1 / 27

Light- Based

Light- Based . Cell-Cell Signaling. VERSATILITY PENETRABILITY ORTHOGONALITY CONTROL. Our System. Coelenterazine. Ca 2+. 470nm Photons. 470nm Photons. eYFP. Renilla Luciferase. CMV Promoter BBa_I712004. Renila. CMV Promoter BBa_I712004. CatCh-eYFP. Humanized Aeq BBa_K548000.

sally
Download Presentation

Light- Based

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Light-Based Cell-Cell Signaling

  2. VERSATILITY PENETRABILITY ORTHOGONALITY CONTROL

  3. Our System Coelenterazine Ca2+ 470nm Photons 470nm Photons eYFP RenillaLuciferase CMV Promoter BBa_I712004 Renila CMV Promoter BBa_I712004 CatCh-eYFP Humanized Aeq BBa_K548000 HEK 293T Cell HEK 293T Cell

  4. The Sender Cell Coelenterazine Ca2+ 470nm Photons 470nm Photons eYFP RenillaLuciferase CMV Promoter BBa_I712004 Renila CMV Promoter BBa_I712004 CatCh-eYFP Humanized Aeq BBa_K548000 HEK 293T Cell HEK 293T Cell

  5. Gaussia and Renilla Luciferase Coelenteramide + CO2 + 470 nm photons Gaussia or Renilla Coelenterazine + O2 http://www.cmarz.org/CMarZ_RHBrown_April06/images_press/index.html http://profmother.files.wordpress.com/2010/06/seapansy3_jpg.jpg

  6. Ivis Lumina II 5.53 E6 5.53 E6 pRL-CMV-Renilla pRRL-CMV-Gaussia 2.79 E5 2.79 E5

  7. Renilla + CZ Luminescence Time Course

  8. The Receiver Cell Coelenterazine Ca2+ 470nm Photons 470nm Photons eYFP RenillaLuciferase CMV Promoter BBa_I712004 Renila CMV Promoter BBa_I712004 CatCh-eYFP Humanized Aeq BBa_K548000 HEK 293T Cell HEK 293T Cell CatCh Aequorin

  9. pRRL–CMV–Aeq Aeq pENTR5-CMV pRRL + + = pENTR-d/TOPO-Aeq pRRL-CMV-Aeq

  10. Characterization of Aequorin

  11. Fura 2-AM Ca2+ Imaging • Ratio-metric Calcium Dye • Emission – 510 nm • Calcium Free Excitation – 380nm • Calcium Bound Excitation – 340 nm • Measure change in emission at 340/380 380nm 340nm 340/380

  12. Fura 2-AM Ca2+ Imaging Ionomycin – Positive Control

  13. Fura 2-AM Ca2+ Imaging pRRL-CMV-CatCh-eYFP

  14. Fura 2-AM Ca2+ Imaging pRRL-CMV-CatCh-eYFP co-cultured with pRL-CMV-Renilla

  15. Human Practices http://www.preciouslife.com/saimages/346.png http://www.dailymail.co.uk/sciencetech/article-1279988/Artificial-life-created-Craig-Venter--wipe-humanity.html

  16. Goals

  17. How Big is Media Influence? • Conducted a survey of over 200 people • Asked a wide variety of questions http://www.aseonline.biz/Portals/0/survey-man.jpg

  18. How Big is Media Influence? Biology-Related Field Non-Biology Related Field Non-Classroom Biology Experience

  19. How Big is Media Influence? Heard of Synthetic Bio Heard of Genetic Eng. Not Heard of Either

  20. How Big is Media Influence? Other Sources 14% 38% News Media (non-scientific) 19% Scientific Literature 28% Science-focused media

  21. How Big is Media Influence? Other Sources 19% 44% News Media (non-scientific) 12% Scientific Literature 25% Science-focused media

  22. What Can We Do? • “The media shapes perception” • “We need to recognize, respect, and understand the fear surrounding Synthetic Biology” • Synthetic Biologists need to join the conversation! • We are not heard in the most widely disseminated forms of media. Valerie Bonham http://bioethics.gov/cms/valerie-bonham http://bioethics.gov/

  23. Discussion • Optimization of interactions between sender & receiver cells Ca2+ Coelenterazine 470nm Photons 470nm Photons eYFP RenillaLuciferase CMV Promoter BBa_I712004 Renila CMV Promoter BBa_I712004 CatCh-eYFP Humanized Aeq BBa_K548000 HEK 293T Cell HEK 293T Cell • Our sender cell can produce a detectable signal • Our receiver cell can respond to bright light

  24. Looking Ahead…

  25. Thanks Everyone! Dr. CasimSarkar Dr. Jordan Miller Dr. Dan Cohen Dr. Anthony Choo Daphne Ng Najaf Shah

  26. Questions?

More Related