets of parameters: Result reproducibility of technical replicates Positive and Negative Controls Stability rating of reference target set.

22. Intermediate Results The Intermediate Results function displays results on : Amplification efficiency Normalization factors Inter Run Calibrators to be selected and linked.

23. Analysis The Analysis function displays : Report table Copy number Correlation plot geNorm Multi-target bar chart Statistics.

24. Project Conclusions The project conclusions tab allows for the annotation of combined total results derived from the summation of individual experiments.

25. FAQs Is the arrangement of Biogazelle with Bio-Rad exclusive?  Yes.  Bio-Rad is the only company to ship qbase PLUS licenses with real-time PCR instruments

26. FAQs How do my CFX96 and CFX384 customers get a copy of qbase PLUS ?  New sales of CFX96 and CFX384 will ship with a quick guide containing a unique activation code  Quick guide has instructions on how to activate the full version license  Go to biogazelle.com  Register  Download qbase PLUS  Install and activate qbase PLUS using the activation code

27. Activation Quick Guide: 10016764 revA Unique activation code will appear here

28. FAQs How long is the license valid?  The license never expires  Software upgrade and premium support valid for 1 year  50% price for an additional premium license  After one year the support and the software upgrades can be purchased for a discounted price

29. FAQs How will customers be trained and supported for qbase PLUS ?  Bio-Rad staff trained on features and benefits, demo skills and functionality  Technical support will be provided by Biogazelle (bigazelle.com)  Biogazelle.com has many online training tools  Products section >> videos  Support section >> tutorials

30.  Included with each CFX96 and CFX384 is a quick guide explaining how to export data from CFX Manager software and import into qbase PLUS  Copy and paste from CFX Manager software data table to excel (.xls format)  Export and improt RDML files Data Transfer Quick Guide: 10016883 revA

31. Conclusions  We are the only who one provide the full scientific solution for Real-Time usres,  Qbase Plus is a great additional value for customers and a sales tool for you  for analyzing and validating real-time PCR and gene expression data

32. Questions? ?