Chapter 16
The Molecular Basis of Inheritance
Note: Using the Interactive Study
Partner CD, review all activites in Chapter 16:
Molecular Basis of Inheritance. These activities
will be covered on the quiz and the test.
I. Introduction
A. 1940's
1. knew than chromosomes = DNA & protein
2. knew that chromosomes carried genes
3. BUT, What was a gene?
a. DNA was usually dismissed as a possibility
(1) physical & chemical properties of
DNA considered monotonous &
repetitive
b. protein seemed more likely to be gene
(1) proteins showed great heterogeneity
(2) proteins showed great specificity
B. case for DNA as the gene
1. 1928-Fred Griffith
a. worked on Streptococcus pneunomoniae
(1) causes pneumonia in mammels
b. grew colonies of Streptococcus pneunomoniae
(1) noted two types of colonies
(a) some appeared smooth (S)
(b) some appeared rough (R)
c. inject live bacteria from the two different
kinds of colonies into mice
(1) bacteria from S colony kills mouse
(2) bacteria from R colony doesn't kill mouse
d. inject dead bacteria into mice
(1) dead S do not kill
(2) dead R do not kill
e. mixed dead S with live R bact - waited -
then injected into mice
(1) kills mice
(2) look in blood
(a) found LIVE S bacteria!
f. Griffith concluded:
(1) "something" in the dead S bacteria transformed the
R bacteria and made them S bacteria
(2) see Fig 16.1 in Campbell et al.
2. 1944 - Avery, McCarty, MacLeod
a. showed that DNA isolated from S bacteria
transformed R form to S form
b. BUT: notion that DNA is genetic material
still not accepted!
3. 1952 - Hershey and Chase
a. showed that DNA is the genetic material of
viruses which attack bacteria
b. use bacteria phage = phage (T2)
c. T2 infects E. coli
(1) 1000's of copies of T2 made when one T2
infects E. coli
(2) E.coli cell bursts, 1000's of phage
released & cause another round infection
d. Hershey and Chase's Question:
Where are the genes which
program E. coli to make T2?
(1) T2's proteins?
(2) T2's DNA?
e. Answer with an experiment (Fig 16.2)
(1) grow T2 with E.coli in presence of S-35
(radioactive S)
(a) proteins contain S
(b) DNA doesn't contain S
(c) proteins get labeled with S-35 and
become radioactive
(2) second batch T2 grown in E.coli in presence
P-32 (radioactive P)
(a) proteins don't contain P
(b) DNA contains lots of P
(3) Experiment 1: T2 with S-35 labeled proteins
allowed to infect E.coli
(a) wait
(b) E.coli suspensions homogenized in
Waring Blendor to knock virus off of cells
(c) centrifuge to pellet cells
(d) look for 35S in pellet (pellet contained
intact bacteria) & in supernatant fluid
(supernatant fluid contained the T2)
(e) only found 35S in supernatant fluid
(no S-35 proteins found in E.coli)
(4) Experiment 2: T2 with P-32 labeled DNA
allowed to infect E.coli
(a) wait
(b) homogenize
(c) spin
(d) find lots of P-32 labeled DNA
inside of E.coli
(5) see the graphic again
(6) Hershey and Chase's conclusions:
(a) T2 injects DNA into E.coli
(b) DNA directs the synthesis of
new phage particles
(c) Finally, scientific community
accepts DNA as
the genetic material
II. DNA structure as a double helix
A. Watson and Crick see x-ray
crystallograph of DNA (Fig 16.4)
B. X-ray crystallograph suggests double helix
1. helix width uniform (2nm)
2. bases stacked 0.34 nm apart
C. Watson and Crick made models DNA molecule
D. proposed the model we use today
1. Fig 16.5 in Campbell et al.
2. see the interactive structure
III. DNA replication
A. Watson & Crick write in their first paper:
"It has not escaped our notice that the
specific pairing we have postulated
immediately suggests a possible copying
mechanism for the genetic material."
B. Semiconservative or conservative replication?
1. the scientific question is: How does DNA replicate?
2. three hypotheses were proposed
3. each hypothesis led to different predictions
4. BUT, how distinguish between old and new DNA?
-answer: use isotopes
-heavier isotopes => heavier DNA molecules
-lighter isotopes => lighter DNA molecules
4b. BUT, how distinguish between light and heavy DNA?
-answer: density of molecules vs
density of suspending solution
5. the Meselson & Stahl experiment
a. Meselson & Stahl experiments
(1) Meselson & Stahl predictions for first generation:
(a) IF replication is conservative, THEN
both strands of parental DNA are old
and both strands of new DNA are new
(b) IF replication is semiconservative, THEN
one strand of a double helix is old
(parental DNA) and other strand is
newly synthesized DNA
(c) IF replication is dispersive, THEN both
strands of DNA will always contain both
old and new DNA
(2) Meselson & Stahl predictions for second generation:
(a) IF replication is conservative, THEN
should see one DNA molecule with
old (parental) DNA and three DNA
molecules with new DNA
(b) IF replication is semiconservative, THEN
two DNA molecule wherein one
strand of a double helix is old
(parental DNA) and other strand is
newly synthesized DNA, AND two DNA
molecules which are all new DNA
(c) IF replication is dispersive, THEN both
all four DNA molecules will have strands
of DNA which contain both
old and new DNA
(3) How tell old strand from new?
(4) answer: use stable isotopes of N
(a) N-14 has 7 protons & 7 neutrons
(b) N-15 has 7 protons & 8 neutrons
(5) the experiment
(a) grow bacteria in N-15 containing
medium for many generations
(20 min/generation in E.coli)
(b) bacteria make DNA containing N-15 DNA
-every N in the bacteria (including
the DNA) is a N-15
-there is very little N-14 left anywhere
in the bacteria
(c) transfer bacteria to N-14 media for one
generation (20 min) or two generations
(40 minutes)
(d) break bacterial cells open
(e) isolate DNA molecules
(f) centrifuge DNA molecules in a
CsCl gradient
-CsCl forms a concentration gradient
when subjected to high speed centrifugation
which gives rise to a "density gradient"
(g) DNA moves down tube until it
"floats" at the concentration of CsCl
which has a density just equal to the
density of DNA
i) but, N-15 DNA is more dense than N-14 DNA
due to the extra neutron of each N atom
(h) results seen in Fig 16.9
-Note: results after 20 minutes (enough time for
the bacteria to replicate DNA once) eliminate the
conservative model (recall the prediction)
-Results after 40 minutes (enough time for
the bacteria to replicate twice) eliminate the
dispersive model
-We are left with the semiconservative model
-semiconservative model best predicts the
experimental outcomes
7. current understanding of DNA replication?
a. Fig 16.7 shows how DNA is replicated
b. two strands unzip
c. each parent strand serves as template for
production new complementary strand
d. get two identical DNA double helices
when DNA replication is complete
C. Some Molecular Details of Replication
1. replication begins at specific sites
on DNA molecule called "origins of replication"
a. origins of replication = "origins"
b. origins are specific sequence of bases
c. bacterial DNA has only one origin
-how many origins are found in
mitochondrial DNA?
-how many origins are found in
chloroplast DNA?
d. mammalian chromosomes have many origins
e. specific proteins recognize origins & bind DNA
-primase and DNA polymerase will find
these specific proteins and will bind to the
template DNA at the correct location
-so, some proteins find the origins and
guide the replication proteins to the right spot
2. replication requires strand separation
a. strand separation begins at origin
b. specific proteins prevent the two
separated DNA strands from coming
back together or knotting up
c. DNA replication requires a RNA primer
(1) primer synthesized by the enzyme primase
(2) primer is a short strand RNA (about 5 bases)
(3) RNA primer is complementary to DNA
d. new DNA synthesized by DNA polymerase chime
(1) DNA polymerase binds to a parent DNA strand
that has a complementary RNA primer
(2) DNA polymerase sequentially adds
deoxyribonucleotides to RNA primer
(a) deoxyribonucleotides added have bases
complementary to parent strand DNA
(3) see Fig 16.11
(4) new strand is DNA synthesis
(the DNA is becoming replicated)
e. DNA polymerase has "editting" function
(1) constantly "checks" to make sure no error
in DNA synthesis
f. new nucleotides added to 3'-OH group of
growing DNA strand by
dehydration synthesis
(1) recall structure of a nucleotide (ATP)
(2) recall structure of a deoxynucleotide
(3) see the 3'-OH
g. rate nucleotide additions
(1) bacteria add about 500 bases/second
(2) mammels add about 50 bases/second
IV. DNA repair
A. many proteins specifically "proofread" and "edit" DNA
B. various types of DNA errors can be found
1. errors due to physical damage
a. uv light
-not too hard to distinguish damaged bases
2. errors caused by faulty replication process
-DNA polymerase makes a mistake such
that the two bases are not complementary
-WHY IS THIS TYPE OF ERROR
A MAJOR PROBLEM?
3. the RNA left over from the primer is part of DNA strand
-easy to distinguish RNA from DNA
-RNA has ribose sugar, DNA has deoxyribose sugar
4. many others sources of errors
C. "incorrect" nucleotides are removed by specific enzymes
D. new deoxy-nucleotides-complementary to
"correct" DNA replace those that are removed by other enzymes
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