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MOLECULAR
BIOLOGY
°FTHE GENE SIXTH EDITIO N
James D . Watson
Tania A. Baker
Stephen P. Bell
Alexander Gan n
Michael Levine
Richard Losick
PART 1
CHEMISTRY AND GENETICS, 1
1 The Mendelian View of the World, 5
2 Nucleic Acids Convey Genetic Information, 1 93 The Importance of Weak Chemical Interactions, 4 34 The Importance of High-Energy Bonds, 5 7
5 Weak and Strong Bonds Determine Macromolecular Structure, 7 1
PART 2
MAINTENANCE OF THE GENOME, 9 5
6 The Structures of DNA and RNA, 10 1
7 Genome Structure, Chromatin, and the Nucleosome, 13 5
8 The Replication of DNA, 19 59 The Mutability and Repair of DNA, 25 7
10 Homologous Recombination at the Molecular Level, 28 311 Site-Specific Recombination and Transposition of DNA, 31 9
PART 3
EXPRESSION OF THE GENOME, 37 1
12 Mechanisms of Transcription, 37 713 RNA Splicing, 41 514 Translation, 45 7
15 The Genetic Code, 52 1
PART 4
REGULATION, 54 1
16 Transcriptional Regulation in Prokaryotes, 54 717 Transcriptional Regulation in Eukaryotes, 58 9
18 Regulatory RNAs, 63 319 Gene Regulation in Development and Evolution, 66 120 Genome Analysis and Systems Biology, 70 3
PART 5
METHODS, 73 3
20 Techniques of Molecular Biology, 73 9
21 Model Organisms, 78 3
Index, 819
PART 1
CHEMISTRY AND GENETICS, 1
CHAPTER 1 • The Mendelian View of the World, 5
Mendel's Discoveries, 6
Chromosome Mapping, 1 2The Principle of Independent Segregation, 6
The Origin of Genetic Variability through Mutations, 1 5ADM( 11) Cov(rPr) Box 1-1 Mendelian Laws, 6
Some Alleles Are neither Dominant nor Recessive, 8
Early Speculations about What Genes Are and Ho w
Principle of Independent Assortment, 8
They Act, 1 6
Chromosomal Theory of Heredity, 8
Preliminary Attempts to Find a Gene-Protei nRelationship, 1 6
Gene Linkage and Crossing Over, 9
SUMMARY, 1 7KEY EXPERIMENTS Box 1-2 Genes Are Linked to
BIBLIOGRAPHY, 1 8Chromosomes, 10
CHAPTER 2 • Nucleic Acids Convey Genetic Information, 1 9
Avery's Bombshell : DNA Can Carry Genetic
The Central Dogma, 3 2Specificity, 20
The Adaptor Hypothesis of Crick, 3 2
Viral Genes Are Also Nucleic Acids, 21
Discovery of Transfer RNA, 32
The Double Helix, 21
The Paradox of the Nonspecific-Appearing Ribosomes, 3 3
Finding the Polymerases That Make DNA, 23
Discovery of Messenger RNA (mRNA), 3 4
KEY EXPERIMENTS Box 2 1 Chargaff's Rules, 24
Enzymatic Synthesis of RNA upon DNA Templates, 35
Experimental Evidence Favors Strand Separation during
Establishing the Genetic Code, 3 6
DNA Replication, 25
Establishing the Direction of Protei n
The Genetic Information within DNA Is Conveyed by the
Synthesis, 3 8
Sequence of Its Four Nucleotide Building Blocks, 28
Start and Stop Signals Are Also Encoded within DNA, 39
KEY EXPERIMENTS Box 2-2, Evidence That Genes Control
The Era of Genomics, 3 9Amino Acid Sequences in Proteins, 29
DNA Cannot Be the Template That Directly Orders Amino
SUMMARY, 40
Acids during Protein Synthesis, 30
BIBLIOGRAPHY, 4 1
RNA Is Chemically Very Similar to DNA, 30
CHAPTER 3 • The Importance of Weak Chemical Interactions, 43
Characteristics of Chemical Bonds, 43
Some Ionic Bonds Are Hydrogen Bonds, 50Chemical Bonds Are Explainable in Quantum-Mechanical
Weak Interactions Demand Complementary Molecula rTerms, 44
Surfaces, 5 1
Chemical-Bond Formation Involves a Change in the Form of
Water Molecules Form Hydrogen Bonds, 5 1Energy, 45
Weak Bonds between Molecules in Aqueous Solutions, 5 1Equilibrium between Bond Making and Breaking, 45
Organic Molecules That Tend to Form Hydrogen Bonds Are
The Concept of Free Energy, 46
Water Soluble, 52
Key Is Exponentially Related to AG, 46
ADVANCED CONCEPTS Box 3-1 The Uniqueness of MolecularShapes and the Concept of Selective Stickiness, 5 3
Covalent Bonds Are Very Strong 46
Hydrophobic "Bonds" Stabilize Macromolecules, 5 4Weak Bonds in Biological Systems, 47
The Advantage of AG between 2 and 5 kcal/mole, 55
Weak Bonds Have Energies between 1 and 7 kcallmol, 47
Weak Bonds Attach Enzymes to Substrates, 55
Weak Bonds Are Constantly Made and Broken at
Weak Bonds Mediate Most Protein-DNA an dPhysiological Temperatures, 47
Protein-Protein Interactions, 55The Distinction between Polar and Nonpolar Molecules, 47
SUMMARY, 56van der Waals Forces, 48
BIBLIOGRAPHY, 56Hydrogen Bonds, 49
CHAPTER 4 • The Importance of High-Energy Bonds, 5 7
Molecules That Donate Energy Are Thermodynamically
Activation of Precursors in Group Transfer Reactions, 63Unstable, 57
ATP Versatility in Group Transfer, 64
Enzymes Lower Activation Energies in Biochemical
Activation of Amino Acids by Attachment of AMP, 6 5
Reactions, 59
Nucleic Acid Precursors Are Activated by the Presence o f0-0, 66
Free Energy in Biomolecules, 60 The Value of 0 -- 0 Release in Nucleic Acid Synthesis, 6 6High-Energy Bonds Hydrolyze with Large Negative AG, 60
0 -O Splits Characterize Most Biosynthetic Reactions, 6 7
High-Energy Bonds in Biosynthetic Reactions, 62
SUMMARY, 68Peptide Bonds Hydrolyze Spontaneously, 62
BIBLIOGRAPHY, 69Coupling of Negative with Positive AG, 6 3
CHAPTER 5 • Weak and Strong Bonds Determine Macromolecular Structure, 7 1
Higher-Order Structures Are Determined by Intra- and
Most Proteins Are Modular, Containing Two or Thre eIntermolecular Interactions, 71
Domains, 82DNA Can Form a Regular Helix, 71
Proteins Are Composed of a Surprisingly Small Number of
RNA Forms a Wide Variety of Structures, 73
Structural Motifs, 82
Chemical Features of Protein Building Blocks, 73
AI> L :' \. (
( r, (
I?
>-' Large Proteins Are Often
The Peptide Bond, 75
Constructed of Several Smaller Polypeptid e
There Are Four Levels of Protein Structure, 75
Chains, 83
a Helices and ß Sheets Are the Common Forms of
Different Protein Functions Arise from Various Domai n
Secondary Structure, 76
Combinations, 8 4
TECHNIQUES Box 5-1 Determination of Protein
Weak Bonds Correctly Position Proteins along DNA an d
Structure, 78
RNA Molecules, 85Proteins Scan along DNA to Locate a Specific DNA-Bind ing
The Specific Conformation of a Protein Results from Its
Site, 8 7Pattern of Hydrogen Bonds, 80
a Helices Come Together to Form Coiled-Coils, 80
Diverse Strategies for Protein Recognition of RNA, 88
Allostery : Regulation of a Protein's Function by Changing
Not All Regulation of Proteins Is Mediated byAllosteric
Its Shape, 90
Events, 93
The Structural Basis of Allosteric Regulation Is Known for
SUMMARY, 93Examples Involving Small Ligands, Protein-Protein
BIBLIOGRAPHY, 94Interactions, and Protein Modification, 9 0
PART 2
MAINTENANCE OF THE GENOME, 95
CHAPTER 6 • The Structures of DNA and RNA, 10 1
DNA Structure,1 02
Nucleosomes Introduce Negative Supercoiling i n
DNA Is Composed of Polynucleotide Chains, 102
Eukaryotes, 12 0
Each Base Has Its Preferred Tautomeric Form, 104
Topoisomerases Can Relax Supercoiled DNA, 72 1
The Two Strands of the Double Helix Are Held Together by
Prokaryotes Have a Special Topoisomerase That Introduce s
Base Pairing in an Antiparallel Orientation, 105
Supercoils into DNA, 12 1
The Two Chains of the Double Helix Have Complementary Topoisomerases Also Unknot and Disentangle DNA
Sequences, 106
Molecules, 12 1
Hydrogen Bonding Is Important for the Specificity of Base
Topoisomerases Use a Covalent Protein-DNA Linkage to
Pairing 106
Cleave and Rejoin DNA Strands, 12 3
Bases Can Flip Out from the Double Helix, 107
Topoisomerases Form an Enzyme Bridge and Pass DNASegments through Each Other, 123
DNA Is Usually a Right-Handed Double Helix, 107DNA Topoisomers Can Be Separated by Electrophoresis, 12 5
The Double Helix Has Minor and Major Grooves, 108
Ethidium Ions Cause DNA to Unwind, 126KEY EXPERIMENTS Box 6-1 DNA Has 10.5 Base Pairs per Tur n
of the Helix in Solution : The Mica Experiment, 108
RNA Structure, 12 7The Major Groove Is Rich in Chemical Information, 109
RNA Contains Ribose and Uracil and Is Usually Single-The Double Helix Exists in Multiple Conformations, 110
Stranded, 12 7
KEY EXPERIMENTS Box 6-2 How Spots on an X-ray Film Reveal
RNA Chains Fold Back on Themselves to Form Local Regionsthe Structure of DNA, 112
of Double Helix Similar to A-Form DNA, 12 7
DNA Can Sometimes Form a Left-Handed Helix, 113
KEY EXPERIMENTS Box 6-3 Proving that DNA Has a Helica l
DNA Strands Can Separate (Denature) and Reassociate, 113
Periodicity of about 10.5 Base Pairs per Turn from the
Some DNA Molecules Are Circles, 116
Topological Properties of DNA Rings, 12 8
RNA Can Fold Up into Complex Tertiary Structures, 12 9DNA Topology, 117
Some RNAs Are Enzymes, 13 0Linking Number Is an Invariant Topological Property of
The Hammerhead Ribozyme Cleaves RNA by the Formatio nCovalently Closed, Circular DNA, 117
of a 2', 3' Cyclic Phosphate, 13 1Linking Number Is Composed of Twist and Writhe, 117
Did Life Evolve from an RNA World?, 132Lk° Is the Linking Number of Fully Relaxed cccDNA under
SUMMARY, 132Physiological Conditions, 11 9
DNA in Cells Is Negatively Supercoiled, 120
BIBLIOGRAPHY, 133
CHAPTER 7 • Genome Structure, Chromatin, and the Nucleosome, 13 5
Genome Sequence and Chromosome Diversity, 136
The E . coli Genome Is Composed Almost Entirely o f
Chromosomes Can Be Circular or Linear, 136
Genes, 14 0
Every Cell Maintains a Characteristic Number of
More Complex Organisms Have Decreased Gene Density,
Chromosomes, 137
1 `w
Genome Size Is Related to the Complexity of the
Genes Make Up Only a Small Proportion of the Eukaryoti c
Organism, 139
Chromosomal DNA, 141
The Majority of Human Intergenic Sequences Are Composed
Higher-Order Chromatin Structure, 16 9of Repetitive DNA, 143
Heterochromatin and Euchromatin, 16 9
Chromosome Duplication and Segregation, 144
Histone H1 Binds to the Linker DNA between Nucleosomes,
Eukaryotic Chromosomes Require Centromeres, Telomeres,
169
and Origins of Replication to Be Maintained during Cell
Nucleosome Arrays Can Form More Complex Structures :Division, 144
The 30-nm Fiber, 170
Eukaryotic Chromosome Duplication and Segregation Occur
The Histone Amino-Terminal Tails Are Required for th ein Separate Phases of the Cell Cycle, 147
Formation of the 30-nm Fiber, 172
Chromosome Structure Changes as Eukaryotic Cells
Further Compaction of DNA involves Large Loops ofDivide, 149
Nucleosomal DNA, 172
Sister-Chromatid Cohesion and Chromosome Condensation
Histone Variants Alter Nucleosome Function, 174Are Mediated by SMC Proteins, 150
Regulation of Chromatin Structure, 17 4Mitosis Maintains the Parental Chromosome Number, 152
The Interaction of DNA with the Histone Octamer I sDuring Gap Phases, Cells Prepare for the Next Cell Cycle
Dynamic, 174Stage and Check That the Previous Stage Is CompletedCorrectly, 152
Nucleosome-Remodeling Complexes Facilitate Nucleosom e
Meiosis Reduces the Parental Chromosome Number, 154
Movement, 175Some Nucleosomes Are Found in Specific Positions :
Different Levels of Chromosome Structure Can Be Observed
Nucleosome Positioning, 17 9by Microscopy, 156
KEY EXPERIMENTS Box 7-3 Determining Nucleosome Positio n
The Nucleosome, 157
in the Cell, 180
Nucleosomes Are the Building Blocks of Chromosomes, 157
Modification of the Amino-Terminal Tails of the Histone s
KEY EXPERIMENTS Box 7-1 Micrococcal Nuclease and the
Alters Chromatin Accessibility, 182
DNA Associated with the Nucleosome, 158
Protein Domains in Nucleosome-Remodeling an d
Histones Are Small, Positively Charged Proteins, 159
-Modifying Complexes Recognize Modified Histones, 18 4
The Atomic Structure of the Nucleosome, 160
Specific Enzymes Are Responsible for Histone Modification ,185
Histones Bind Characteristic Regions of DNA within the
Nucleosome Modification and Remodeling Work TogetherNucleosome, 162
Many DNA Sequence-Independent Contacts Mediate
to Increase DNA Accessibility, 18 6
the Interaction between the Core Histones and
Nucleosome Assembly, 187DNA, 162
Nucleosomes Are Assembled Immediately after DNAThe Histone Amino-Terminal Tails Stabilize DNA Wrapping
Replication, 18 7around the Octamer, 165
Assembly of Nucleosomes Requires Histone "Chaperones, "Wrapping of the DNA around the Histone Protein Core
189Stores Negative Superhelicity, 166
SUMMARY, 192KEY EXPERIMENTS Box 7-2 Nucleosomes and Superhelica l
Density, 166
BIBLIOGRAPHY, 193
CHAPTER 8 • The Replication of DNA, 19 5
The Chemistry of DNA Synthesis, 196
Primer:Template Junction, 202DNA Synthesis Requires Deoxynucleoside Triphosphates
MEDICAL CONNECTIONS Box 8-2 Anticancer and Antivira land a Primer:Template Junction, 196
Agents Target DNA Replication, 20 3DNA Is Synthesized by Extending the 3 ' End of the
DNA Polymerases Are Processive Enzymes, 20 7Primer, 197
Exonucleases Proofread Newly Synthesized DNA, 20 8Hydrolysis of Pyrophosphate Is the Driving Force for DNA
The Replication Fork, 20 9Synthesis, 198Both Strands of DNA Are Synthesized Together at th e
The Mechanism of DNA Polymerase, 198
Replication Fork, 209DNA Polymerases Use a Single Active Site to Catalyze DNA
The Initiation of a New Strand of DNA Requires an RNASynthesis, 198
Primer, 210TECHNIQUES Box 8-1 Incorporation Assays Can Be
RNA Primers Must Be Removed to Complete DNAUsed to Measure Nucleic Acid and Protein
Replication, 21 1Synthesis, 200
DNA Helicases Unwind the Double Helix in Advance of th eDNA Polymerases Resemble a Hand That Grips the
Replication Fork, 211
TECHNIQUES Box 8-3 Determining the Polarity of a DNA
Protein-Protein and Protein-DNA Interactions Direct th eHelicase, 212
Initiation Process, 235
DNA Helicase Pulls Single-Stranded DNA through a Central
Al .( /I) (()\( /PT.s Box 8-6 The Replication FactoryProtein Pore, 214
Hypothesis, 23 7
Single-Stranded DNA-Binding Proteins Stabilize ssDNA prior
Eukaryotic Chromosomes Are Replicated Exactly Once pe rto Replication, 215
Cell Cycle, 23 9
Topoisomerases Remove Supercoils Produced by DNA
Prereplicative Complex Formation Is the First Step in th eUnwinding at the Replication Fork, 216
Initiation of Replication in Eukaryotes, 24 0
Replication Fork Enzymes Extend the Range of DNA
Pre-RC Formation and Activation Are Regulated to AllowPolymerase Substrates, 217
Only a Single Round of Replication during Each Cel lCycle, 24 1
The Specialization of DNA Polymerases, 218
Similarities between Eukaryotic and Prokaryotic DNADNA Polymerases Are Specialized for Different Roles in the
Replication Initiation, 24 4Cell, 218
gill qv(II ( O,vr i'l :, S( 11 li-7 E . colt DNA Replication IsSliding Clamps Dramatically Increase DNA Polymerase
Regulated by DnaA•ATP Levels and SeqA, 24 4Processivity, 219
Sliding Clamps Are Opened and Placed on DNA by Clamp
Finishing Replication, 246
Loaders, 222
Type II Topoisomerases Are Required to Separate Daughter
AEA' ;\'CfD CC)NCFPln Box 8-4 ATP Control of Protein
DNA Molecules, 24 6
Function : Loading a Sliding Clamp, 223
I.9gging-Strand Synthesis Is Unable to Copy the Extreme Ends
DNA Synthesis at the Replication Fork, 225
of Linear Chromosomes, 24 7
Telomerase Is a Novel DNA Polymerase That Does No tInteractions between Replication Fork Proteins Form the E .
Require an Exogenous Template, 248colt Replisome, 228
Telomerase Solves the End Replication Problem by Extendin g
Initiation of DNA Replication, 230
the 3' End of the Chromosome, 25 0
Specific Genomic DNA Sequences Direct the Initiation of
MEDICAL CONNECTIONS Box 8-8 Aging, Cancer, and the
DNA Replication, 230
Telomere Hypothesis, 25 1
The Replicon Model of Replication Initiation, 230
Telomere-Binding Proteins Regulate Telomerase Activity andTelomere Length, 252
Replicator Sequences Include Initiator Binding Sites andEasily Unwound DNA, 231
Telomere-Binding Proteins Protect Chromosome
KEY EXPERIMENTS Box 8-5 The Identification of Origins of
Ends, 253
Replication and Replicators, 232
SUMMARY, 255
Binding and Unwinding : Origin Selection and Activation
BIBLIOGRAPHY, 256
by the Initiator Protein, 235
CHAPTER 9 • The Mutability and Repair of DNA, 25 7
Replication Errors and Their Repair, 258
Direct Reversal of DNA Damage, 270
The Nature of Mutations, 258
Base Excision Repair Enzymes Remove Damaged Bases by aSome Replication Errors Escape Proofreading, 259
Base Flipping Mechanism, 270
MEDICAL CONNECTIONS Box 9-1 Expansion of Triple Repeats
Nucleotide Excision Repair Enzymes Cleave Damaged DN ACauses Disease, 259
on Either Side of the Lesion, 273
Mismatch Repair Removes Errors That Escape
Recombination Repairs DNA Breaks by Retrieving SequenceProofreading, 260
Information from Undamaged DNA, 275
DSBs in DNA Are Also Repaired by Direct Joining of Broke nDNA Damage, 265
Ends, 275DNA Undergoes Damage Spontaneously from Hydrolysis
MEDICAL CONNECTIONS Box 9-3 Nonhomologous En dand Deamination, 265
Joining, 276DNA Is Damaged by Alkylation, Oxidation, and
Translesion DNA Synthesis Enables Replication to ProceedRadiation, 265
across DNA Damage, 278MEDICAL CONNECTIONS Box 9-2 The Ames T e s t , 266
vx is n ; ' o'r 13rn. 9-4 The Y Family of DNAMutations Are Also Caused by Base Analogs and
Polymerases, 280Intercalating Agents, 268
SUMMARY, 28 1
Repair of DNA Damage, 269
BIBLIOGRAPHY, 282
CHAPTER 10 • Homologous Recombination at the Molecular Level, 283
DNA Breaks Are Common and Initiate Recombination, 284
Homologous Recombination Is Required for Chromosom eModels for Homologous Recombination, 284
Segregation during Meiosis, 304
Strand Invasion Is a Key Early Step in Homologous
Programmed Generation of Double-Stranded DNA Break s
Recombination, 286
Occurs during Meiosis, 305
Resolving Holliday Junctions Is a Key Step to Finishing
MRX Protein Processes the Cleaved DNA Ends fo r
Genetic Exchange, 288
Assembly of the RecA-like Strand-Exchange
The Double-Strand Break-Repair Model Describes Many
Proteins, 30 7
Recombination Events, 288
Dmc1 Is a RecA-like Protein That Specifically Functions inMeiotic Recombination, 308
Homologous Recombination Protein Machines, 291
Many Proteins Function Together to Promote Meioti cc
hrr 10-1 How to Resolve a
Recombination, 308
Recombination Intermediate with Two Holliday
MEDICAL CONNECTIONS Box 10-2 The Product of the TumorJunctions, 292
Suppressor Gene BRCA2 Interacts with Rad5I ProteinThe RecBCD Helicase/Nuclease Processes Broken DNA
and Controls Genome Stability, 309
Molecules for Recombination, 293
Chi Sites Control RecBCD, 296
Mating Type Switching, 31 0
RecA Protein Assembles on Single Stranded DNA and
Mating-Type Switching Is Initiated by a Site-Specific Double -
Promotes Strand Invasion, 297
Strand Break, 31 1
Mating-Type Switching Is a Gene Conversion Event and No tNewly Base-Paired Partners Are Established within the RecA
Associated with Crossing Over, 312Filament, 299
RecA Homologs Are Present in All Organisms, 301
Genetic Consequences of the Mechanism o f
The RuvAB Complex Specifically Recognizes Holliday
Homologous Recombination, 31 4Junctions and Promotes Branch Migration, 301
One Cause of Gene Conversion Is DNA Repair during
RuvC Cleaves Specific DNA Strands at the Holliday Junction
Recombination, 315
to Finish Recombination, 302
SUMMARY, 31 6Homologous Recombination in Eukaryotes, 303
BIBLIOGRAPHY, 31 7Homologous Recombination Has Additional Functions i n
Eukaryotes, 303
CHAPTER 11 • Site-Specific Recombination and Transposition of DNA, 31 9
Conservative Site-Specific Recombination, 320
The Hin Recombinase Inverts a Segment of DNA Allowing
Site-Specific Recombination Occurs at Specific DNA
Expression of Alternative Genes, 33 1
Sequences in the Target DNA, 320
Hin Recombination Requires a DNA Enhancer, 332
Site-Specific Recombinases Cleave and Rejoin DNA Using a
Recombinases Convert Multimeric Circular DNA Molecule sCovalent Protein-DNA Intermediate, 322
into Monomers, 333
Serine Recombinases Introduce Double-Strand Breaks in DNA
There Are Other Mechanisms to Direct Recombination t oand Then Swap Strands to Promote Recombination, 324
Specific Segments of DNA, 334
Structure of the Serine Recombinase-DNA Complex
Transposition, 33 4Indicates That Subunits Rotate to Achieve Stran dExchange, 325
Some Genetic Elements Move to New Chromosoma l
Tyrosine Recombinases Break and Rejoin One Pair of DNA
Locations by Transposition, 334
Strands at a Time, 326
ADVANCED CONCEPTS Box 11-2 The Xer Recombinase
Structures of Tyrosine Recombinases Bound to DNA Reveal
Catalyzes the Monomerization of Bacteria l
the Mechanism of DNA Exchange, 327
Chromosomes and of Many Bacterial Plasmids, 335
There Are Three Principal Classes of TransposableMEDICAL . CONNECTIONS Box 11-1 Application of Site-Specific Elements, 338Recombination to Genetic Engineering, 327
DNA Transposons Carry a Transposase Gene, Flanked byBiological Roles of Site-Specific Recombination, 328
Recombination Sites, 339
? Integrase Promotes the Integration and Excision of a Viral
Transposons Exist as Both Autonomous an dGenome into the Host-Cell Chromosome, 329
Nonautonomous Elements, 339
Bacteriophage 7l Excision Requires a New DNA-Bending
Virus-like Retrotransposons and RetrovirusesProtein, 331
Carry Terminal Repeat Sequences and Two Genes
Important for Recombination, 340
KEY EXPERIMENTS Box 11-4 Maize Elements and th e
Poly-A Retrotransposons Look Like Genes, 340
Discovery of Transposons, 356
DNA Transposition by a Cut-and-Paste Mechanism, 340
Tn 10 Transposition Is Coupled to Cellular DNA Replication ,
The Intermediate in Cut-and-Paste Transposition Is Finished
358
by Gap Repair, 342
Phage Mu Is an Extremely Robust Transposon, 35 9
There Are Multiple Mechanisms for Cleaving the
Mu Uses Target Immunity to Avoid Transposing into Its Ow n
Nontransferred Strand during DNA Transposition, 343
DNA, 359
DNA Transposition by a Rep/icative Mechanism, 345
\ ' ! ' ? N 'l t l'TS Box 11-5 Mechanism of Transposition
Virus-like Retrotransposons and Retroviruses Move Using an
Target Immunity, 36 1
RNA Intermediate, 347
Tc1/mariner Elements Are Extremely Successful DNA
Bo.\ 11-3 The Pathway of Retroviral
Elements in Eukaryotes, 362
cDNA Formation, 349
Yeast Ty Elements Transpose into Safe Havens in th e
DNA Transposases and Retroviral lntegrases Are Members of
Genome, 362
a Protein Superfamily, 351
LINEs Promote Their Own Transposition and Even Transpos e
Poly-A Retrotransposons Move by a "Reverse Splicing"
Cellular RNAs, 36 3
Mechanism, 352
V(D)J Recombination, 36 5
Examples of Transposable Elements and Their
The Early Events in V(D)J Recombination Occur by a
Regulation, 354
Mechanism Similar to Transposon Excision, 36 7
IS4-Family Transposons Are Compact Elements with Multiple
SUMMARY, 369Mechanisms for Copy Number Control, 355
BIBLIOGRAPHY, 369
PART 3
EXPRESSION OF THE GENOME, 37 1
C H APT E R 12 • Mechanisms of Transcription, 37 7
RNA Polymerases and the Transcription Cycle, 378
a!nAwoI ) .,\(1!'r, Box 12-2 The Single-Subunit RNA
RNA Polymerases Come in Different Forms but Share Many
Polymerases, 393
Features, 378
RNA Polymerase Can Become Arrested and Nee d
Transcription by RNA Polymerase Proceeds in a Series of
Removing, 394
Steps, 380
Transcription Is Terminated by Signals within the RN A
Transcription Initiation Involves Three Defined Steps, 382
Sequence, 39 4
The Transcription Cycle in Bacteria, 383
Transcription in Eukaryotes, 396
Bacterial Promoters Vary in Strength and Sequence but Have
RNA Polymerase II Core Promoters Are Made Up of
Certain Defining Features, 383
Combinations of Four Different Sequence Elements, 39 7
The a Factor Mediates Binding of Polymerase to the
RNA Polymerase Il Forms a Preinitiation Complex with
Promoter, 384
General Transcription Factors at the Promoter, 39 8
Transition to the Open Complex Involves Structural
Promoter Escape Requires Phosphorylation of the
Changes in RNA Polymerase and in the Promoter
Polymerase ''Tail," 39 8
DNA, 386
TBP Binds to and Distorts DNA Using a I Sheet Inserted into
TECHNIQUES Box 12-1 Consensus Sequences, 388
the Minor Groove, 400
Transcription Is Initiated by RNA Polymerase without the
The Other General Transcription Factors Also Have Specifi c
Need for a Primer, 388
Roles in Initiation, 40 1
During Initial Transcription, RNA Polymerase Remains
In Vivo, Transcription Initiation Requires Additional Proteins ,Stationary and Pulls Downstream DNA into Itself, 389
Including the Mediator Complex, 402
Promoter Escape Involves Breaking Polymerase-Promoter
Mediator Consists of Many Subunits, Some Conserved fro m
Interactions and Polymerase Core-a Interactions, 390
Yeast to Human, 403
The Elongating Polymerase Is a Processive Machine That
A New Set of Factors Stimulate Pol II Elongation and RNA
Synthesizes and Proofreads RNA, 391
Proofreading, 404
Elongating RNA Polymerase Must Deal with Histones in Its
RNA Pol I and Pol III Recognize Distinct Promoters, UsingPath, 405
Distinct Sets of Transcription Factors, but Still Requir eElongating Polymerase Is Associated with a New Set of
TBP, 41 0Protein Factors Required for Various Types of RNA
Pol Ill Promoters Are Found Downstream of TranscriptionProcessing, 406
Start Site, 412Transcription Termination Is Linked to RNA Destruction by a
SUMMARY, 41 3Highly Processive RNase, 410BIBLIOGRAPHY, 41 4
Transcription by RNA Polymerases I and III, 41 0
CHAPTER 13 • RNA Splicing, 41 5
The Chemistry of RNA Splicing, 417
Several Mechanisms Exist to Ensure Mutually ExclusiveSequences within the RNA Determine Where Splicing
Splicing, 43 5Occurs, 417
The Curious Case of the Drosophila Dscam Gene : MutuallyThe Intron Is Removed in a Form Called a Lariat as the
Exclusive Splicing on a Grand Scale, 43 6Flanking Exons Are Joined, 418
Mutually Exclusive Splicing of Dscam Exon 6 Cannot B eKEY EXPERIMENTS Box 13-1 Adenovirus and the Discovery
Accounted for by Any Standard Mechanism and Instea dof Splicing, 419
Uses a Novel Strategy, 43 7
Exons from Different RNA Molecules Can Be Fused by
Alternative Splicing Is Regulated by Activators an dtrans-Splicing, 421
Repressors, 43 9Regulation of Alternative Splicing Determines the Sex
The Spliceosome Machinery, 422
of Flies, 44 1RNA Splicing Is Carried Out by a Large Complex Called
KEY EXPERIMENTS Box 13-3 Identification of Docking Site andthe Spliceosome, 422
Selector Sequences, 442
Splicing Pathways, 424
MEDICAL CONNECTIONS Box 13-4 Defects in Pre-mRNA
Assembly, Rearrangements, and Catalysis within the
Splicing Cause Human Disease, 44 5
Spliceosome: The Splicing Pathway, 424
Exon Shuffling, 446Self-Splicing Introns Reveal That RNA Can Catalyze RNA
Exons Are Shuffled by Recombination to Produce GenesSplicing, 426
Encoding New Proteins, 44 6Group I Introns Release a Linear Intron Rather Than a
Lariat, 426
RNA Editing, 448
KEY EXPERIMENTS Box 13-2 Converting Group I Introns into
RNA Editing Is Another Way of Altering the Sequence of a n
Ribozymes, 428
mRNA, 44 8
How Does the Spliceosome Find the Splice Sites
Guide RNAs Direct the Insertion and Deletion of Uridines, 45 0
Reliably?, 430
MEDICAL CONNECTIONS Box 13-5 Deaminases and HIV4450
A Small Group of Introns Are Spliced by an Alternative
mRNA Transport, 452Spliceosome Composed of a Different Set of snRNPs,432
Once Processed, mRNA Is Packaged and Exported from th eNucleus into the Cytoplasm for Translation, 45 2
Alternative Splicing, 432
SUMMARY, 454Single Genes Can Produce Multiple Products by Alternative
Splicing 432
BIBLIOGRAPHY, 45 5
CHAPTER 14 • Translation, 45 7
Messenger RNA, 458
Transfer RNA, 46 1
Polypeptide Chains Are Specified by Open Reading
tRNAs Are Adaptors between Codons and AminoFrames, 458
Acids, 46 1Prokaryotic mRNAs Have a Ribosome-Binding Site That
ADVANCED CONCEPTS Box 14-1 CCA-Adding Enzymes :Recruits the Translational Machinery, 459
Synthesizing RNA without a Template, 462
Eukaryotic mRNAs Are Modified at Their 5 ' and 3 ' Ends to
tRNAs Share a Common Secondary Structure Tha tFacilitate Translation, 460
Resembles a Cloverleaf, 462
tRNAs Have an L-shaped Three-Dimensional
The Ribosome Uses Multiple Mechanisms to Select agains tStructure, 463
Incorrect Aminoacyl-tRNAs, 488
Attachment of Amino Acids to tRNA, 464
The Ribosome Is a Ribozyme, 49 1
tRNAs Are Charged by the Attachment of an Amino
Peptide Bond Formation and the Elongation Facto r
Acid to the 3'-Terminal Adenosine Nucleotide via
EF G Drive Translocation of the tRNAs and the
a High-Energy Acyl Linkage, 464
mRNA, 492
Aminoacyl tRNA Synthetases Charge tRNAs in Two
EF-G Drives Translocation by Displacing the tRNA Bound to
Steps, 464
the A Site, 494
Each Aminoacyl-tRNA Synthetase Attaches a Single Amino
EF-Tu-GDP and EF-G-GDP Must Exchange GDP fo r
Acid to One or More tRNAs, 466
GTP prior to Participating in a New Round ofElongation, 495
tRNA Synthetases Recognize Unique Structural Features of
A Cycle of Peptide Bond Formation ConsumesCognate tRNAs, 466Two Molecules of GTP and One Molecule o f
Aminoacyl-tRNA Formation Is Very Accurate, 468
ATP, 49 5Some Aminoacyl-tRNA Synthetases Use an Editin g
Pocket to Charge tRNAs with High Accuracy, 468
Termination of Translation, 496
The Ribosome Is Unable to Discriminate between
Release Factors Terminate Translation in Response to Stop
Correctly and Incorrectly Charged tRNAs, 469
Codons, 496
Short Regions of Class I Release Factors RecognizeThe Ribosome, 469
Stop Codons and Trigger Release of the PeptidylADVANCED CONCEPTS Box 14-2 Selenocysteine, 470
Chain, 496
The Ribosome Is Composed of a Large and a Small
11)V AN( I U r O y(I rr' hO 14-4 GTP-Binding Proteins,Subunit, 471
Conformational Switching, and the Fidelity an dThe Large and Small Subunits Undergo Association
Ordering of the Events of Translation, 498and Dissociation during Each Cycle of
GDP/GTP Exchange and GTP Hydrolysis Control theTranslation, 472
Function of the Class 11 Release Factor, 49 9New Amino Acids Are Attached to the Carboxyl
The Ribosome Recycling Factor Mimics a tRNA, 50 0Terminus of the Growing Polypeptide Chain, 474
MEDICAL CONNECTIONS Box 14-5 Antibiotics Arres tPeptide Bonds Are Formed by Transfer of the Growing
Cell Division by Blocking Specific Steps inPolypeptide Chain from One tRNA to Another, 474
Translation, 50 2
Ribosomal RNAs Are Both Structural and Catalytic
Regulation of Translation, 50 3Determinants of the Ribosome, 47 5
The Ribosome Has Three Binding Sites for tRNA, 475
Protein or RNA Binding Near the Ribosome-Bindin gSite Negatively Regulates Bacterial Translatio n
Channels through the Ribosome Allow the mRNA and
Initiation, 504Growing Polypeptide to Enter and/or Exit the
Regulation of Prokaryotic Translation : Ribosoma lRibosome, 476
Proteins Are Translational Repressors of Their Own
Initiation of Translation, 479
Synthesis, 505
Prokaryotic mRNAs Are Initially Recruited to the Small
Global Regulators of Eukaryotic Translation Target Key
Subunit by Base Pairing to rRNA, 480
Factors Required for mRNA Recognition and Initato r
A Specialized tRNA Charged with a Modified
tRNA Ribosome Binding 508
Methionine Binds Directly to the Prokaryotic
Spatial Control of Translation by mRNA-SpecificSmall Subunit, 480
4E-BPs, 51 0
Three Initiation Factors Direct the Assembly of an
An Iron-Regulated RNA-Binding Protein Control s
Initiation Complex That Contains mRNA and the
Translation of Ferritin, 51 1
Initiator tRNA, 481
Translation of thet Yeast Transcriptional Activator Gcn4 I s
Eukaryotic Ribosomes Are Recruited to the mRNA by the
Controlled by Short Upstream ORFs and Ternary
5' Cap, 482
Complex Abundance, 51 2
The Start Codon Is Found by Scanning Downstream from
Translation-Dependent Regulation of mRNA and Protei nthe 5' End of the mRNA, 483
Stability, 51 4ADVANCED CC?\CEPTS Box 14- uORFs and IRESs : Exceptions
The SsrA RNA Rescues Ribosomes That Translate Broke nThat Prove the Rule, 485
mRNAs, 51 4Translation Initiation Factors Hold Eukaryotic mRNAs in
Eukaryotic Cells Degrade mRNAs That Are Incomplete o rCircles, 487
Have Premature Stop Codons, 51 6
Translation Elongation, 487
SUMMARY, 51 8Aminoacyl-tRNAs Are Delivered to the A Site by Elongation
BIBLIOGRAPHY, 51 9Factor EF-Tu, 488
CHAPTER 15 • The Genetic Code, 52 1
The Code Is Degenerate, 521
Three Kinds of Point Mutations Alter the Genetic Code, 53 1
Perceiving Order in the Makeup of the Code, 522
Genetic Proof That the Code Is Read in Units of Three, 532
Wobble in the Anticodon, 523
Suppressor Mutations Can Reside in the Same or aThree Codons Direct Chain Termination, 525
Different Gene, 532How the Code Was Cracked, 525
Intergenic Suppression Involves Mutant tRNAs, 533Stimulation of Amino Acid Incorporation by Synthetic
Nonsense Suppressors Also Read Normal Terminatio nmRNAs, 526
Signals, 535Poly-U Codes for Polyphenylalanine, 527
Proving the Validity of the Genetic Code, 535Mixed Copolymers Allowed Additional Codon Assignments ,
527
The Code Is Nearly Universal, 536
Transfer RNA Binding to Defined Trinucleotide Codons, 528
SUMMARY, 538Codon Assignments from Repeating Copolymers, 529
BIBLIOGRAPHY, 538
Three Rules Govern the Genetic Code, 530
PART 4
REGULATION, 54 1
C H A P T E R 16 • Transcriptional Regulation in Prokaryotes, 54 7
Principles of Transcriptional Regulation, 547
Combinatorial Control: CAP Controls Other Genes As Well,
Gene Expression Is Controlled by Regulatory
560
Proteins, 547
KEY EXPERIMENTS Box 16-2 Jacob, Monod, and the Ideas
Most Activators and Repressors Act at the Level of
Behind Gene Regulation, 56 1
Transcription Initiation, 548
Alternative 6 Factors Direct RNA Polymerase to Alternative
Many Promoters Are Regulated by Activators That Help RNA
Sets of Promoters, 563
Polymerase Bind DNA and by Repressors That Block That
NtrC and MerR: Transcriptional Activators That Work by
Binding, 548
Allostery Rather than by Recruitment, 564
Some Activators and Repressors Work by Allostery and
NtrC Has ATPase Activity and Works from DNA Sites Fa r
Regulate Steps in Transcriptional Initiation after RNA
from the Gene, 564Polymerase Binding, 550
MerR Activates Transcription by Twisting Promoter
Action at a Distance and DNA Looping, 551
DNA, 565
Cooperative Binding and Allostery Have Many Roles in Gene
Some Repressors Hold RNA Polymerase at the Promoter
Regulation, 552
Rather than Excluding It, 566
Antitermination and Beyond: Not All of Gene Regulation
AraC and Control of the araBAD Operon by Antiactivation, 567
Targets Transcription Initiation, 552The Case of Bacteriophage X : Layers of Regulation, 568
Regulation of Transcription Initiation : Examples from
Alternative Patterns of Gene Expression Control Lytic an d
Prokaryotes, 553
Lysogenic Growth, 569
An Activator and a Repressor Together Control the lac
Regulatory Proteins and Their Binding Sites, 570
Genes, 553
k. Repressor Binds to Operator Sites Cooperatively, 57 1
CAP and Lac Repressor Have Opposing Effects on RNA
AIA,xv (~) iB » 16-S Concentration, Affinity, andPolymerase Binding to the lac Promoter, 554
Cooperative Binding, 572CAP Has Separate Activating and DNA-Binding
Repressor and Cro Bind in Different Patterns to Control LyticSurfaces, 555
and Lysogenic Growth, 573CAP and Lac Repressor Bind DNA Using a Common
Lysogenic Induction Requires Proteolytic Cleavage of AStructural Motif, 556
Repressor, 574KEY EXPERIMENTS Box 16-1 Activator Bypass Experiments, 557
Negative Autoregulation of Repressor Requires Long-
The Activities of Lac Repressor and CAP Are Controlled
Distance Interactions and a Large DNA Loop, 575
Allosterically by Their Signals, 559
Another Activator. A Cll. Controls the Decision between
Lytic and Lysogenic Growth upon Infection of a New
KEY EXPERIMENTS Box 16-5 Genetic Approaches That IdentifiedHost, 577
Genes Involved in the LyticlLysogenic Choice, 58 1
The Number of Phage Particles Infecting a Given Cell
Transcriptional Antitermination in Development, 58 2Affects Whether the Infection Proceeds Lytically or
Retroregulation : An Interplay of Controls on RNA SynthesisLysogenically, 578
and Stability Determines int Gene Expression, 584Growth Conditions of E . coil Control the Stability of CI
SUMMARY, 585Protein and thus the Lytic/Lysogenic Choice, 578
KEY EXPERIMENTS Box 16-4 Evolution of the X Switch, 579
BIBLIOGRAPHY, 586
CHAPTER 17 • Transcriptional Regulation in Eukaryotes, 589
Conserved Mechanisms of Transcriptional Regulation
KEY EXPERIMENTS Box 17-3 Evolvability of a Regulatoryfrom Yeast to Mammals, 591
Circuit, 612
Activators Have Separate DNA-Binding and Activating
Transcriptional Repressors, 61 3Functions, 59 1
Eukaryotic Regulators Use a Range of DNA-Binding
Signal Transduction and the Control of Transcriptiona l
Domains, but DNA Recognition Involves the Same
Regulators, 61 5
Principles as Found in Bacteria, 593
Signals Are Often Communicated to Transcriptiona l
TECHNIQUES Box 17-1 The Two-Hybrid Assay, 594
Regulators through Signal Transduction
Activating Regions Are Not Well-Defined Structures, 596
Pathways, 61 5
Signals Control the Activities of Eukaryotic Transcriptiona lRecruitment of Protein Complexes to Genes by
Regulators in a Variety of Ways, 61 7Eukaryotic Activators, 597
Activators and Repressors Sometimes Come in Pieces, 61 9Activators Recruit the Transcriptional Machinery to th e
Gene, 597
Gene "Silencing" by Modification of Histones an d
Activators Also Recruit Nucleosome Modifiers That Help the
DNA, 620
Transcriptional Machinery Bind at the Promoter or
Silencing in Yeast Is Mediated by Deacetylation an dInitiate Transcription, 598
Methylation of Histones, 62 1
Activators Recruit an Additional Factor Needed for Efficient
In Drosophila, HP1 Recognizes Methylated Histones an d
Initiation or Elongation at Some Promoters, 600
Condenses Chromatin, 622
Action at a Distance : Loops and Insulators, 601
ADC \u II)
1'1 5 Box 17-4 Is There a Histon e
Appropriate Regulation ofSome Groups ofGenes Requires
Code?, 62 3
Locus Control Regions, 603
DNA Methylation Is Associated with Silenced Genes in
KEY EXPERIMENTS Box 17-2 Long-Distance Interactions on the
Mammalian Cells, 62 4
Same and Different Chromosomes, 604
MEDKÜI CY)\\r( nr,\S Box 17-5 Transcriptional Repressionand Human Disease, 626
Signal Integration and Combinatorial Control, 605
Activators Work Synergistically to Integrate Signals, 605
Epigenetic Gene Regulation, 626
Signal Integration: The HO Gene Is Controlled by Two
Some States of Gene Expression Are Inherited through CellDivision Even When the Initiating Signal Is No LongerRegulators-One Recruits Nucleosome Modifiers and the
Present, 62 7Other Recruits Mediator, 60 7
Signal Integration: Cooperative Binding ofActivators at the
MEDIG41 CONNECTIONS Box 17-6 Using Transcriptio nFactors to Reprogram Somatic Cells into EmbryonicHuman 8-Interferon Gene, 608
Stem Cells, 62 9Combinatorial Control Lies at the Heart of the Complexity
and Diversity of Eukaryotes, 610
SUMMARY, 630
Combinatorial Control of the Mating-Type Genes from S .
BIBLIOGRAPHY, 63 1cerevisiae, 61 1
CHAPTER 18 • Regulatory RNAs, 63 3
Regulation by RNAs in Bacteria, 633
in Secondary Structure, 635
Riboswitches Reside within the Transcripts of Genes
r n cO c r i' - Box 18-1 Amino Acid BiosyntheticWhose Expression They Control through Changes
Operons Are Controlled by Attenuation, 639
RNA Interference Is a Major Regulatory Mechanism in
Small RNAs Can Transcriptionally Silence Genes by DirectingEukaryotes, 641
Chromatin Modification, 64 9
Short RNAs That Silence Genes Are Produced from a Variety
KEY EXPERIMENTS Box 18-2 History of miRNAs and RNA1, 65 0of Sources and Direct the Silencing of Genes in Thre eDifferent Ways, 641
The Evolution and Exploitation of RNAi, 652Did RNAi Evolve As an Immune System?, 652
Synthesis and Function of miRNA Molecules, 643
RNAi Has Become a Powerful Tool for Manipulating Gen emiRNAs Have a Characteristic Structure That Assists in
Expression, 65 4Identifying Them and Their Target Genes, 643
MEDICAL CONNECTIONS Box 18-3 RNAi and HumanAn Active miRNA Is Generated through a Two-Step
Disease, 65 6Nucleolytic Processing, 645
Dicer Is the Second RNA-Cleaving Enzyme Involved in
Regulatory RNAs and X inactivation, 65 7miRNA Production, 646
X-inactivation Creates Mosaic Individuals, 65 7
Incorporation of a Guide Strand RNA into RISC Makes the
Xist Is an RNA Regulator That Inactivates a Single XMature Complex That Is Ready to Silence Gene
Chromosome in Female Mammals, 65 7Expression, 647
SUMMARY, 65 9siRNAs Are Regulatory RNAs Generated from Long Double-
BIBLIOGRAPHY, 660Stranded RNAs, 64 9
CHAPTER 19 • Gene Regulation in Development and Evolution, 66 1
TECHNIQUES Box 19-1 Microarray Assays : Theory and
A Morphogen Gradient Controls Dorsoventral Patterning o fPractice, 662
the Drosophila Embryo, 679
Three Strategies by Which Cells Are Instructed to
Segmentation Is Initiated by Localized RNAs at the Anterio r
Express Specific Sets of Genes during Development, 663
and Posterior Poles of the Unfertilized Egg, 682
Some mRNAs Become Localized within Eggs and Embryos
Bicoid and Nanos Regulate hunchback 683
because of an Intrinsic Polarity in the Cytoskeleton, 663
KEY EXPERIMENTS Box 19-5 The Role of Activator Synergy i n
Cell-to-Cell Contact and Secreted Cell-Signaling Molecules
Development, 684
Both Elicit Changes in Gene Expression in Neighboring
MEDICAL CONNECTIONS Box 19 6 Stem Cells, 68 6
Cells, 664
The Gradient of Hunchback Repressor Establishes Different
Gradients of Secreted Signaling Molecules Can Instruct Cells
Limits of Gap Gene Expression, 68 7
to Follow Different Pathways of Development Based on
Hunchback and Gap Proteins Produce Segmentation Stripes
Their Location, 665
of Gene Expression, 688
Gap Repressor Gradients Produce Many Stripes of GeneExamples of the Three Strategies for Establishing
Expression, 68 9Differential Gene Expression, 666
KEY EXPERIMENTS Box 19-7 cis-Regulatory Sequences inThe Localized Ash l Repressor Controls Mating Type in Yeast
Animal Development and Evolution, 690by Silencing the HO Gene, 666
Short-Range Transcriptional Repressors Permit Differen tADVANCED CONCEPfS Box 19-2 Review of Cytoskeleton :
Enhancers to Work Independently of One Another withi nAsymmetry and Growth, 669
the Complex eve Regulatory Region, 692
A Localized mRNA Initiates Muscle Differentiation in the Sea
Homeotic Genes: An Important Class of DevelopmentalSquirt Embryo, 670
Regulators, 69 3\LNANCED CONCEPTS Box 19-3 Overview of Cion a
Development, 671
Changes in Homeotic Gene Expression Are Responsible for
Arthropod Diversity, 695Cell-to-Cell Contact Elicits Differential Gene Expression in
Arthropods Are Remarkably Diverse, 69 5the Sporulating Bacterium, Bacillus subtilis, 672
A Skin-Nerve Regulatory Switch Is Controlled by Notch
Changes in Ubx Expression Explain Modification of Limb s
Signaling in the Insect Central Nervous System, 673
among the Crustaceans, 695
A Gradient of the Sonic Hedgehog Morphogen Controls the
ADVANCED CONCEPTS Box 19-8 Homeotic Genes of Drosophil a
Formation of Different Neurons in the Vertebrate Neural
Are Organized in Special Chromosome Clusters, 696
Tube, 674
Why Insects Lack Abdominal Limbs, 69 8
The Molecular Biology of Drosophila Embryogenesis, 676
Modification of Flight Limbs Might Arise from the Evolutio nof Regulatory DNA Sequences, 699
An Overview of Drosophila Embryogenesis, 676
v>In c()N( LPTti Box 19-4 Overview of Drosophila
SUMMARY, 70 1
Development, 677
BIBLIOGRAPHY, 702
CHAPTER 20 • Genome Analysis and Systems Biology, 70 3
Genomics Overview, 703
Negative Autoregulation Dampens Noise and Allows a Rapid
Bioinformatics Tools Facilitate the Genome-wide
Response Time, 71 7
Identification of Protein-Coding Genes, 703
Gene Expression Is Noisy, 718
Whole-Genome Tiling Arrays Are Used to Visualize the
Positive Autoregulation Delays Gene Expression, 720
Transcriptome, 704
Some Regulatory Circuits Lock in Alternative Stabl e
Regulatory DNA Sequences Can Be Identified by Using
States, 720Specialized Alignment Tools, 706
Feed-Forward Loops Are Three-Node Networks with
The ChIP-Chip Assay Is the Best Method for Identifying
Beneficial Properties, 722Enhancers, 708
KEY EXPERIMENTS Box 20-2 Bistability and Hysteresis, 722TECHNIQUES Box 20-1 Bioinformatics Methods for the
Feed-Forward Loops Are Used in Development, 725Identification of Complex Enhancers, 708
Some Circuits Generate Oscillating Patterns of Gen eDiverse Animals Contain Remarkably Similar Sets of
Expression, 72 7Genes, 711
Synthetic Circuits Mimic Some of the Features of NaturalMany Animals Contain Anomalous Genes, 712
Regulatory Networks, 729
Synteny Is Evolutionarily Ancient, 713
Prospects, 730Deep Sequencing Is Being Used to Explore Human
SUMMARY, 73 0Origins, 715
BIBLIOGRAPHY, 73 1Systems Biology, 71 5
Transcription Circuits Consist of Nodes and Edges, 716
PART 5
METHODS, 73 3
CHAPTER 21 • Techniques of Molecular Biology, 73 9
Nucleic Acids, 740
Nested Sets of DNA Fragments Reveal Nucleotid e
Electrophoresis through a Gel Separates DNA and RNA
Sequences, 753
Molecules according to Size, 740
KEY EXPERIMENTS Box 21-2 Sequenators Are Used for High -
Restriction Endonucleases Cleave DNA Molecules at
Throughput Sequencing, 75 7
Particular Sites, 742
Shotgun Sequencing a Bacterial Genome, 757
DNA Hybridization Can Be Used to Identify Specific DNA
The Shotgun Strategy Permits a Partial Assembly of Larg e
Molecules, 743
Genome Sequences, 758
Hybridization Probes Can Identify Electrophoretically
The Paired-End Strategy Permits the Assembly of Large-
Separated DNAs and RNAs, 744
Genome Scaffolds, 760
Isolation of Specific Segments of DNA, 746
The $1000 Human Genome Is within Reach, 762
DNA Cloning, 746
Proteins, 764Cloning DNA in Plasmid Vectors, 746
Specific Proteins Can Be Purified from Cell Extracts, 764Vector DNA Can Be Introduced into Host Organisms by
Purification of a Protein Requires a Specific Assay, 764Transformation, 748
Preparation of Cell Extracts Containing Active Proteins, 765Libraries of DNA Molecules Can Be Created by
Proteins Can Be Separated from One Another Using ColumnCloning, 748
Chromatography, 765Hybridization Can Be Used to Identify a Specific Clone in a
Affinity Chromatography Can Facilitate More Rapid ProteinDNA Library, 749
Purification, 767Chemically Synthesized Oligonucleotides, 750
Separation of Proteins on Polyacrylamide Gels, 768The Polymerase Chain Reaction Amplifies DNAs by Repeated
Antibodies Are Used to Visualize Electrophoreticall yRounds of DNA Replication in Vitro, 751
Separated Proteins, 769TECHNIQUES Box 21-1 Forensics and the Polymerase Chain
Protein Molecules Can Be Directly Sequenced, 769Reaction, 753
Proteomics, 771
Nucleic Acid-Protein Interactions, 77 5
Combining Liquid Chromatography with Mass
The Electrophoretic Mobility of DNA Is Altered by ProteinSpectrometry Identifies Individual Proteins
Binding, 776within a Complex Extract, 771
DNA-Bound Protein Protects the DNA from Nucleases an dProteome Comparisons Identify Important Differences
Chemical Modification, 77 7beween Cells, 773
Chromatin Immunoprecipitation Can Detect Protei nMass Spectrometry Can Also Monitor Protein Modification
Association with DNA in the Cell, 778States, 773
In Vitro Selection Can Be Used to Identify a Protein's DNA -Protein-Protein Interactions Can Yield Information about
or RNA-Binding Site, 78 0Protein Function, 774
BIBLIOGRAPHY, 782
CHAPTER 22 • Model Organisms, 783
Bacteriophage, 784
Arabidopsis Is Easily Transformed for Reverse Genetics, 799
Assays of Phage Growth, 786
Arabidopsis Has a Small Genome That Is ReadilyThe Single-Step Growth Curve, 787
Manipulated, 800
Phage Crosses and Complementation Tests, 787
Epigenetics, 80 1
Transduction and Recombinant DNA, 788
Plants Respond to the Environment, 80 1Development and Pattern Formation, 80 2
Bacteria, 78 9Assays of Bacterial Growth, 789
The Nematode Worm, Caenorhabditis elegans, 80 2
Bacteria Exchange DNA by Sexual Conjugation, Phage-
C . elegans Has a Very Rapid Life Cycle, 803Mediated Transduction, and DNA-Mediated
C. elegans Is Composed of Relatively Few, Well-Studied CellTransformation, 790
Lineages, 804Bacterial Plasmids Can Be Used as Cloning Vectors, 791
The Cell Death Pathway Was Discovered in C . elegans, 805
Transposons Can Be Used to Generate Insertional Mutations
RNA1 Was Discovered in C . elegans, 805and Gene and Operon Fusions, 79 1
Studies on the Molecular Biology of Bacteria Have Been
The Fruit Fly, Drosophila melanogaster, 806
Enhanced by Recombinant DNA Technology, Whole-
Drosophila Has a Rapid Life Cycle, 80 6Genome Sequencing, and Transcriptional Profiling, 793
The First Genome Maps Were Produced in Drosophila, 807
Biochemical Analysis Is Especially Powerful in Simple Cells
Genetic Mosaics Permit the Analysis of Lethal Genes in Adultwith Well-Developed Tools of Traditional and Molecular
Flies, 809Genetics, 793
The Yeast FLP Recombinase Permits the Efficient Productio nBacteria Are Accessible to Cytological Analysis, 793
of Genetic Mosaics, 809Phage and Bacteria Told Us Most of the Fundamental Things
It Is Easy to Create Transgenic Fruit Flies that Carry Foreignabout the Gene, 794
DNA, 810
Baker 's Yeast, Saccharomyces cerevisiae, 795
The House Mouse, Mus musculus, 81 2The Existence of Haploid and Diploid Cells Facilitate Genetic
Mouse Embryonic Development Depends on StemAnalysis of S . cerevisiae, 795
Cells, 81 3Generating Precise Mutations in Yeast Is Easy, 796
It Is Easy to Introduce Foreign DNA into the Mous eS . cerevisiae Has a Small, Well-Characterized Genome, 796
Embryo, 81 3
S . cerevisiae Cells Change Shape as They Grow, 797
Homologous Recombination Permits the Selective Ablatio nof Individual Genes, 81 4
Arabidopsis, 798
Mice Exhibit Epigenetic Inheritance, 81 6Arabidopsis Has a Fast Life Cycle with Haploid and Diploid
BIBLIOGRAPHY, 81 8Phases, 79 8
Index, 819