Table of Contents
PART I INTRODUCTION
Chapter 1. A BRIEF HISTORY

1.1 Transmission Genetics

Mendel's Laws of Inheritance
The Chromosome Theory of Inheritance
Genetic Recombination and Mapping
Physical Evidence for Recombination

1.2 Molecular Genetics

The Discovery of DNA
The Composition of Genes
The Relationship Between Genes and Proteins
Activities of Genes

How Genes Are Replicated
How Genes Direct the Production of Polypeptides
How Genes Accumulate Mutations
Gene Cloning

Box 1.1 Cell Structure
Box 1.2 Cell Cycle and Mitosis
Box 1.3 Meiosis
Chapter 2. THE MOLECULAR NATURE OF GENES

2.1 The Nature of Genetic Material

Transformation in Bacteria

DNA: The Transforming Material
Further Confirmation

The Chemical Nature of Polynucleotides

RNA

2.2 DNA Structure

Experimental Background
The Double Helix

2.3 Genes Made of RNA
2.4 Physical Chemistry of Nucleic Acids

A Variety of DNA Structures

A form
Separating the Two Strands of a DNA Double Helix
Reuniting the Separated DNA Strands
Hybridization of Two Different Polynucleotide Chains

DNAs of Various Sizes and Shapes

The Relationship between DNA Size and Genetic Capacity
DNA Content and the C-Value Paradox
Chapter 3. AN INTRODUCTION TO GENE FUNCTION

3.1 Storing Information

Overview of Gene Expression
Protein Structure
Protein Function

The Relationship between Genes and Proteins

Discovery of Messenger RNA
Transcription
Translation

Ribosomes: Protein-Synthesizing Machines
Transfer RNA: The Adapter Molecule
Initiation of Protein Synthesis

Translation Elongation
Termination of Translation and mRNA Structure

3.2 Replication
3.3 Mutations

Sickle-Cell Disease
PART II METHODS IN MOLECULAR BIOLOGY
Chapter 4. MOLECULAR CLONING METHODS

4.1 Gene Cloning

The Role of Restriction Endonucleases
Vectors

Plasmids as Vectors
Phages as Vectors

λ Phage Vectors
Cosmids
M13 phage vectors
Phagemids

Eukaryotic Vectors

Identifying a Specific Clone with a Specific Probe

Polynucleotide Probes

4.2 The Polymerase Chain Reaction (PCR)

cDNA Cloning

Using RT-PCR in cDNA Cloning

Rapid Amplification of cDNA Ends (RACE)

4.3 Methods of Expressing Cloned Genes

Expression Vectors

Expression Vectors with Strong Promoters
Inducible Expression Vectors
Expression Vectors that Produce Fusion Proteins
Eukaryotic Expression Systems

Other Eukaryotic Vectors
Using the Ti Plasmid to Transfer Genes to Plants

Box 4.1 Jurassic Park: More than a Fantasy?
Chapter 5. MOLECULAR TOOLS FOR STUDYING GENES AND GENE ACTIVITY

5.1 Molecular Separations

Gel Electrophoresis
Two-dimensional Gel Electrophoresis
Ion Exchange Chromatography
Gel Filtration Chromatography

5.2 Labeled Tracers

Autoradiography
Phosphorimaging
Liquid Scintillation Counting
Non-radioactive Tracers

5.3 Using Nucleic Acid Hybridization

Southern Blots: Identifying Specific DNA Fragments
DNA Fingerprinting and DNA Typing
Forensic Uses of DNA Fingerprinting and DNA Typing

Northern Blots: Measuring Gene Activity
In situ Hybridization: Locating Genes in Chromosomes

DNA Sequencing

The Sanger Chain-termination Sequencing Method
ATP (ddATP)
Maxam-Gilbert Sequencing

DNA Sequencing: The Ultimate in Genetic Mapping
Restriction Mapping
Protein Engineering with Cloned Genes: Site-directed Mutagenesis

5.4 Mapping and Quantifying Transcripts

S1 Mapping
Primer Extension
Run-off Transcription and G-less Cassette Transcription

5.5 Measuring Transcription Rates In Vivo

Nuclear Run-on Transcription
Reported Gene Transcription

5.6 Assaying DNA-Protein Interactions

Filter Binding
Gel Mobility Shift
DNase Footprinting
DMS Footprinting and Other Footprinting Methods

5.7 Knockouts
PART III TRANSCRIPTION IN PROKARYOTES Chapter 6. THE TRANSCRIPTION APPARATUS OF PROKARYOTES

6.1 RNA Polymerase Structure

σ as a Specificity Factor

6.2 Promoters

Binding of RNA Polymerase to Promoters
Promoter Structure

6.3 Transcription Initiation

The Function of σ

s Stimulates Transcription Initiation
Re-use of σ
Local DNA Melting at the Promoter

Specific Bases Involved in Polymerase Binding to Promoters
Structure of σ

Region 1
Region 2
Region 3
Region 4

Destabilizing Non-specific Polymerase-DNA Interactions
The Role of the α Subunit in UP Element Recognition

6.4 Elongation

Core Polymerase Functions

The Role of β in Phosphodiester Bond Formation
The Role of β and β in DNA Binding

The Role of α in Polymerase Assembly
Structure of the Elongation Complex

The RNA-DNA Hybrid
Structural Studies
Topology of Elongation

6.5 Termination of Transcription

Rho-independent (Intrinsic) Termination

Inverted Repeats and Hairpins
The Structure of an Intrinsic Terminator

A Model for Termination
Rho-dependent Termination

Rho Affects Chain Elongation but Not Initiation
Rho Causes Production of Shorter Transcripts
Rho Releases Transcripts from the DNA Template
The Mechanism of Rho
Chapter 7. OPERONS: FINE CONTROL OF PROKARYOTIC TRANSCRIPTION

7.1 The lac Operon

Negative Control of the lac Operon
Discovery of the Operon
Repressor-operator Interactions
The Mechanism of Repression
Positive Control of the lac Operon

Catabolite Activator Protein (CAP)

Mechanism of CAP Action

Recruitment

7.2 The mal Regulon

The Role of CAP in the mal Regulon
Evidence for the Role of CAP in the mal Regulon

7.3 The ara Operon

The ara Operon Repression Loop
Evidence for the ara Operon Repression Loop
Autoregulation of araC

7.4 The trp Operon

Tryptophan's Role in Negative Control of the trp Operon
Control of the trp Operon by Attenuation
Defeating Attenuation
Chapter 8. MAJOR SHIFTS IN PROKARYOTIC TRANSCRIPTION

8.1 Modification of the Host RNA Polymerase
8.2 The RNA Polymerase Encoded in Phage T7
8.3 Control of Transcription During Sporulation
8.4 Genes with Multiple Promoters

The B. subtilis spoVG Gene
The Anabaena Glutamine Sythetase Gene
The E. coli glnA Gene

8.5 The E. coli Heat Shock Genes
8.6 Infection of E. coli by Phage λ

Lytic Reproduction of Phage λ

Antitermination

Establishing Lysogeny
Autoregulation of the cI Gene During Lysogeny

RNA Polymerase-repressor Interaction

Determining the Fate of a λ Infection: Lysis or Lysogeny
Lysogen Induction
Chapter 9. DNA-PROTEIN INTERACTIONS IN PROKARYOTES

9.1 The Lambda Family of Repressors

High-resolution Analysis of l Repressor-operator Interactions

General Structural Features
Interactions with Bases
Amino Acid-DNA Backbone Interactions
Confirmation of Biochemical and Genetic Data

High-resolution Analysis of Phage 434 Repressor-operator Interactions

Repressor-operator Contacts

Contacts with Double Helical Backbone
Contacts with Base Pairs

Effects of DNA Conformation
Genetic Test of the Model

9.2 The trp Repressor

The Role of Tryptophan

trp Repressor-operator Interactions

9.3 General Considerations on Protein-DNA Interactions

Hydrogen Bonding Capabilities of the Four Different Base Pairs
The Role of DNA Shape in Specific Binding to Proteins
The Importance of Multimeric DNA-binding Proteins

9.4 DNA-binding Proteins: Action at a Distance

The gal Operon
Duplicated λ Operators
The lac Operon
Enhancers

Box 9.1 X-ray Crystallography
PART IV TRANSCRIPTION IN EUKARYOTES
Chapter 10. EUKARYOTIC RNA POLYMERASES AND THEIR PROMOTERS

10.1 Multiple Forms of Eukaryotic RNA Polymerase

Indications of Multiple Eukaryotic Polymerases
Separation of the Three Nuclear Polymerases
The Roles of the Three RNA Polymerases
RNA Polymerase Subunit Structures

Polymerase II Structure

Core Subunits
Common Subunits
Non-essential Subunits

Heterogeneity of the RPB1 Subunit
The RPB1 Subunit of α-Amanitin-Sensitivity
The Shape of RNA Polymerase II

10.2 Promoters

Class II Promoters

The TATA Box
Upstream Elements
Initiators and Downstream Elements

Class I Promoters
Class III Promoters

Class III Genes with Internal Promoters
Class III Genes with Polymerase II-like Promoters

10.3 Enhancers and Silencers

Enhancers
Silencers
Chapter 11. GENERAL TRANSCRIPTION FACTORS IN EUKARYOTES

11.1 Class II Factors

The Class II Preinitiation Complex
Structure and Function of THIID

The TATA Box-binding Protein (TBP)
The Versatility of TBP
The TBP-associated Factors (TAF_IIs)
The Non-universality of TAFs and TBP

Structure and Function of TFIIA and TFIIB
Structure and Function of TFIIF
Structure and Function of TFIIE and TFIIH
Elongation Factors

IIS Stimulates Elongation
IIS Stimulates Proofreading of Transcripts

The Polymerase II Holoenzyme

11.2 Class I Factors

SL1
UBF
Structure and Function of SL1

11.3 Class III Factors

TFIIIA
TFIIIB and C
The Role of TBP
Chapter 12. TRANSCRIPTION ACTIVATORS IN EUKARYOTES

12.1 Categories of Activators

DNA-binding Domains
Transcription-activation Domains

12.2 Structure of the DNA-Binding Motifs of Activators

Zinc Fingers

Finger Structure
Interaction with DNA
Comparison with Other DNA-binding Proteins

The GAL4 Protein

The DNA-binding Motif
The Dimerization Motif

The Nuclear Receptors
Homeodomains
The bZIP and bHLH Domains

12.3 Independence of the Domains of Activators
12.4 Functions of Transcription-Activation Domains

Recruitment of TFIID
Recruitment of TFIIB
Recruitment of Other General Transcription Factors
Recruitment of the Holoenzyme

12.5 Interaction Among Activators

Dimerization

The Jun-Fos Dimer

Action at a Distance
Multiple Enhancers
Architectural Transcription Factors
Insulators
Mediators

12.6 Regulation of Transcription Factors

Signal Transduction Pathways
Chapter 13. CHROMATIN STRUCTURE AND ITS EFFECTS ON TRANSCRIPTION
13.1 Histones
13.2 Nucleosomes

The Nucleosome Filament
The 30 nm Fiber
The Role of Histone H1 in Chromatin Folding
Higher-Order Chromatin Folding

13.3 Chromatin Structure and Gene Activity
The Effects of Histones on 5S rRNA Gene Transcription
The Effects of Histones on Transcription of Class II Genes
Core Histones
Histone H1
Nucleosome Positioning
Nucleosome-free Zones
DNase Hypersensitivity
Detecting Positioned Nucleosomes
Histone Acetylation
Histone Deacetylation
Chromatin Remodeling
Heterochromatin and Silencing
Nucleosomes and Transcription Elongation
PART V POST-TRANSCRIPTIONAL EVENTS
Chapter 14. POST-TRANSCRIPTIONAL EVENTS I: SPLICING

14.1 Genes in Pieces
Evidence for Split Genes
RNA Splicing
Splicing Signals

14.2 The Mechanism of Splicing of Nuclear mRNA Precursors
A Branched Intermediate
A Signal at the Branch
Spliceosomes
Snurps

U1 snRNP
U6 snRNP
U2 snRNP
U5 snRNP
U4 snRNP
A Summary of snRNP Involvement in mRNA Splicing

Spliceosome Assembly and Function

The Spliceosome Cycle
3¢ Splice Site Selection
Commitment
Commitment in Yeast

Alternative Splicing

14.3 Self-Splicing RNAs

Group I Introns
Group II Introns
14.4 tRNA Splicing
Chapter 15. POST-TRANSCRIPTIONAL EVENTS II: CAPPING AND POLYADENYLATION
15.1 Capping
Cap Structure
Cap Synthesis
Functions of Caps
Protection
Translatability
Transport of mRNA

15.2 Polyadenlation
Poly(A)
Function of Poly(A)
Protection of mRNA
Translatability of mRNA

Basic Mechanism of Polyadenylation
Polyadenylation Signals
Cleavage and Polyadenylation of a Pre-mRNA

Pre-mRNA Cleavage
Initiation of Polyadneylation
Elongation of Poly(A)

Poly(A) Polymerase
Turnover of Poly(A)

Cytospasmic Polyadenylation

15.3 The Effects of the Cap and Poly(A) on Splicing

Dependence of Splicing on the Cap
Effect of Poly(A) on Splicing
Chapter 16. POST-TRANSCRIPTIONAL EVENTS III: OTHER EVENTS

16.1 Ribosomal RNA Processing

Eukaryotic rRNA Processing
Prokaryotic rRNA Processing

16.2 Transfer RNA Processing

Cutting Apart Polycistronic Precursors
Forming Mature 5¢-ends
Forming Mature 3¢-ends

16.3 Trans-Splicing

The Mechanism of Trans-Splicing
Polycistronic Arrangement of Coding Regions in Trypanosomes

16.4 RNA Editing

Mechanism of Editing

16.5 Post-Transcriptional Control of Gene Expression

Casein mRNA Stability
Transferrin Receptor mRNA Stability

Iron Response Elements
The Rapid Turnover Determinant
TfR mRNA Stability
The TfR mRNA Degradation Pathway

16.6 Post-Transcriptional Gene Silencing (RNA Interference)
PART VI TRANSLATION
Chapter 17. THE MECHANISM OF TRANSLATION I: INITIATION

17.1 Initiation of Translation in Prokaryotes

tRNA Charging
Dissociation of Ribosomes
Formation of the 30S Initiation Complex

The First Codon and the First Aminoacyl tRNA
Binding mRNA and fMET-tRNAfMET to the 30S Ribosomal Subunit
Binding fMET-tRNAfMET to the 30S Initiation Complex

Formation of the 70S Initiation Complex
Summary of Initiation in Prokaryotes

17.2 Initiation in Eukaryotes

The Scanning Model of Initiation
Eukaryotic Initiation Factors
Overview of Translation in Initiation in Eukaryotes

Function of eIF4F
Function of eIF4A and -4B
Functions of eIF4G
Functions of eIF1 and eIF1A

17.3 Control of Initiation

Prokaryotic Translation Control
Eukaryotic Translation Control

Phosphorylation of Initiation Factor eIF2I
Phosphorylation of an eIF4E-Binding Protein
Simulation by an mRNA-Binding Protein
Chapter 18. THE MECHANISM OF TRANSLATION II: ELONGATION AND TERMINATION

18.1 The Direction of Polypeptide Synthesis and of mRNA Translation
18.2 The Genetic Code

Nonoverlapping Codons
No Gaps in the Code
The Triplet Code
Breaking the Codes
Unusual Base Pairs Between Codon and Anticodon
The (Almost) Universal Code

18.3 The Elongation Mechanism

Overview of Elongation
A Three-site Model of the Ribosome
Elongation Step 1: Binding an Aminoacyl-tRNA to the A Site of the Ribosome

Proof-reading

Elongation Step 2: Peptide Bond Formation
Elongation Step 3: Translocation

Three-nucleotide Movement of mRNA During Translocation
Role of GTP and EF-G

Structures of EF-Tu and EF-G

18.4 Termination

GTPases and Translation
Termination Codons
Stop Codon Suppression
Release Factors
Chapter 19. RIBOSOMES AND TRANSFER RNA

19.1 Ribosomes

Gross Ribosomal Structure
Fine Structure of the 70S Ribosome
Ribosome Composition
Ribosome Assembly
Fine Structure of the 30S Subunit

Interaction of the 30S Subunit with Antibiotics

Fine Structure of the 50S Subunit
Polysomes

19.2 Transfer RNA

The Discovery of tRNA
tRNA Structure
Recognition of tRNA by Aminoacyl-tRNA Synthetase: The Second Genetic Code

The Acceptor Stem
The Anticodon
Structures and Synthetase-tRNA Complexes

Proofreading and Editing by Aminoacyl-tRNA Synthetases
PART VII DNA REPLICATION, RECOMBINATION, AND TRANSPOSITION
Chapter 20. DNA REPLICATION I: BASIC MECHANISM AND ENZYMOLOGY

20.1 General Features of DNA Replication

Semiconservative Replication
Semidiscontinuous Replication
Priming of DNA Synthesis
Bidirectional Replication
Unidirectional Replication
Rolling Circle Replication

20.2 Enzymology of DNA Replication

Strand Separation

Helicase

Single-strand DNA-binding Proteins
Topoisomerases

The mechanism of type II topoisomerases

Three DNA Polymerases in E. coli

Pol I
Pol II and Pol III
Pol III

Fidelity of Replication
Mulitple Eukaryotic DNA Polumerases

20.3 DNA Damage and Repair

Damage Caused by Alkylation of Bases
Damage Caused by Ultraviolet Radiation
Damage Caused by Gamma and X Rays
Directly Undoing DNA Damage
Excision Repair in Prokaryotes

Base Excision Repair
Nucleotide Excision Repair

Excision Repair in Eukaryotes

Global Genome NER
Transcription-coupled NER

Mismatch Repair
Failure of Mismatch Repair in Humans
Coping with DNA Damage Without Repairing It

Recombination Repair
Error-Prone Bypass
Error-Prone and Error-Free Bypass in Humans
Chapter 21. DNA REPLICATION II: DETAILED MECHANISM

21.1 Speed of Replication
21.2 Initiation

Priming in E. coli

The Origin of Replication in E. coli

Priming in Eukaryotes

The Origin of Replication in SV40
The Origin of Replication in Yeast

21.3 Elongation

The Pol III Holoenzyme and Processivity of Replication

The β Clamp
The Clamp Loader
Lagging Strand Synthesis

21.4 Termination

Decatenation: Disentangling Daughter DNAs
Termination in Eukaryotes

Telomere Maintenance
Telomere Structure

Box 21.1 Telomeres, the Hayflick Limit, and Cancer
Chapter 22. HOMOLOGOUS RECOMBINATION

22.1 Models for Homologous Recombination

The Holliday Model
The Meselson-Radding Model
The RecBCD Pathway

22.2 Experimental Support for the RecBCD Pathway

RecA

Presynapsis
Synapsis: Alignment of Complementary Sequences

Double Helix

Postsynapsis: Strand Exchange

RecBCD
RuvA and RuvB
RuvC

22.3 Meiotic Recombination

The Mechanism of Meiotic Recombination: Overview
The Double-stranded DNA Break
Creation of Single-stranded Ends at DSBs

22.4 Gene Conversion
Chapter 23. SITE-SPECIFIC RECOMBINATION AND TRANSPOSITION

23.1 Site Specific Recombination

Lambda Phage Integration and Excision

The DNA Sites
Mechanism

Bacterial Use of Site-Specific Recombination

23.2 Bacterial Transposons

Discovery of Bacterial Transposons
Insertion Sequences: The Simplest Transposons
More Complex Transposons
Mechanisms of Transposition

23.3 Eukaroytic Transposons

The First Examples of Transposable Elements, Ds and Ac of Maize
P Elements
Rearrangement of Immunoglobulin Genes

Recombination Signals
The Recombinase
Mechanism of V(D)J Recombination

Retrotransposons

Retroviruses
Retrotransposons

Non-LRT Retrotransposons
Non-Autonomous Retrotransposons
Group II Introns
Part VIII GENOMES
Chapter 24. GENOMICS

24.1 The First Sequenced Genomes

The Human Genome Project
Vectors for Large-Scale Genome Projects

Yeast Artificial Chromosomes
Bacterial Artificial Chromosomes

The Clone-by-Clone Strategy

Restriction Fragment Length Polymorphisms (RFLPs)
Variable Number of Tandem Repeats (VNTRs)
Sequence Tagged Sites (STSs)
Microsatellites
Radiation Hybrid Mapping

Shotgun Sequencing
Sequencing Standards
Progress in Sequencing the Human Genome

Chromosome 22
Chromosome 21

24.2 Applications of Genomics

Functional Genomics Techniques

DNA Microarrays and Microchips
Serial Analysis of Gene Expression (SAGE)

Positional Cloning

Exon Traps
CG Islands

Applications of Functional Genomics

Huntington's Disease
Cystic Fibrosis

Other Applications
Bioinformatics
Proteomics

Protein Separations
Protein Analysis

Box 24.1 Problems in Genetic Screening

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		Molecular Biology 2nd Edition Robert F. Weaver
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