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

TABLE OF CONTENTS
1 Introduction ………………………………………………………………….. 1
1.1 Is there a common coordinating factor? ………………………………... 2
1.1.1 Is there a common coordinating mechanism? ………………………… 3
1.2 Global transcription coordination - current knowledge …………….. 4
1.2.1 Gene promoters ………………………………………………………. 5
1.2.2’Gene promoters respond distinctly to changes of supercoiling ………… 7
1.2.3 DNA Supercoiling and mechanisms regulating it ……………………. 9
1.2.4 Supercoiling and growth transitions …………………………………. 11
1.2.5 The nucleoid associated proteins – major players in global gene expression
regulation …………………………………………………………….. 12
1.2.6 Composition of RNAP also changes with growth …………………… 14
1.3 Organisation of global transcription - coordinated view …………… 15
2 Materials and methods ……………………………………………………… 18
2.1 Materials ……………………………………………………………….. 18
2.1.1 Enzymes & chemicals ………………………………………………… 18
2.1.2 Antibiotics ……………………………………………………………. 19
2.1.3 Media …………………………………………………………………. 19
2.1.4 Buffers and stock solutions …………………………………………… 20
2.1.5 Bacterial strains ………………………………………………………. 21
2.1.6 Plasmids ……………………………………………………………… 22
2.1.7 Primers ……………………………………………………………….. 23
2.2 Methods ………………………………………………………………… 24
2.2.1 Agarose gel electrophoresis ………………………………………….. 24
2.2.2 Small and medium scale isolation of plasmid DNA using Qiagen kit … 24
2.2.3 Restriction Digestion ………………………………………………… 25
2.2.4 Polymerase chain reaction …………………………………………… 25
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