Back to the Basics Purdue iGEM 2013

The Biomakers

Back to the Basics What Do We Mean? Complex Projects Lack of Fundamental Control

The Taguchi Method Cheaper, Faster Characterization

Explanation of Problem Increase Robustness of Genetic Circuits Assess with Factorial Experiment

The Taguchi Method Sources: (2013). IGCSE and GCSE Industry. Retrieved from Venil, C. K., & Lakshmanapermalsamy, P. (2009). Taguchi Experiment Design.... Journal of Science & Technology Used in Industry – Maintain quality Our Project – Apply Taguchi to synthetic biology

Comparison of Methods Partial-Factorial Method Fewer Combinations Taguchi Method All Possible Combinations Often Not Feasible Full Factorial

Experimental Design Parameters Orthogonal Arrays Response Variable: GFP Venil, C. K., & Lakshmanapermalsamy, P. (2009). Taguchi experimental design for medium optimization…. Journal of Science & Technology

Intended Data Analysis Determining GFP Expressed Per Cell – Growth curve – Coulter counter JMP Statistical Analysis Actual DataHypothetical Data

Challenges Transformations – Competent cells – Altered growing conditions 3A Assembly – Not feasible

Future Work Complete Assembly Measure GFP per Cell for All Combinations Compare Results of Taguchi vs. Full Factorial

Bicistronic Design Increasing Reliability of Expression

Explanation of Problem Variation in Protein Expression Increase Reliability of Genetic Constructs

BCD Background Bicistronic Design Sources:Mutalik, V., Endy, D., Guimaraes, J., Cambray, G., Lam, C., Juul, M., Tran, A., & Paull, M. (2013). Precise and reliable gene expression via standard transcription and translation initiation elements. Nature, 10(4)

Monocistronic Design

Bicistronic Design

Supporting Data Protein Expression Variability Decreased Increase Protein Expression Reliability

Monocistronic DesignBicistronic Design Sources:Mutalik, V., Endy, D., Guimaraes, J., Cambray, G., Lam, C., Juul, M., Tran, A., & Paull, M. (2013). Precise and reliable gene expression via standard transcription and translation initiation elements. Nature, 10(4)

Design BCD Parts Designed Golden Gate Assembly

Design

Methods Synthesized Constructs Placed Ptrc* into pSB1C3 Growth Rate Assay Re-Assembled Constructs into pSB1C3 Proof of Function

Data Promoter Submitted to the Registry Growth Rate Assay Fluorescence Intensity

Conclusion Promoter Submitted to Registry Implementation Increase Reliability

Future Work Compare Biobrick BCDs to Original BCDs Submit BCDs to iGEM HQ

Standardized Datasheets Revolutionizing Characterization

Explanation of Problem Lack of Characterization

Explanation of Problem Lack of Characterization Vague Requirements Difficult to Judge

Background Research Standardized Protocol Researched Past Protocols Found s, Sent Survey Survey Results Changed Direction

First Draft Prior Art – Drew Endy

First Draft Prior Art – Drew Endy – Boston iGEM

First Draft Prior Art – Drew Endy – Boston iGEM First Draft – Design

Feedback Prior Art – Drew Endy – Boston iGEM First Draft – Design – Implementation

Feedback Prior Art – Drew Endy – Boston iGEM First Draft – Design – Implementation Second Survey and Video Conferences Ideas Generated

Final Design and Feedback BostonU iGEM Visit Final Draft

Final Design and Feedback BostonU iGEM Visit Final Draft Third Survey and Feedback

Conclusion and Future Work Implementation Continue Collaboration Bba_K

Human Practices

Intro and Overview Outreach to Producers Outreach to High School Girl Scouts Workshop Biomaker Bench

High School Outreach Teaching modeling to AP Biology Teachers – Educating next generation of Synthetic Biologists

Girl Scout Outreach Generate interest in STEM Workshop Curriculum – Lecture – Experiment – Scientific method

Biomaker Bench Creating a Community Lab in Noblesville, IN – Business plan – Nonprofit organization

Conclusion

Accomplishments Statistical analysis method to fit synthetic biology Updated Drew Endy's Bicistronic Design Constructs Designed a Datasheet Standard for the Registry Submitted a New Promoter and 4 BCD Constructs Collaborated with over 75 iGEM Teams Performed Outreach to Local Farmers and Producers Taught High School Teachers About Modeling Worked with the Girls Scouts of America Community Lab Space in Noblesville, Indiana

Attributions and Acknowledgements Advisers:Purdue University: Dr. Jenna Rickus Dr. Fernandez Dr. Kari Clase Dr. Dilkes Soo Ha Dr. Akridge Janie Brennan Jen Kahn Bindley Bioscience Center:Research Support: Rajitarun Madangopal Boston University iGEM Dr. Larisa Avramova Drew Endy and the Endy Lab Dr. Lake Paul Dr. Tony Pedley

Sponsors

Questions?