Molecular cloning and characterization of Anaerobiosis-inducible promoters from Escherichia coli and Salmonella typhimurium.

• Other Authors: Wong, Kwong-Kwok.
• Format: Others
• Language: English
• Published: Chinese University of Hong Kong 1990
• Subjects: Molecular cloning; Promoters (Genetics); Escherichia coli; Salmonella typhimurium
• Online Access: http://library.cuhk.edu.hk/record=b5886597; http://repository.lib.cuhk.edu.hk/en/item/cuhk-318559

by Kwong-Kwok Wong. === Thesis (Ph.D)--Chinese University of Hong Kong, 1990. === Bibliography: leaves 171-183. === TITLE PAGE --- p.I === ABSTRACT --- p.II === STATEMENT --- p.V === ACKNOWLEGEMENTS --- p.VI === ABBREVIATIONS --- p.VII === TABLE OF CONTENTS --- p.VIII === LIST OF TABLES --- p.XIII === LIST OF FIGURES --- p.XVI === Chapter Chapter 1. --- Introduction and Literature Review --- p.1 === Chapter I . --- Introduction --- p.1 === Chapter A. --- General introduction --- p.1 === Chapter B. --- Purpose of study --- p.5 === Chapter II. --- Literature review --- p.6 === Chapter A. --- Global control of aerobic-anaerobic shift --- p.6 === Chapter B. --- Identified anaerobiosis-inducible genes --- p.8 === Chapter C. --- Genetics of anaerobic regulation --- p.15 === Chapter i. --- Redox control --- p.15 === Chapter ii. --- DNA conformation --- p.15 === Chapter iii. --- fnr (oxrA) regulatory gene --- p.16 === Chapter iv. --- narL gene --- p.18 === Chapter v. --- Other regulatory genes --- p.19 === Chapter vi. --- Proposed FNR and NarL recognition sequences --- p.20 === Chapter D. --- future prospect --- p.23 === Chapter Chapter 2. --- Isolation of Anaerobiosis-inducible Promoters --- p.25 === Chapter I . --- Introduction --- p.25 === Chapter A. --- Properties of promoter-probe plasmid pKK232.8 --- p.26 === Chapter B. --- Properties of promoter-probe plasmid pFZYl --- p.28 === Chapter II. --- Materials and methods --- p.30 === Chapter A. --- Bacterial strains and plasmids --- p.30 === Chapter B. --- Media --- p.30 === Chapter C. --- Solutions --- p.31 === Chapter D. --- Small scale prepartaion of plasmid DNA --- p.32 === Chapter E. --- Large scale preparation of plasmid DNA --- p.33 === Chapter F. --- Digestion of DNA with restriction endonucleases --- p.35 === Chapter G. --- Analysis of DNA samples with agarose gel electrophoresis --- p.36 === Chapter H. --- Dephosphorylation of DNA fragments --- p.37 === Chapter I. --- Partial digestion of Chromosomal DNA with restriction enzyme Sau3A. --- p.37 === Chapter J. --- Ligation of DNA --- p.38 === Chapter K. --- Preparation of competent cells --- p.38 === Chapter L. --- Transformation --- p.40 === Chapter M. --- Chloramphenicol resistance levels test for promoter clones with plasmid pKK232.8 --- p.41 === Chapter N. --- Preparation of crude cell extract for chloramphenicol acetyltransferase (CAT) assays --- p.41 === Chapter O. --- CAT assay --- p.42 === Chapter P. --- Protein assay --- p.42 === Chapter Q. --- β-galactosidase assay --- p.43 === Chapter III . --- Results --- p.45 === Chapter A. --- Molecular cloning of anaerobiosis-inducible promoters with promoter-probe plasmid pKK232.8 --- p.45 === Chapter B. --- Molecular cloning of anaerobiosis-inducible promoters with promoter-probe plasmid pFZYl --- p.54 === Chapter IV. --- Summary and Discussion --- p.70 === Chapter A. --- Cloning with promoter-probe plasmid pKK232.8 --- p.70 === Chapter B. --- Cloning with promoter-probe plasmid pFZYl --- p.71 === Chapter C. --- Number of anaerobiosis inducible promoters --- p.73 === Chapter Chapter 3. --- Subcloning and Sequencing --- p.74 === Chapter I . --- Introduction --- p.74 === Chapter II. --- Materials and methods --- p.74 === Chapter A. --- Bacterial strains and bacteriophages --- p.74 === Chapter B. --- Preparation of M13mp RF plasmid --- p.75 === Chapter C. --- DNA sequencing by the chain termination method --- p.75 === Chapter D. --- Polymerase chain reaction (PCR) for the amplification of DNA fragments cloned in plasmid pFZYl --- p.79 === Chapter E. --- Using Exonuclease III to construct unidirectional deletions to generate nested clones --- p.80 === Chapter F. --- Direct gel electrophoresis --- p.81 === Chapter G. --- C-testiscreening for the orientation of insert in M13 phage --- p.81 === Chapter III . --- Results --- p.82 === Chapter A. --- Subcloning and sequencing of pFE29 and pFE117 --- p.82 === Chapter B. --- Subcloning and sequencing of pHSKl --- p.90 === Chapter C. --- Subcloning and sequencing of pHSK8 --- p.100 === Chapter D. --- "Subclonig and sequencing of pFSl, pFS22 and pFS3 4" --- p.109 === Chapter IV. --- Summary and Discussion --- p.113 === Chapter A. --- Trimming down size of DNA fragments to smaller fragments which still contained anaerobiosis-inducible promoters --- p.113 === Chapter B. --- Nucleotide sequencing --- p.113 === Chapter C. --- Sucloning and sequencing strategy --- p.115 === Chapter Chapter 4. --- Expression of Anaerobiosis-inducible Promoters --- p.120 === Chapter I . --- Introduction --- p.120 === Chapter II. --- Materials and methods --- p.122 === Chapter A. --- Bacterial strains and phages --- p.122 === Chapter B. --- Media --- p.125 === Chapter C. --- Transformation in Salmonella typhimurium --- p.125 === Chapter D. --- Genetic techniques --- p.126 === Chapter III. --- Results --- p.129 === Chapter A. --- Expression of Escherichia coli qlpT promoter --- p.129 === Chapter B. --- "Expression of Salmonella typhimurium anaerobiosis-inducible promoters cloned in pHSK8, pFS22 and pFS34" --- p.134 === Chapter IV. --- Summary and Discussion --- p.137 === Chapter A. --- A pair of divergent promoters were both regulated by anaerobiosis and glucose. --- p.137 === Chapter B. --- fnr(oxrA) dependent and independent promoters --- p.137 === Chapter C. --- Effect of nitrate on anaerobiosis expression. --- p.138 === Chapter Chapter 5. --- Analysis of Anaerobiosis-inducible Promoter-containing DNA sequences and Final Discussion --- p.141 === Chapter I. --- Analysis of anaerobiosis-inducible promoter-containing DNA sequences --- p.141 === Chapter A. --- "Search for initiation codon, conserved ""-10"" and ""-35"" regions" --- p.141 === Chapter B. --- Search for FNR binding sites and NarL binding sites. --- p.151 === Chapter C. --- Homology search among the promoter sequences of all anaerobiosis-inducible genes. --- p.156 === Chapter II. --- Final Discussions. --- p.164 === Chapter A. --- Summary of the properties of the sequenced and characterized promoters cloned in this study --- p.164 === Chapter B. --- Further studies. --- p.167 === REFERENCES --- p.168