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A Synthetic Biology Approach To Engineered Competency in E. coli

A Synthetic Biology Approach To Engineered Competency in E. coli. Samantha Simpson Genomics 4/8/09. Why Would You Want To Insert New DNA into E. coli ?. Insulin Gene Extracted. Plasmid. My Focus. Insulin Gene Extracted. Plasmid. A Closer Look at Taking Up New DNA.

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A Synthetic Biology Approach To Engineered Competency in E. coli

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  1. A Synthetic Biology Approach To Engineered Competency in E. coli Samantha Simpson Genomics 4/8/09

  2. Why Would You Want To Insert New DNA into E. coli? Insulin Gene Extracted Plasmid

  3. My Focus Insulin Gene Extracted Plasmid

  4. A Closer Look at Taking Up New DNA

  5. What Is Competency?

  6. How does natural competency work?

  7. How does ComK activate late competency genes? ComK Late Competency Gene K-box promoter

  8. How are cells made competent? • Natural Competency in B. subtilis • Controlled by ComK • Induced Competency in E. coli Both 48 genes Microarray 57 genes K-boxes 1014

  9. Is natural competency feasible in E. coli?

  10. Is natural competency feasible in E. coli? K-Box to gene distance not consistent.

  11. Building a Testable System } 600 bps ComK ComK MWM High Copy AmpR 1018 bp 506 bp 396 bp 344 bp 298 bp

  12. Building a Testable System RBS ComK GFP pKBox Low Copy High Copy AmpR KanR

  13. Testing Fluorescence p < 0.02

  14. Building a Testable System RBS RBS ComK GFP pKBox Low Copy High Copy AmpR KanR

  15. RBS’s: Not Consistent

  16. Promoters: Consistent

  17. Building a Competent System ComK Low Copy KanR

  18. Testing Competency ComK / Kan Pellet Cells RFP / Amp LB + Kan/Amp

  19. Conclusions In E. coli: In B. subtilis: pKBox + ComK

  20. Acknowledgments • Davidson Research Initiative • National Science Foundation • Howard Hughes Medical Institute • James G. Martin Genomics Program • Dr. Heyer, Dr. Campbell and Dr. Denham • Fellow lab workers • Friends and Family

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