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Chapter 14: DNA: The Genetic Material
I. Introduction
A. Patterns of Heredity Explained by
B. Enhanced the Study of Humans as Biological Organisms
II. Where Do Cells Store Hereditary Information?
A. Hammerling's Experiments with Acetabularia
1. Initial experiment used a single genus as model organism
a. Large green alga cell with distinct foot, stalk and cap
b. Cap lacking nucleus amputated:
c. Foot with nucleus amputated:
d. Concluded hereditary information in foot
2. Second experiment used species that looked different
a. A. crenulata: disk-shaped cap, A. mediterranea: flower-shaped
b. A. crenulata stalk onto A. mediterranea foot
c. Regenerated cap looked similar to A. crenulata
d. Amputated regenerated cap, next cap looked like A. mediterranea
e. Further supported that hereditary information in foot
B. Frog Nucleus Transplant Experiments
1. Removed nucleus from frog egg:
2. Added nucleus from another egg:
3. Concluded nucleus directed development
C. Carrot Experiments
1. Mature carrot tissue fragmented
2. Individual cells developed roots, became adult plants
3. Concluded each cell has full set of genetic material, can generate entire
adult
III. Which Component of the Nucleus Contains the Hereditary Information?
A. Genes Hold Hereditary Information
B. The Griffith-Avery Experiments: Transforming Principle Is DNA
1. Mice injected with various strains of bacteria
a. Virulent, coated bacteria
b. Nonvirulent, coatless strain
c. Dead coated bacteria
2. Dead coated and live coatless bacteria mixed and injected
a. Mice
b.
passed from one strain to other
3. Transforming principle isolated, resembled DNA
a. Activity unaffected by protein-digesting enzymes
b. Activity lost in presence of
C. The Hershey-Chase Experiment: Some Viruses Direct Their Heredity with DNA
1. Bacteriophage viruses attack bacteria, possess either
a. Lytic virus injects viral genetic material into bacteria
b. Causes
2. Genetic material DNA or protein
a. Labeled T2 bacteriophage DNA with 32P and protein coat with
b. Viruses infect bacteria, attached viruses shaken off
c. Agitation removed 35S from bacterial preparation
d. Found 32P injected into bacterial cells
3. Concluded genetic material in bacteriophages was DNA
D. The Fraenkel-Conrat Experiment: Other Viruses Direct Their Heredity with
1. Some viruses possess
a. Tobacco mosaic virus (TMV)
b. Holmes ribgrass virus (HRV)
2. Genetic material
a. Tobacco infected with hybrid: TMV protein coat and HRV RNA
b. Observed lesions characteristic of HRV
3. Concluded hereditary material was
4. Other viruses also contain RNA, not DNA
a. Most copy own and insert into cell's
b. Retroviruses make intermediate double-stranded DNA
IV. The Chemical Nature of Nucleic Acids
A. Nucleic Acid First Isolated from Cell Nuclei
B. Composed of Nucleotides (P.A. Levine)
1. General structure
a.
b.
c.
1)
= adenine, guanine
2)
= thymine, cytosine
2. Numbering scheme for sugar structure
a. A prime ( ') indicates that the carbon is located on the sugar
b.
attaches to 5' carbon
c. Base attaches to carbon
d. OH attaches to carbon
C. Nucleotides Strung Together in Chains
1. Phosphate at 5 C, hydroxyl at 3 C allow chains to form
2. Sugars linked by
3. Nucleotide chain possesses definite
a. One end of chain with free
phosphate group
b. Other end of chain with free
hydroxyl group
c. Sequences conventionally written in
direction
D. Base Composition in Nucleotide Chains
1. Initially thought all four bases were in equal amounts
a. Assumed DNA a polymer of four repeating units
b. DNA had structural role and protein had hereditary role
2. Later found base amounts differed, depended on source
a. DNA not a simple repeating polymer
b. Chargaff's rules
1) Proportion of adenine (A) equal to
2) Proportion of guanine (G) equal to
3) Proportion of
(A + G) equal to
V. The Three-Dimensional Structure of DNA
A. Franklin's X-Ray Crystallography
1. Pattern of diffractions caused by DNA fibers
2. Not precise since DNA sample was in fibers not true crystals
3. Initial analysis of DNA
a. Spring-like spiral with helical diameter of 2 nanometers
b. Complete turn made every 3.4 nanometers
B. Watson-Crick Analysis
1. Constructed models to determine shape
2.
fit all known data
a. Bases pointed
toward one another
b. Large purine always paired with
c. Hydrogen bonds between bases stabilize antiparallel strands
d. Model explained Chargaff's results
1) Adenine, thymine form bonds
2) Guanine, cytosine form bonds
3.
backbone
a. joined by a
bond
4. Antiparallel strands
a. 3’ (OH) -------------- 5’ (PO4 )
5’ (PO4 ) ------------ 3’ (OH)
VI. How DNA Replicates
A. Model Dependent on Complementarity of Strands
1. Sequence of one chain determines
of its partner
2. Each chain is
of other
B. Replication Is
1. DNA replication model based on Meselson-Stahl experiments
a. Double strands
from one another
b. Separated strand serves as
for new strand
c. Each strand is copied to make new double helices
2. Labeled generations of bacteria with heavy nitrogen 15N
3. Transferred onto media containing lighter nitrogen 14N
a. Initial bacteria all heavy: two heavy strands
b. Later ones intermediate: one heavy, one light strand
c. Later grouped into intermediate and light classes
1) Intermediate group had one strand of each
2) Light group had two light strands
C. Two Strands of DNA Are Replicated in
Directions
1. Replication begins at
origins of replication
a. DNA duplex opened and untwisted by
enzyme
b. Forms replication bubbles where DNA strands are separated
2. Actual replication occurs at Y shaped ends of
a. Catalyzed by DNA
b. RNA primer constructs initial 10 sequence RNA
c.
recognizes primer and adds to it
d. RNA nucleotides replaced with
3. Replication occurs only in
direction
4. Strands are elongated by different mechanisms
5. Replication of leading strand,
strand
a. New strand grows from
end
b. Elongates towards
6. Lagging strand, strand replication
a. Elongates
from replication fork
b. Synthesized discontinuously in
batches
c. 5' 3' synthesis catalyzed by
d. Segments called
e.
attaches fragment to lagging strand
7. Overall replication process is termed
D. Stages of Replication in a Prokaryote
1. Initiation
a. at an
rich area within
b. primed with
replication
2. elongation
a. pol III adds complementary
b. only one primer is needed on the
strand
c. continuous priming on the
strand
3. termination
a. site roughly across from
b. intertwining is prevented
c. process not completely understood
E. Other Enzymatic Activities
1. DNA Primase – synthesizes the primer and continuously primes the
strand
a. creates a complementary short
2. Helicase – opens up the
a. uses energy from the hydrolysis of
3. DNA gyrase- reduces tortional strain – prevents
4. ssb – single strand binding protein – increases stability in an
environment
5. DNA pol I – removes primers and replaces them with
6. DNA ligase – joins
fragments “link em with ligase”
F. Comparing Prokaryotic and Eukaryotic DNA Replication
1. Bacterial DNA double helix in form of
a. Duplex nicked at single site
1)
on one side form one replication fork
2) Displace strand on two sides form
replication forks
b. Forks proceed around circle creating a daughter
c. When complete,
circles of DNA are present
2. Eukaryote DNA is not circular, but in
a. Each chromosome has replication forks
b. Each zone replicated as discrete replication unit
c. Zones average 100,000 base pairs in length
d. Advantage of this method is
1. number of origins that “fire” are adjusted during
e. Large amount of DNA requires sophisticated controls
f. uses DNA
VII. The Eukaryotic Chromosome
A. Nucleus Contains a Large Amount of DNA
1. Too fragile to stay extended at all times
2. Need efficient packaging to fit inside
B. Histones Package DNA into
1. Single DNA molecule wrapped around cluster of
2. Cluster binds to 146 nucleotide base pairs to form a
3. Resembled beads (nucleosomes) on a string (linker DNA)
4. H1 histone protein further condenses material into
C. Euchromatin and Heterochromatin
1. Both found in cell during
2. Heterochromatin is tightly packaged,
3. Less densely packaged euchromatin
D. The Chromosome
1. Further condensing occurs at beginning of
a. Probably assisted by H1 histones
b. Most transcriptionally inactive form of DNA
c. Packaging ensures surviving mitotic process
2. Chromosome has centromere and telomeres at ends of DNA
3. Full complement of chromosomes seen in karyotype
a. Stained chromosomes show
pattern
b.
identified in related species
E. How Many Genes Are on a Chromosome?
1. Example: Saccharomyces, brewer's yeast chromosome III
a. Identified 182 genes, half with no known function
b. Most genes are transcribed since 160 different mRNA's detected
2. Requires far more to identify gene functions than to map chromosome
VIII. Genes: The Units of Hereditary Information
A. Garrod Investigated Alkaptonuria, a Genetic Disorder
1. Abnormal urine turns black on exposure to air
a. Contains homogensic acid that oxidizes and blackens
b. Acid in normal urine broken down by
2. Postulated that affected patients lack
3. Concluded that information in DNA coded for
B. The One Gene-One Enzyme Hypothesis
1. Beadle and Tatum examined bread mold
2. Set out to create mutations in chromosomes
3. Creating genetic differences
a. Used X-rays to induce
in mold spores
b. Allowed progeny to grow on complete medium
1) Contains all possible nutrients
2) Strains unable to produce own nutrients still grew
4. Identifying mutant strains
a. Grow progeny on minimal medium to test for deficiencies
b. Cells unable to make
would not grow
c. Identified numerous growth-deficient mutants
5. Pinpointing biochemical deficiencies
a. Individually replace chemicals to determine deficiency
b. Determine enzymes involved in deficiencies
1) Arginine mutants clustered in three areas
2) Each site coded for different
in pathway
c. Postulated one gene-one enzyme (now polypeptide) hypothesis
C. How DNA Encodes Proteins
1. Sanger identified amino acid sequence of insulin
a. First demonstration of protein structure
b. Information for enzymes is ordered list of
2. Ingram analyzed normal and sickle-cell hemoglobin
a.
between hemoglobins
b. Alleles for genes altered in only
amino acid