Download Ch 16

Students
-Grades updated & posted….what mistakes have I made?
-Get graded Pocket Mouse worksheet from the counter
-Absent yesterday? Turn in sickle cell video packet
-Genetics problems DUE TOMORROW
-Tomorrow – report to Mr. Mercer’s room
- Rm 232
- Beside computer lab next to Media Center
-Phones into bins….off or muted….please & thank you!
Chapter 16: The Molecular Basis of Inheritance
Ch 14 – traits carried in gametes
Ch15 – genes found on chromosomes
Ch 16 – DNA is the molecule of inheritance
How did we learn this?
Chapter 16: The Molecular Basis of Inheritance
1. What evidence did Frederick Griffith have that DNA was the genetic material?
Transformation – external DNA is incorporated by a cell which
changes the cell’s genotype and phenotype
EXPERIMENT Bacteria of the “S” (smooth) strain of Streptococcus pneumoniae are pathogenic because they
have a capsule that protects them from an animal’s defense system. Bacteria of the “R” (rough) strain lack a
capsule and are nonpathogenic. Frederick Griffith injected mice with the two strains as shown below:
Living S
(control) cells
Living R
Heat-killed
Mixture of heat-killed S cells
(control) cells (control) S cells and living R cells
RESULTS
Mouse dies Mouse healthy Mouse healthy
Mouse dies
Living S cells
are found in
blood sample.
CONCLUSION Griffith concluded that the living R bacteria had been transformed into pathogenic S bacteria
by an unknown, heritable substance from the dead S cells.
Chapter 16: The Molecular Basis of Inheritance
1. What evidence did Frederick Griffith have that DNA was the genetic material?
2. How did Hershey & Chase determine that DNA was the genetic material?
- Bacteriophage – viruses that infect bacteria
Phage
head
Tail
Tail fiber
Bacterial
cell
100 nm
DNA
Chapter 16: The Molecular Basis of Inheritance
1. What evidence did Frederick Griffith have that DNA was the genetic material?
2. How did Hershey & Chase determine that DNA was the genetic material?
EXPERIMENT
In their famous 1952 experiment, Alfred Hershey and Martha Chase used radioactive sulfur
and phosphorus to trace the fates of the protein and DNA, respectively, of T2 phages that infected bacterial cells.
1
2 Agitated in a blender to 3
Mixed radioactively
separate phages outside
labeled phages with
the bacteria from the
bacteria. The phages
bacterial cells.
infected the bacterial cells.
Phage
Centrifuged the mixture 4 Measured the
so that bacteria formed
radioactivity in
a pellet at the bottom of
the pellet and
the test tube.
the liquid
Radioactivity
(phage protein)
in liquid
Radioactive Empty
protein
protein shell
Bacterial cell
Batch 1: Phages were
grown with radioactive
sulfur (35S), which was
incorporated into phage
protein (pink).
Batch 2: Phages were
grown with radioactive
phosphorus (32P), which
was incorporated into
phage DNA (blue).
DNA
Phage
DNA
Centrifuge
Radioactive
DNA
Pellet (bacterial
cells and contents)
Centrifuge
Pellet
Radioactivity
(phage DNA)
in pellet
Chapter 16: The Molecular Basis of Inheritance
1. What evidence did Frederick Griffith have that DNA was the genetic material?
2. How did Hershey & Chase determine that DNA was the genetic material?
RESULTS
Phage proteins remained outside the bacterial cells during infection, while
phage DNA entered the cells. When cultured, bacterial cells with radioactive phage DNA
released new phages with some radioactive phosphorus.
CONCLUSION
Hershey and Chase concluded that DNA, not protein, functions as the
T2 phage’s genetic material.
Chapter 16: The Molecular Basis of Inheritance
1. What evidence did Frederick Griffith have that DNA was the genetic material?
2. How did Hershey & Chase determine that DNA was the genetic material?
Nitrogenous
Sugar-phosphate
3. What was Chargaff’s contribution?
bases
backbone
- Nucleotide composition
5 end
CH
O
5
- Human
O
H
O P O CH
O 1
4 H
N
O
N
- A – 30.3%
H
H
H
H
O
2
3
- T – 30.3%
H
Thymine (T)
- C – 19.9%
O
H
H
O P O CH
N
- G – 19.5%
O
N
–
3
2
–
2
O–
Chargaff’s rules
A=T
C=G
H
H
H
Adenine (A)
H
H
O
P O CH2
O–
Note:
H
H
O
H
H
N H
N
H
H
5’ end – phosphate group
3’ end - -OH
N
N
H
O
H
N
H
H
N
O
Cytosine (C)
O
5
H
O P O CH2
N
O
O
1
–
4 H
O
N
H
Phosphate H
H
N H
2
3
N
H
OH
N H
Sugar (deoxyribose)
H
3 end
Guanine (G)
DNA nucleotide
Chapter 16: The Molecular Basis of Inheritance
1.
2.
3.
4.
What evidence did Frederick Griffith have that DNA was the genetic material?
How did Hershey & Chase determine that DNA was the genetic material?
What was Chargaff’s contribution?
What experiment did Watson & Crick do?
- NONE – compiled everyone else’s data
(a) Rosalind Franklin
(b) Franklin’s X-ray diffraction
Photograph of DNA Double
helix
Chapter 16: The Molecular Basis of Inheritance
1.
2.
3.
4.
What evidence did Frederick Griffith have that DNA was the genetic material?
How did Hershey & Chase determine that DNA was the genetic material?
What was Chargaff’s contribution?
What experiment did Watson & Crick do?
- NONE – compiled everyone else’s data
- Determined double helix from Rosalind Franklin’s data
Purine + Purine: too wide
Pyrimidine + pyrimidine: too narrow
Purine + pyrimidine: width
Consistent with X-ray data
Chapter 16: The Molecular Basis of Inheritance
1.
2.
3.
4.
What evidence did Frederick Griffith have that DNA was the genetic material?
How did Hershey & Chase determine that DNA was the genetic material?
What was Chargaff’s contribution?
What experiment did Watson & Crick do?
- NONE – compiled everyone else’s data
- Determined double helix from Rosalind Franklin’s data
N
N
N
N
Sugar
O
H
H
CH3
N
N
N
O
Sugar
Thymine (T)
Adenine (A)
H
O
N
N
Sugar
N
H
H
N
N
N
N
N
H
Guanine (G)
H
O
Sugar
Cytosine (C)
Figure 16.7 The double helix
G
5 end
C
O
A
T
–O
T
OH
P
Hydrogen bond
3 end
OH
O
A
O
H2C
O
C
P
3.4 nm
G
–O
O
G
O
P
–O
A
A
–O
A
T
O
CH2
O
O
O
T
O
C
O
O–
P
O
O
H2C
T
O–
P
O
O
T
G
CH2
O
H2C
A
O
O
C
G
C
A
T
1 nm
O
G
C
O
O
P
CH2
O
O
P
O
O
H2C
O–
A
O
T
T
A
O
OH
CH2
O
3 end
G
O
C
O–
P
O
0.34 nm
A
T
(a) Key features of DNA structure
5 end
(b) Partial chemical structure
(c) Space-filling model
DNA is antiparallel
Hydrogen bonds hold double helix together
Now that we know how DNA is shaped & how it fits together…..
Chapter 16: The Molecular Basis of Inheritance
1.
2.
3.
4.
5.
What evidence did Frederick Griffith have that DNA was the genetic material?
How did Hershey & Chase determine that DNA was the genetic material?
What was Chargaff’s contribution?
What experiment did Watson & Crick do?
How does DNA replicate itself?
The basic concept
T
A
T
A
T
A
C
G
C
G
C
T
A
T
A
T
A
A
T
A
T
A
T
G
C
G
C
G
C
G
A
T
A
T
A
T
C
G
C
G
C
G
T
A
T
A
T
A
T
A
T
A
T
C
G
C
G
C
A
G
(a) The parent molecule has two
complementary strands of DNA.
Each base is paired by hydrogen
bonding with its specific partner,
A with T and G with C.
(b) The first step in replication is
separation of the two DNA
strands.
(c) Each parental strand now
serves as a template that
determines the order of
nucleotides along a new,
complementary strand.
(d) The nucleotides are connected
to form the sugar-phosphate
backbones of the new strands.
Each “daughter” DNA
molecule consists of one parental
strand and one new strand.
But is replication done in a conservative, semi-conservative or dispersive manner?
Students
- Place Genetics problems in box
-include “scratch work” if you only provided the answers
- Lab notebooks due Monday
- Learning logs due Tuesday (MC & math test)
- Review session Tuesday 7 AM
- Phones in bin…muted or off…please & thank you
Figure 16.10 Three alternative models of DNA replication
Parent cell
Conservative
- Original strands
return together
(a) Conservative
model. The two
parental strands
reassociate
after acting as
templates for
new strands,
thus restoring
the parental
double helix.
Semi-conservative
-One original &
one new strand
(b) Semiconservative model.
The two strands
of the parental
molecule
separate,
and each functions
as a template
for synthesis of
a new, complementary strand.
Dispersive
-Original strands
are dispersed
through new
strands
(c) Dispersive
model. Each
strand of both
daughter molecules contains
a mixture of
old and newly
synthesized
DNA.
First
replication
Second
replication
The Meselson – Stahl Experiment showed how DNA replicates.
Figure 16.11 Does DNA replication follow the conservative, semiconservative,
or dispersive model?
EXPERIMENT Matthew Meselson and Franklin Stahl cultured E. coli bacteria for several generations
on a medium containing nucleotide precursors labeled with a heavy isotope of nitrogen, 15N. The bacteria
incorporated the heavy nitrogen into their DNA. The scientists then transferred the bacteria to a medium with
only 14N, the lighter, more common isotope of nitrogen. Any new DNA that the bacteria synthesized would be
lighter than the parental DNA made in the 15N medium. Meselson and Stahl could distinguish DNA of different
densities by centrifuging DNA extracted from the bacteria.
1 Bacteria
2 Bacteria
cultured in
medium
containing
15N
transferred to
medium
containing
14N
RESULTS
3 DNA sample
centrifuged
after 20 min
(after first
replication)
4 DNA sample
centrifuged
after 40 min
(after second
replication)
Less
dense
More
dense
The bands in these two centrifuge tubes represent the results of centrifuging two DNA samples from the flask
in step 2, one sample taken after 20 minutes and one after 40 minutes.
CONCLUSION Meselson and Stahl concluded that DNA replication follows the semiconservative
model by comparing their result to the results predicted by each of the three models in Figure 16.10.
The first replication in the 14N medium produced a band of hybrid (15N–14N) DNA. This result eliminated
the conservative model. A second replication produced both light and hybrid DNA, a result that eliminated
the dispersive model and supported the semiconservative model.
First replication
Conservative
model
Semiconservative
model
Dispersive
model
Second replication
Chapter 16: The Molecular Basis of Inheritance
1.
2.
3.
4.
5.
6.
What evidence did Frederick Griffith have that DNA was the genetic material?
How did Hershey & Chase determine that DNA was the genetic material?
What was Chargaff’s contribution?
What experiment did Watson & Crick do?
How does DNA replicate itself?
How is DNA replicated?
- Origin of replication - sequence of DNA
Chapter 16: The Molecular Basis of Inheritance
1.
2.
3.
4.
5.
6.
What evidence did Frederick Griffith have that DNA was the genetic material?
How did Hershey & Chase determine that DNA was the genetic material?
What was Chargaff’s contribution?
What experiment did Watson & Crick do?
How does DNA replicate itself?
How is DNA replicated?
- Origin of replication – sequence of DNA
- Elongation
- DNA polymerase
- 500 bp/sec bacteria, 50 bp/sec in us
- New base added to 3’end (-OH)…..5’  3’ direction
- dNTPs provide energy
Figure 16.13 Incorporation of a nucleotide into a DNA strand
New strand
Template strand
Sugar
A
Base
Phosphate
3’ end
5’ end
3’ end
5’ end
T
A
T
C
G
C
G
G
C
G
C
T
A
OH
DNA polymerase
A
P
OH
P
Pyrophosphate 3’ end
C
C
OH
Nucleoside
triphosphate
2 P
5’ end
5’ end
Chapter 16: The Molecular Basis of Inheritance
1.
2.
3.
4.
5.
6.
What evidence did Frederick Griffith have that DNA was the genetic material?
How did Hershey & Chase determine that DNA was the genetic material?
What was Chargaff’s contribution?
What experiment did Watson & Crick do?
How does DNA replicate itself?
How is DNA replicated?
- Origin of replication – sequence of DNA
- Elongation
- Dealing with antiparallel strands
Figure 16.14 Synthesis of leading and lagging strands during DNA replication
1 DNA pol Ill elongates
DNA strands only in the
5
3 direction.
3
5
Parental DNA
2 One new strand, the leading strand,
can elongate continuously 5
3
as the replication fork progresses.
5
3
Okazaki
fragments
2
1
3
5
3 The other new strand, the
lagging strand must grow in an overall
3
5 direction by addition of short
segments, Okazaki fragments, that grow
5
3 (numbered here in the order
they were made).
DNA pol III
Template
strand
3
Leading strand
Lagging strand
2
Template
strand
1
DNA ligase
Overall direction of replication
4 DNA ligase joins Okazaki
fragments by forming a bond between
their free ends. This results in a
continuous strand.
Figure 16.15 Synthesis of the lagging strand
1
Primase joins RNA nucleotides
into a primer.
3
5
5
3
Template
strand
2
RNA primer
3
5
3
DNA pol III adds DNA nucleotides to the
primer, forming an Okazaki fragment.
5
3
1
After reaching the next
RNA primer (not shown),
DNA pol III falls off.
Okazaki
fragment
3
3
5
1
5
4
After the second fragment is
primed. DNA pol III adds DNA
nucleotides until it reaches the
first primer and falls off. 5
3
5
3
2
5
1
DNA pol 1 replaces the
RNA with DNA, adding to
the 3 end of fragment 2.
5
3
6
3
2
DNA ligase forms a bond
between the newest DNA
and the adjacent DNA of
fragment 1.
5
3
2
5
1
7
The lagging strand
in this region is now
complete.
3
1
Overall direction of replication
5
Figure 16.16 A summary of bacterial DNA replication
Overall direction of replication
1 Helicase unwinds the
parental double helix.
2 Molecules of singlestrand binding protein
stabilize the unwound
template strands.
3 The leading strand is
synthesized continuously in the
5 3 direction by DNA pol III.
Leading
strand
Origin of replication
Lagging
strand
Lagging
strand
Leading
strand
OVERVIEW
DNA pol III
Leading
strand
5
3
Parental DNA
Replication fork
Primase
DNA pol III
Primer
4 Primase begins synthesis
of RNA primer for fifth
Okazaki fragment.
5
DNA pol III is completing synthesis of
the fourth fragment, when it reaches the
RNA primer on the third fragment, it will
dissociate, move to the replication fork,
and add DNA nucleotides to the 3 end
of the fifth fragment primer.
4
6
DNA ligase
DNA pol I
Lagging
strand
2
1
3
DNA pol I removes the primer from the 5 end
of the second fragment, replacing it with DNA
nucleotides that it adds one by one to the 3’ end
of the third fragment. The replacement of the
last RNA nucleotide with DNA leaves the sugarphosphate backbone with a free 3 end.
3
5
7
DNA ligase bonds
the 3 end of the
second fragment to
the 5 end of the first
fragment.
Chapter 16: The Molecular Basis of Inheritance
1.
2.
3.
4.
5.
6.
7.
What evidence did Frederick Griffith have that DNA was the genetic material?
How did Hershey & Chase determine that DNA was the genetic material?
What was Chargaff’s contribution?
What experiment did Watson & Crick do?
How does DNA replicate itself?
How is DNA replicated?
How is DNA repaired?
- Proofreading – mismatch repair – DNA polymerase fixes mistakes made
during copying
- Excision repair
Figure 16.17 Nucleotide excision repair of DNA damage
1 A thymine dimer
distorts the DNA molecule.
2 A nuclease enzyme cuts
the damaged DNA strand
at two points and the
damaged section is
removed.
Nuclease
3 Repair synthesis by
a DNA polymerase
fills in the missing
nucleotides.
DNA
polymerase
DNA
ligase
4 DNA ligase seals the
Free end of the new DNA
To the old DNA, making the
strand complete.
Thymine dimers
from UV damage
Chapter 16: The Molecular Basis of Inheritance
1.
2.
3.
4.
5.
6.
7.
8.
What evidence did Frederick Griffith have that DNA was the genetic material?
How did Hershey & Chase determine that DNA was the genetic material?
What was Chargaff’s contribution?
What experiment did Watson & Crick do?
How does DNA replicate itself?
How is DNA replicated?
How is DNA repaired?
What happens at the end of chromosomes (telomere)?
- Recall DNA polymerase works 5’ → 3’ direction
- After RNA primer is removed, chromosome is shortened
Figure 16.18 Shortening of the ends of linear DNA molecules
5
End of parental
DNA strands
Leading strand
Lagging strand
3
Last fragment
Previous fragment
RNA primer
Lagging strand
5
3
Primer removed but
cannot be replaced
with DNA because
no 3 end available
for DNA polymerase
Removal of primers and
replacement with DNA
where a 3 end is available
5
3
Second round
of replication
5
New leading strand 3
New lagging strand 5
3
Further rounds
of replication
Shorter and shorter
daughter molecules
Telomerase
-active in germ line cells & newborns
to prevent extensive shortening
-extends telomere a bit
-has its own RNA
- complementary to telomere
- serves as primer
primer
2009 Nobel Prize in Medicine - for the discovery of how chromosomes are protected
by telomeres and the enzyme telomerase