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| Preface | |
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| Acknowledgments | |
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| Introduction to the cell | |
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| The bacterial cell: a quick overview | |
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| How do cells grow? | |
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| What is genetics? | |
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| Summary | |
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| The bacterial DNA molecule | |
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| The structure of DNA and RNA | |
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| Deoxyribonucleosides and deoxyribonucleotides | |
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| DNA is only polymerized 5' to 3' | |
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| Double-stranded DNA | |
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| Supercoiling double-stranded DNA | |
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| Replication of the Escherichia coli chromosome | |
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| Constraints that influence DNA replication | |
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| The replication machinery | |
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| DNA polymerases | |
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| DnaG primase | |
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| DnaA, DnaB, and DnaC | |
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| Replication of both strands | |
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| Theta mode replication | |
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| Minimizing mistakes in DNA replication | |
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| The DNA replication machinery as molecular tools | |
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| Summary | |
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| Mutations | |
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| Phenotype and genotype | |
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| Classes of mutations | |
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| Point mutations and their consequences | |
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| Measuring mutations: rate and frequency | |
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| Spontaneous and induced mutations | |
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| Errors during DNA replication: incorporation errors | |
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| Errors due to tautomerism | |
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| Spontaneous alteration by depurination | |
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| Spontaneous alteration by deamination | |
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| Alterations by spontaneous genetic rearrangement | |
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| Alterations caused by transposition | |
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| Induced mutations | |
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| Chemicals that mimic normal DNA bases: base analogs | |
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| Chemicals that react with DNA bases: base modifiers | |
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| Chemicals that bind DNA bases: intercalators | |
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| Mutagens that physically damage the DNA: ultraviolet light and ionizing radiation | |
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| Mutator strains | |
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| Reverting mutations | |
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| Suppression | |
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| Ames test | |
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| How have we exploited bacterial mutants? | |
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| Summary | |
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| DNA repair | |
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| Lesions that constitute DNA damage | |
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| Reverse, excise or tolerate? | |
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| Mechanisms that reverse DNA damage | |
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| Photoreactivation | |
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| O6-methylguanine or O4-methylthymine methyltransferase | |
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| Mechanisms that excise DNA damage | |
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| UvrABC directed nucleotide excision repair | |
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| MutHLS methyl directed mismatch repair | |
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| Very short patch repair | |
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| Glycosylases | |
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| Uracil-N-glycosylase coupled with AP excision repair | |
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| Deaminated bases removed by DNA glycosylase | |
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| Alkylated bases removed by DNA glycosylase | |
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| MutM/MutY: oxidative damage | |
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| N-glycosylases specific for pyrimidine dimers | |
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| Mechanisms that tolerate DNA damage | |
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| Transdimer synthesis | |
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| Post replication/recombinational repair (PRR) | |
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| Introduction to the SOS regulon | |
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| Summary | |
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| Recombination | |
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| Homologous recombination | |
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| Models for homologous recombination | |
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| The Holliday or double-strand invasion model of recombination | |
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| An alternative to the Holliday model: the single-strand invasion model of Meselson and Radding | |
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| Further enzymatic considerations | |
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| Site-specific recombination | |
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| A typical site-specific recombinational event | |
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| Bacteriophage [lambda]: a model for site-specific recombination | |
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| Other microbial examples of site-specific recombination | |
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| Illegitimate recombination | |
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| Summary | |
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| Transposition | |
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| The structure of transposons | |
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| The frequency of transposition | |
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| The two types of transposition reactions | |
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| The transposition machinery | |
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| Accessory proteins encoded by the transposon | |
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| Accessory proteins encoded by the host | |
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| Non-replicative transposition | |
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| Replicative transposition | |
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| Does the formation of a cointegrate predict the transposition mechanism? | |
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| The fate of the donor site | |
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| Target immunity | |
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| Transposons as molecular tools | |
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| Summary | |
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| Bacteriophage | |
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| The structure of phage | |
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| The lifecycle of a bacteriophage | |
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| Lytic--lysogenic options | |
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| The [lambda] lifecycle | |
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| [lambda] adsorption | |
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| [lambda] DNA injection | |
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| Protecting the [lambda] genome in the bacterial cytoplasm | |
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| What happens to the [lambda] genome after it is stabilized? | |
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| [lambda] and the lytic--lysogenic decision | |
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| The [lambda] lysogenic pathway | |
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| The [lambda] lytic pathway | |
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| DNA replication during the [lambda] lytic pathway | |
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| Making [lambda] phage | |
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| Getting out of the cell--the [lambda] S and R proteins | |
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| Induction of [lambda] by the SOS system | |
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| Superinfection | |
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| Restriction and modification of DNA | |
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| The lifecycle of M13 | |
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| M13 adsorption and injection | |
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| Protection of the M13 genome | |
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| M13 DNA replication | |
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| M13 phage production and release from the cell | |
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| The lifecycle of P1 | |
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| Adsorption, injection, and protection of the genome | |
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| P1 DNA replication and phage assembly | |
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| The location of the P1 prophage in a lysogen | |
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| P1 transducing particles | |
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| The lifecycle of T4 | |
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| T4 adsorption and injection | |
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| T4rII mutations and the nature of the genetic code | |
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| Summary | |
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| Transduction | |
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| Generalized transduction vs. specialized transduction | |
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| P1 as a model for generalized transducing phage | |
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| Packaging the chromosome | |
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| Moving pieces of the chromosome from one cell to another | |
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| Identifying transduced bacteria: selection vs. screening | |
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| Carrying out a transduction | |
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| Uses for transduction | |
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| Two-factor crosses to determine gene linkage | |
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| Mapping the order of genes--three-factor crosses | |
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| Strain construction | |
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| Localized mutagenesis | |
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| Specialized transducing phage | |
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| Making merodiploids with specialized transducing phage | |
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| Moving mutations from plasmids to specialized transducing phage to the chromosome | |
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| Summary | |
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| Natural plasmids | |
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| Origins of replication | |
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| Plasmid copy number | |
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| Setting the copy number | |
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| Plasmid incompatibility | |
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| Plasmid amplification | |
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| Other genes that can be carried by plasmids | |
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| Plasmids can be circular or linear DNA | |
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| Broad host range plasmids | |
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| Moving plasmids from cell to cell | |
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| Summary | |
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| Conjugation | |
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| The F factor | |
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| The R factors | |
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| The conjugation machinery | |
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| Transfer of the DNA | |
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| Surface exclusion | |
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| F, Hfr, or F-prime | |
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| Formation of the Hfr | |
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| Transfer of DNA from an Hfr to another cell | |
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| Formation of F-primes | |
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| Transfer of F-primes from one cell to another | |
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| Genetic uses of F-primes | |
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| Genetic uses of Hfr strains--mapping genes on the E. coli chromosome using Hfr crosses | |
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| The 50% rule | |
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| Using several Hfr strains to cover the chromosome | |
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| Mobilization of non-conjugatible plasmids by R and F | |
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| Conjugation from prokaryotes to eukaryotes | |
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| Summary | |
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| Transformation | |
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| Natural competency | |
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| The process of natural transformation | |
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| The machinery of naturally transformable cells | |
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| Artificial transformation | |
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| Transformation as a genetic tool: gene mapping | |
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| Transformation as a molecular tool | |
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| Summary | |
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| Gene expression and regulation | |
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| The players in the regulation game | |
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| Operons and regulons | |
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| Repression of the lac operon | |
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| Activation of the lac operon by cyclic AMP and the CAP protein | |
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| Regulation of the tryptophan biosynthesis operon by attenuation | |
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| Regulation of the heat-shock regulon by an alternate sigma factor, mRNA stability, and proteolysis | |
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| Regulation of the SOS regulon by proteolytic cleavage of the repressor | |
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| Two-component regulatory systems: signal transduction and the cps regulon | |
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| Summary | |
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| Plasmids, bacteriophage, and transposons as tools | |
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| What is a cloning vector? | |
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| Why not use naturally occurring plasmids as vectors? | |
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| The importance of copy number | |
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| An example of how a cloning vector works--pBR322 | |
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| Multiple cloning sites | |
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| Determining which plasmids contain an insert | |
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| Expression vectors | |
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| Vectors for purifying the cloned gene product | |
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| Vectors for localizing the gene product | |
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| Vectors for studying gene expression | |
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| Shuttle vectors | |
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| Artificial chromosomes | |
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| Constructing phage vectors | |
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| Suicide vectors | |
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| Phage display vectors | |
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| Combining phage vectors and transposons | |
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| Summary | |
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| DNA cloning | |
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| Isolating DNA from cells | |
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| Plasmid DNA isolation | |
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| Chromosomal DNA isolation | |
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| Cutting DNA molecules | |
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| Type I restriction-modification systems | |
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| Type II restriction-modification systems | |
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| Type III restriction-modification systems | |
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| Restriction-modification as a molecular tool | |
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| Generate double-stranded breaks in DNA by shearing the DNA | |
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| Joining DNA molecules | |
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| Manipulating the ends of molecules | |
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| Visualizing the cloning process | |
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| Constructing libraries of clones | |
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| DNA detection--Southern blotting | |
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| DNA amplification--polymerase chain reaction | |
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| Adding novel DNA sequences to the ends of a PCR amplified sequence | |
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| Site-directed mutagenesis using PCR | |
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| Cloning and expressing a gene | |
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| DNA sequencing using dideoxy sequencing | |
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| DNA sequence searches | |
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| Summary | |
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| Bioinformatics and proteomics | |
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| Bioinformatics | |
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| Strategies for sequencing genomes | |
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| Bacterial genomes | |
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| Analyzing genomes | |
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| The E. coli K12 genome | |
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| Proteomics | |
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| Techniques for examining the proteome--SDS-PAGE and 2-D PAGE | |
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| Techniques for examining the proteome--microarray technology | |
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| Summary | |
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| Glossary | |
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| Further reading | |
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| Index | |