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Back to the Basics

Back to the Basics

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Back to the Basics

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  1. Purdue iGEM 2013 Back to the Basics

  2. The Biomakers

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

  4. The Taguchi Method Cheaper, Faster Characterization

  5. Explanation of Problem • Increase Robustness of Genetic Circuits • Assess with Factorial Experiment

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

  7. Comparison of Methods • Full Factorial • All Possible Combinations • Often Not Feasible

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

  9. Intended Data Analysis • Determining GFP Expressed Per Cell • Growth curve • Coulter counter • JMP Statistical Analysis Actual Data Hypothetical Data

  10. Challenges • Transformations • Competent cells • Altered growing conditions • 3A Assembly • Not feasible

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

  12. Bicistronic Design Increasing Reliability of Expression

  13. Explanation of Problem • Variation in Protein Expression • Increase Reliability of Genetic Constructs

  14. 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)

  15. Monocistronic Design

  16. Bicistronic Design

  17. Supporting Data • Protein Expression Variability Decreased • Increase Protein Expression Reliability

  18. Monocistronic Design 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)

  19. Design • BCD Parts Designed • Golden Gate Assembly

  20. Design

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

  22. Data • Promoter Submitted to the Registry • Growth Rate Assay •  Fluorescence Intensity

  23. Conclusion • Promoter Submitted to Registry • Implementation • Increase Reliability

  24. Future Work • Compare Biobrick BCDs to Original BCDs • Submit BCDs to iGEM HQ

  25. Standardized Datasheets Revolutionizing Characterization

  26. Explanation of Problem • Lack of Characterization

  27. Explanation of Problem • Lack of Characterization • Vague Requirements • Difficult to Judge

  28. Background Research • Standardized Protocol • Researched Past Protocols • Found Emails, Sent Survey • Survey Results Changed Direction

  29. First Draft • Prior Art • Drew Endy

  30. First Draft • Prior Art • Drew Endy • Boston iGEM

  31. First Draft • Prior Art • Drew Endy • Boston iGEM • First Draft • Design

  32. Feedback • Prior Art • Drew Endy • Boston iGEM • First Draft • Design • Implementation

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

  34. Final Design and Feedback • BostonUiGEM Visit • Final Draft

  35. Final Design and Feedback • BostonUiGEM Visit • Final Draft • Third Survey and Feedback

  36. Conclusion and Future Work • Implementation • Continue Collaboration • Bba_K1225000

  37. Human Practices

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

  39. High School Outreach • Teaching modeling to AP Biology Teachers • Educating next generation of Synthetic Biologists

  40. Girl Scout Outreach • Generate interest in STEM • Workshop Curriculum • Lecture • Experiment • Scientific method

  41. Biomaker Bench • Creating a Community Lab in Noblesville, IN • Business plan • Nonprofit organization

  42. Conclusion

  43. Accomplishments • Statistical analysis method to fit synthetic biology • Updated Drew Endy'sBicistronicDesign 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

  44. 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: RajitarunMadangopal Boston University iGEM Dr. Larisa Avramova Drew Endy and the Endy Lab Dr. Lake Paul Dr. Tony Pedley