Coordinated regulation of gene expression by E ... - Jacobs University

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RESULTS techniques. Potassium permanganate reacts with bases, especially thymine, where the base-stacking in the DNA duplex is disrupted [Gralla et al., 1993]. In these experiments Figure 20: in vitro transcription from tyrTwild-type and tyrTD at different supercoiling. (A) In vitro transcription at different superhelical densities from tyrT wild-type, tyrTD and reference bla promoters is shown. (B) Fold activation of transcription by FIS from tyrT wild-type and tyrT D at different superhelical densities. RNA polymerase was used in large excess to promoter DNA and the complexes were equilibrated for 30 minutes prior to a short exposure to permanganate ion. Figure 21A (lanes 1 and 2) shows that addition of RNA polymerase to the wild-type promoter did not alter the reactivity pattern on native supercoiled plasmid DNA (σ = −0.068). By 55
RESULTS contrast the tyrTD promoter mutant showed significantly increased reactivity within the −10 region at positions T-8, A-10, T-11 and A-12 (lanes 4, 5). In addition bases T-3 and T-4 within the discriminator as well as bases T+3 and T+4 were reactive suggesting that DNA unwinding had extended from the −10 region to the start point in this mutant. No enhanced signals were observed in the other mutants in these positions (C-3, C-4). For both the tyrTS and tyrT35U increased reactivity was observed within the −10 region at positions T-8, A-10, T-11 and A-12 (lanes 7, 8 and 10, 11) although the effect was much for the former mutant. Unlike the tyrTD mutant neither of the tyrTS nor the tyrT35U mutant promoters exhibited increased reactivity at positions T+3, T+4. Figure 21: Potassium permanganate footprinting of ptyrT and its derivatives. (A) Footprinting performed in the absence of nucleoside triphosphates. The positions of permanganate reactive thymine bases are indicated. (B) Footprinting performed in the presence of initiating nucleoside triphosphates GTP and CTP. FIS and RNAP when added is indicated by a ‘+’. Wt, D, S and 35U are the wild-type and mutants of ptyrT. 56
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Coordinated regulation of gene expr
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Parts of this work has been publish
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ABSTRACT promoter region determines
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ACKNOWLEDGEMENTS first impression o
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TABLE OF CONTENTS 2.2.5 Preparation
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LIST OF TABLES List of tables Table
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LIST OF FIGURES List of Figures Fig
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ABBREVIATIONS Abbreviations ∆61
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ABBREVIATIONS TBE Tris TS tyrT35U t
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INTRODUCTION Salmonella: “In more
Page 21 and 22: INTRODUCTION Figure 1: Schematic il
Page 23 and 24: INTRODUCTION UP element -35 -10 Spa
Page 25 and 26: INTRODUCTION can also influence elo
Page 27 and 28: INTRODUCTION have more base-pairs t
Page 29 and 30: INTRODUCTION Figure 6: Graphical re
Page 31 and 32: INTRODUCTION shown that H-NS reache
Page 33 and 34: INTRODUCTION formed as a repercussi
Page 35 and 36: MATERIALS AND METHODS 2 Materials a
Page 37 and 38: MATERIALS AND METHODS NaCl 10 gm YT
Page 39 and 40: MATERIALS AND METHODS LZ41 3 pyrF28
Page 41 and 42: MATERIALS AND METHODS 2.2 Methods 2
Page 43 and 44: MATERIALS AND METHODS given using a
Page 45 and 46: MATERIALS AND METHODS phenol: chlor
Page 47 and 48: MATERIALS AND METHODS Filtered (by
Page 49 and 50: MATERIALS AND METHODS free water (A
Page 51 and 52: RESULTS 3 Results Growth phase-depe
Page 53 and 54: RESULTS Neither the fis nor the hns
Page 55 and 56: RESULTS these identified genes (by
Page 57 and 58: RESULTS furthermore the colour of t
Page 59 and 60: RESULTS During the entire growth cy
Page 61 and 62: RESULTS transcripts of a growth pha
Page 63 and 64: RESULTS phosphorylation, are associ
Page 65 and 66: RESULTS Table 4: RT-PCR analysis of
Page 67 and 68: RESULTS Figure 16: Schematic repres
Page 69 and 70: RESULTS parC) which allows the supe
Page 71: RESULTS Figure 19: In vitro transcr
Page 75 and 76: DISCUSSION 4 Discussion The idea of
Page 77 and 78: DISCUSSION In a study by Balke & Gr
Page 79 and 80: DISCUSSION cells maintain viability
Page 81 and 82: DISCUSSION writhe in the UAS by FIS
Page 83 and 84: DISCUSSION 4.3.2 Mechanism of trans
Page 85 and 86: CONCLUSION 5 Conclusion Regulation
Page 87 and 88: REFERENCES Bordes, P., Conter, A.,
Page 89 and 90: REFERENCES Gruber, T.M., and Gross,
Page 91 and 92: REFERENCES molecular basis for sele
Page 93 and 94: REFERENCES Pan, C.Q., Finkel, S.E.,
Page 95 and 96: REFERENCES Schroder, O., and Wagner
Page 97 and 98: REFERENCES Willenbrock, H., and Uss
Page 99 and 100: APPENDIX I gene blattner ratio p-va
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APPENDIX I gene blattner ratio p-va
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APPENDIX I Supplementary table - VI
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APPENDIX I fabB b2323 0.6336 0.0043
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APPENDIX I ibpA b3687 2.4960 0.0181
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APPENDIX I pepP b2908 1.2182 0.0368
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APPENDIX I speG b1584 0.4151 0.0001
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APPENDIX I ycjX b1321 4.0841 0.0141
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APPENDIX I yhfY b3382 1.4063 0.0453
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APPENDIX I gene blattner ratio A/B
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Statement of sources Declaration I
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