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Development of the Theophylline Riboswitch

Development of the Theophylline Riboswitch. 1 on 1 Meeting 09/03/13. Original Riboswitch Design ( Desaia and Gallivan , 2004). 5’-GGTGATACCAGCATCGTCTTGATGCCCTTGGCAGCACC-3’. 5 bp upstream. B- galactosidase : Translated Region. ATG. RBS. TGA.

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Development of the Theophylline Riboswitch

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  1. Development of the Theophylline Riboswitch 1 on 1 Meeting 09/03/13

  2. Original Riboswitch Design (Desaia and Gallivan, 2004) 5’-GGTGATACCAGCATCGTCTTGATGCCCTTGGCAGCACC-3’ 5 bp upstream B-galactosidase: Translated Region ATG RBS TGA

  3. Original Riboswitch Design (Desaia and Gallivan, 2004) B-galactosidase: Translated Region Kpn1 Aptamer 5 bp 25 bp of RBS Fwd Primer RBS Untranslated region 8bp spacer Untranslated region

  4. Results

  5. Results

  6. Paper published on mechanistic Insights into Riboswithces(Lynch et al., 2007)

  7. Selection

  8. Results

  9. Results 8.1 8.2

  10. Final TheophylineRiboswitch(Topp et al., 2010) • 5 Riboswitches • Experiment with Mutation in the RBS to increase base pairing with Aptamer • Experiment with changing positions of the random sequencing of spacers

  11. C A B C2 D E

  12. RBS’ of Riboswitches RBS---Shine-Dalgarnosequence: AGGAGGU Generated a library of randomized RBS sequences to see which one based paired to Aptamer

  13. Results

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