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Lab Meeting

Lab Meeting. Catherine Doyle 09/13/13. RBS. Aptamer. Mutate for 6,7,or 8 base paring with RNA polymerase. Add Riboswitch w/o apatamer. Complete Riboswitch. Mutated RBS. Randomize 8 bp between RBS and aptamer. Add two 8 bp spacers and ATG to RBS. 65,536 Riboswitches.

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Lab Meeting

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  1. Lab Meeting Catherine Doyle 09/13/13

  2. RBS Aptamer Mutate for 6,7,or 8 base paring with RNA polymerase Add Riboswitch w/o apatamer Complete Riboswitch Mutated RBS Randomize 8 bp between RBS and aptamer Add two 8 bp spacers and ATG to RBS 65,536 Riboswitches Predict free energy of aptamer M-fold Riboswitch w/o Aptamer Riboswtiches with Calculated Free Energy Filter out riboswitches too large of free energy Riboswitches with greater free energy than aptamer Putative Riboswitches

  3. Caffeine Metabolism

  4. c.Dog RBS eCDM8 psB4C5 GFP Low Copy psB1A8 High Copy

  5. c.Dog RBS eCDM8 psB1A8 GFP High Copy psB4C5 Low Copy

  6. eCDM8 c.Dog RBS GFP psB1A8 High Copy

  7. Inoculate in media with different metabolites + caffeine + 3-methyl - caffeine + caffeine + caffeine + xanthine - caffeine + 3-methyl - caffeine + xanthine Grow o/n @ 37C and measure GFP expression

  8. Future Work • Implement UnaFold and generate putative riboswitches

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