LV (Lyu) Hexin ( 吕和鑫 ), Ph.D.

Associate Professor

Office: 2#-107

Wechat/Mobile:86-15822765529

Email: [email protected]

32 credit hours
Course arrangement
1st to 12th week

• Lesson 1: Introduction of Molecular Genetics

• Lesson 2: Transcription in Bacteria
• Lesson 3: Transcription in Eukaryotes
• Lesson 4: Post-Transcriptional Events
• Lesson 5: Translation
• Lesson 6: DNA Replication, Recombination, and T
ransposition, Genomes
Lesson 1: Genetics and Methods in
Molecular Biology

In 1865, Gregor Mendel published his findings on the inheritance of

seven different traits in the garden pea. Before Mendel ’s research,
Garden pea flowers. Flower color (purple
scientists thought inheritance occurred through a blending of each or white) was one of the traits Mendel
trait of the parents in the offspring. Mendel concluded instead that studied in his classic examination of
inheritance is particulate. That is, each parent contributes particles, or inheritance in the pea plant.
genetic units, to the offspring. We now call these particles genes.
Furthermore, by carefully counting the number of progeny plants
having a given phenotype, or observable characteristic (e.g., yellow
seeds, white flowers), Mendel was able to make some important
generalizations. The word phenotype, by the way, comes from the
same Greek root as phenomenon, meaning appearance. Thus, a tall
pea plant exhibits the tall phenotype, or appearance. Phenotype can
also refer to the whole set of observable characteristics of an
organism.

Gregor Mendel
Mendel ’s Laws of Inheritance
Mendel ’s Laws of Inheritance
• Mendel concluded that the allele for green seeds must have been pr
eserved in the F 1 generation, even though it did not affect the seed
color of those peas. His explanation was that each parent plant carri
ed two copies of the gene; that is, the parents were diploid, at least f
or the characteristics he was studying. According to this concept, h
omozygotes have two copies of the same allele, either two alleles f
or yellow seeds or two alleles for green seeds. Heterozygotes have
one copy of each allele. The two parents in the fi rst mating were h
omozygotes; the resulting F 1 peas were all heterozygotes. Further,
Mendel reasoned that sex cells contain only one copy of the gene; t
hat is, they are haploid. Homozygotes can therefore produce sex cel
ls, or gametes, that have only one allele, but heterozygotes can prod
uce gametes having either allele.
 Purple X white

P W

F1 PP WW
P P W W
F2 PP 1 PW 1 WW 1 WP 1

3:1
• Genes can exist in several differ
ent forms, or alleles. One allele
can be dominant over another, s
o heterozygotes having two diff
erent alleles of one gene will ge
nerally exhibit the characteristic
dictated by the dominant allele.
The recessive allele is not lost; i
t can still exert its infl uence wh
en paired with another recessive
allele in a homozygote
The Chromosome Theory
of Inheritance

• Other scientists either did not know about or uni

formly ignored the implications of Mendel ’s wo
rk until 1900 when three botanists, who had arri
ved at similar conclusions independently, redisc
overed it.
• After 1900, most geneticists accepted the particu
late nature of genes, and the field of genetics be Thomas Hunt Morgan
gan to blossom. One factor that made it easier fo
r geneticists to accept Mendel ’s ideas was a gro
wing understanding of the nature of chromosom
es, which had begun in the latter half of the nine
teenth century.
• Every gene has its place, or locus,
on a chromosome.

• Location of genes on chromosomes

. (a) A schematic diagram of a chro
mosome, indicating the positions of
three genes: A, B, and C. (b) A sche
matic diagram of a diploid pair of c
hromosomes, indicating the positio
ns of the three genes — A, B, and
C —on each, and the genotype (A o
r a; B or b; and C or c ) at each locu
s.
• Recombination in Drosophila. The two X chromosomes of
the female are shown schematically. One of them (red) carr
ies two wild-type genes: (m 1), which results in normal wi
ngs, and ( w 1), which gives red eyes. The other (blue) carr
ies two mutant genes: miniature ( m ) and white ( w ). Duri
ng egg formation, a recombination, or crossing over, indica
ted by the crossed lines, occurs between these two genes o
n the two chromosomes. The result is two recombinant chr
omosomes with mixtures of the two parental alleles. One is
m 1 w, the other is m w 1.
 Physical Evidence for
Recombination
• Barbara McClintock ( Figure 1.5 )
and Harriet Creighton provided a di
rect physical demonstration of reco
mbination in 1931. By examining
maize chromosomes microscopicall
y, they could detect recombinations
between two easily identifiable feat
ures of a particular chromosome (a
knob at one end and a long extensi
on at the other).

Photomicrographs (a and b) of chromosomes from Zea mays, or

maize. Diagrams of the chromosomes are shown in c and d. From
(Coe and Kass, 2005).

Barbara McClintock
 Molecular Genetics
• The studies just discussed tell us important things about th
e transmission of genes and even about how to map genes
on chromosomes, but they do not tell us what genes are m
ade of or how they work. This has been the province of mo
lecular genetics, which also happens to have its roots in M
endel ’s era. Friedrich Miescher

• The Discovery of DNA

In 1869, Friedrich Miescher discovered in the cell nucleus a mixtur
e of compounds that he called nuclein. The major component of nuclei
n is deoxyribonucleic acid (DNA). By the end of the nineteenth centur
y, chemists had learned the general structure of DNA and of a related c
ompound, ribonucleic acid (RNA). Both are long polymers — chains
of small compounds called nucleotides. Each nucleotide is composed o
f a sugar, a phosphate group, and a base. The chain is formed by linkin
g the sugars to one another through their phosphate groups.
The Composition of Genes

• By the time the chromosome theory of inheritance was gen

erally accepted, geneticists agreed that the chromosome
must be composed of a polymer of some kind. This would a
gree with its role as a string of genes. But which polym
er is it? Essentially, the choices were three: DNA, RNA,
and protein. Protein was the other major component of Mi
escher ’s nuclein; its chain is composed of links calle
d amino acids. The amino acids in protein are joined by
peptide bonds, so a single protein chain is called a pol
ypeptide
• Oswald Avery and his colleagues demonstrated in 1944 that DNA is th
e right choice . These investigators built on an experiment performed
earlier by Frederick Griffith in which he transferred a genetic trait fro
m one strain of bacteria to another.
 Griffith's experiment

Frederick Griffith laid the foundation for the identification of DNA as th

e genetic material in 1928 with his experiments on transformation in the
bacterium pneumococcus, now known as Streptococcus pneumoniae. Th
e wild-type organism is a spherical cell surrounded by a mucous coat call
ed a capsule. The cells form large, glistening colonies, characterized as s
mooth (S) . These cells are virulent, that is, capable of causing lethal infe
ctions upon injection into mice. A certain mutant strain of S. pneumoniae
has lost the ability to form a capsule. As a result, it grows as small, rough
(R) colonies. More importantly, it is avirulent; because it has no protecti
ve coat, it is engulfed by the host ’s white blood cells before it can prolif
erate enough to do any damage.
 DNA is genetic material

• Oswald Avery, Colin MacLeod, and Maclyn McCarty suppli

ed the missing piece in 1944. They used a transformation te
st similar to the one that Griffith had introduced, and they to
ok pains to define the chemical nature of the transforming s
ubstance from virulent cells. First, they removed the protein
from the extract with organic solvents and found that the ext Oswald Avery
ract still transformed. Next, they subjected it to digestion wi
th various enzymes. Trypsin and chymotrypsin, which destr
oy protein, had no effect on transformation. Neither did ribo
nuclease, which degrades RNA. These experiments ruled ou
t protein or RNA as the transforming material. On the other
hand, Avery and his coworkers found that the enzyme deox
yribonuclease (DNase), which breaks down DNA, destroye
d the transforming ability of the virulent cell extract. These r
esults suggested that the transforming substance was DNA.
• Direct physical-chemical analysis supported the hypothesis that the purifi ed transforming su
bstance was DNA. The analytical tools Avery and his colleagues used were the following:
• 1. Ultracentrifugation They spun the transforming substance in an ultracentrifuge (a very hig
h-speed centrifuge) to estimate its size. The material with transforming activity sedimented ra
pidly (moved rapidly toward the bottom of the centrifuge tube), suggesting a very high molec
ular weight, characteristic of DNA.
• 2. Electrophoresis They placed the transforming substance in an electric fi eld to see how rapi
dly it moved. The transforming activity had a relatively high mobility, also characteristic of D
NA because of its high charge-to-mass ratio.
• 3. Ultraviolet Absorption Spectrophotometry They placed a solution of the transforming subst
ance in a spectrophotometer to see what kind of ultraviolet (UV) light it absorbed most strong
ly. Its absorption spectrum matched that of DNA. That is, the light it absorbed most strongly
had a wavelength of about 260 nanometers (nm), in contrast to protein, which absorbs maxim
ally at 280 nm.
• 4. Elementary Chemical Analysis This yielded an average nitrogen-to-phosphorus ratio of 1.6
7, about what one would expect for DNA, which is rich in both elements, but vastly lower tha
n the value expected for protein, which is rich in nitrogen but poor in phosphorus. Even a slig
ht protein contamination would have raised the nitrogen-tophosphorusratio.
• These findings should have settled the issue of the nature of the gene,
but they had little immediate effect. The mistaken notion, from early c
hemical analyses, that DNA was a monotonous repeat of a four nucleo
tide sequence, such as ACTG-ACTG-ACTG, and so on, persuaded ma
ny geneticists that it could not be the genetic material. Furthermore, c
ontroversy persisted about possible protein contamination in the transf
orming material, whether transformation could be accomplished with
other genes besides those governing R and S, and even whether bacter
ial genes were like the genes of higher organisms.
• Yet, by 1953, when James Watson and Francis Crick published the
double-helical model of DNA structure, most geneticists agreed th
at genes were made of DNA. What had changed? For one thing, E
rwin Chargaff had shown in 1950 that the bases were not really fo
und in equal proportions in DNA, as previous evidence had sugges
ted, and that the base composition of DNA varied from one specie
s to another. In fact, this is exactly what one would expect for gene
s, which also vary from one species to another. Furthermore, Rolli
n Hotchkiss had refined and extended Avery ’s fi ndings. He purifi
ed the transforming substance to the point where it contained only
0.02% protein and showed that it could still change the genetic ch
aracteristics of bacterial cells. He went on to show that such highl
y purifi ed DNA could transfer genetic traits other than R and S.
• In 1952, A. D. Hershey and Martha Chase performed another experiment tha
t added to the weight of evidence that genes were made of DNA. This experi
ment involved a bacteriophage (bacterial virus) called T2 that infects the bac
terium Escherichia coli.
• During infection, the phage genes enter the host cell and direct the synthesis
of new phage particles. The phage is composed of protein and DNA only. Th
e question is this: Do the genes reside in the protein or in the DNA? The Her
shey –Chase experiment answered this question by showing that, on infectio
n, most of the DNA entered the bacterium, along with only a little protein. T
he bulk of the protein stayed on the outside. Because DNA was the major co
mponent that got into the host cells, it likely contained the genes. Of course,
this conclusion was not unequivocal; the small amount of protein that entere
d along with the DNA could conceivably have carried the genes. But taken t
ogether with the work that had gone before, this study helped convince genet
icists that DNA, and not protein, is the genetic material.
• The Hershey –Chase experiment depended on radioactive labels on th
e DNA and protein —a different label for each. The labels used were
phosphorus-32 ( 32 P) for DNA and sulfur-35 ( 35 S) for protein. Thes
e choices make sense, considering that DNA is rich in phosphorus but
phage protein has none, and that protein contains sulfur but DNA does
not. Hershey and Chase allowed the labeled phages to attach by their t
ails to bacteria and inject their genes into their hosts. Then they remov
ed the empty phage coats by mixing vigorously in a blender. Because
they knew that the genes must go into the cell, their question was: Wh
at went in, the 32 P-labeled DNA or the 35 S-labeled protein? As we h
ave seen, it was the DNA. In general, then, genes are made of DNA.
On the other hand, as we will see later in this chapter, other experimen
ts showed that some viral genes consist of RNA.
• The Hershey —Chase experiment. Phage T2 contains genes that allow it to replicate in E. coli. Because t
he phage is composed of DNA and protein only, its genes must be made of one of these substances. To dis
cover which, Hershey and Chase performed a two-part experiment. In the fi rst part (a), they labeled the p
hage protein with 35 S (red), leaving the DNA unlabeled (black). In the second part (b), they labeled the p
hage DNA with 32 P (red), leaving the protein unlabeled (black). Since the phage genes must enter the cel
l, the experimenters reasoned that the type of label found in the infected cells would indicate the nature of
the genes. Most of the labeled protein remained on the outside and was stripped off the cells by use of a b
lender (a), whereas most of the labeled DNA entered the infected cells (b). The conclusion was that the ge
nes of this phage are made of DNA.
The Relationship Between
Genes and Proteins
• How do genes work?
• In 1902. Archibald Garrod noticed that the human disease alcaptonuria se
emed to behave as a Mendelian recessive trait. It was likely, therefore, th
at the disease was caused by a defective, or mutant, gene. Moreover, the
main symptom of the disease was the accumulation of a black pigment in
the patient ’s urine, which Garrod believed derived from the abnormal bu
ildup of an intermediate compound in a biochemical pathway.
• By this time, biochemists had shown that all living things carry out count
less chemical reactions and that these reactions are accelerated, or catalyz
ed, by proteins called enzymes. Many of these reactions take place in seq
uence, so that one chemical product becomes the starting material, or sub
strate, for the next reaction. Such sequences of reactions are called path
ways, and the products or substrates within a pathway are called interme
diates.
• Garrod postulated that an intermediate accumulated to abnormally hig
h levels in alkaptonuria because the enzyme that would normally conv
ert this intermediate to the next was defective. Putting this idea togeth
er with the finding that alcaptonuria behaved genetically as a Mendeli
an recessive trait, Garrod suggested that a defective gene gives rise to
a defective enzyme. To put it another way: A gene is responsible for th
e production of an enzyme.
One-gene/one-enzyme
hypothesis
• By using Neurospora, scientists are not limited to the mutations that n
ature provides, but can use mutagens to introduce mutations into gen
es and then observe the effects of these mutations on biochemical path
ways.
• Beadle and Tatum found many instances where they could create Neu
rospora mutants and then pin the defect down to a single step in a bioc
hemical pathway, and therefore to a single enzyme (see Chapter 3). Th
ey did this by adding the intermediate that would normally be made b
y the defective enzyme and showing that it restored normal growth. B
y circumventing the blockade, they discovered where it was. In these
same cases, their genetic experiments showed that a single gene was i
nvolved. Therefore, a defective gene gives a defective (or absent) enz
yme. In other words, a gene seemed to be responsible for making one
enzyme. This was the one-gene/one-enzyme hypothesis.
• This hypothesis was actually not quite right for at least three reasons:
(1) An enzyme can be composed of more than one polypeptide chain
, whereas a gene has the information for making only one polypeptid
e chain. (2) Many genes contain the information for making polypept
ides that are not enzymes. (3) As we will see, the end products of so
me genes are not polypeptides, but RNAs. A modern restatement of t
he hypothesis would be: Most genes contain the information for mak
ing one polypeptide. This hypothesis is correct for prokaryotes and l
ower eukaryotes, but must be qualifi ed for higher eukaryotes, such a
s humans, where a gene can give rise to different polypeptides throu
gh an alternative splicing mechanism
Activities of Genes
How do genes work?

• First, they are replicated faithfully;

• Second, they direct the production of RNAs and proteins;
• Third, they accumulate mutations and so allow evolution.
The Three Domains of Life

• In the early part of the twentieth century, scientists divided all li

fe into two kingdoms: animal and plant. Bacteria were consider
ed plants, which is why we still refer to the bacteria in our guts
as intestinal “fl ora. ” But after the middle of the century, this cl
assifi cation system was abandoned in favor of a fi ve-kingdom
system that included bacteria, fungi, and protists, in addition to
plants and animals
• Then in the late 1970s, Carl Woese performed sequencing studies on
the ribosomal RNA genes of many different organisms and reached
a startling conclusion: A class of organisms that had been classifi ed
as bacteria have rRNA genes that are more similar to those of eukar
yotes than they are to those of classical bacteria like E. coli. Thus,
Woese named these organisms archaebacteria, to distinguish them fr
om true bacteria, or eubacteria. However, as more and more molecul
ar evidence accumulated, it became clear that the archaebacteria, de
spite a superfi cial resemblance, are not really bacteria. They represe
nt a distinct domain of life, so Woese changed their name to archaea
. Now we recognize three domains of life: bacteria, eukaryota, and a
rchaea. Like bacteria, archaea are prokaryotes— organisms without
nuclei—but their molecular biology is actually more like that of euk
aryotes than that of bacteria.
• The archaea live in the most inhospitable regions of the earth. Some
of them are thermophiles ( “heat-lovers ”) that live in seemingly unbe
arably hot zones at temperatures above 100 0C near deep-ocean geoth
ermal vents or in hot springs such as those in Yellowstone National P
ark. Others are halophiles (halogen-lovers) that can tolerate very high
salt concentrations that would dessicate and kill other forms of life. S
till others are methanogens ( “methane-producers ”) that inhabit envir
onments such as a cow ’s stomach, which explains why cows are suc
h a good source of methane.
The Chemical Nature
of Polynucleotides

The bases of DNA and RNA. The parent bases, purine a

nd pyrimidine, on the left, are not found in DNA and R
NA. They are shown for comparison with the other five
bases.
• The sugars of nucleic acids. Note the OH in the 2-p
osition of ribose and its absence in deoxyribose. • Two examples of nucleosides.
 Three nucleotides.
• The 5 9-nucleotides of deoxyadenosine are formed by phosphorylating the 5
9-hydroxyl group. The addition of one phosphate results in deoxyadenosine-5
9-monophosphate (dAMP). One more phosphate yields deoxyadenosine-5 9-di
phosphate (dADP). Three phosphates (designated a, b, g) give deoxyadenosin
e-5 9-triphosphate (dATP).
• A trinucleotide. This little piece of
DNA contains only three nucleoti
des linked together by phosphodie
ster bonds (red) between the 5’- a
nd 3’-hydroxyl groups of the suga
rs. The 5’-end of this DNA is at th
e top, where a free 5’-phosphate g
roup (blue) is located; the 3’-end i
s at the bottom, where a free 3’-hy
droxyl group (also blue) appears.
The sequence of this DNA could
be read as 5’pdTpdCpdA3’. This
would usually be simplified to TC
A.
• Shorthand DNA notation. (a) The nucleotide dATP. This illustration highlights four featu
res of this DNA building block: (1) The deoxyribose sugar is represented by the vertical
black line. (2) At the top, attached to the 1’-position of the sugar is the base,
• adenine (green). (3) In the middle, at the 3’-position of the sugar is a hydroxyl group (O
H, orange). (4) At the bottom, attached to the 5’-position of the sugar is a triphosphate gr
oup (purple). (b) A short DNA strand.
DNA Structure

Franklin ’s x-ray picture of DNA.

The regularity of this pattern
indicated that DNA is a helix. The
spacing between the bands at the top • The Double Helix
and bottom of the X gave the spacing
between elements of the helix (base
pairs) as 3.32 Å. The spacing
between neighboring bands in the
pattern gave the overall repeat of the
helix (the length of one helical turn)
as 33.2 Å.
• Watson and Crick published the outline of their model in t
he journal Nature, back-to-back with papers by Wilkins an
d Franklin and their coworkers showing the x-ray data. Th
e Watson –Crick paper is a classic of simplicity —only 90
0 words, barely over a page long. It was published very ra
pidly, less than a month after it was submitted. Actually, C
rick wanted to spell out the biological implications of the
model, but Watson was uncomfortable doing that. They co
mpromised on a sentence that is one of the greatest underst
atements in scientifi c literature: “It has not escaped our no
tice that the specifi c base pairing we have proposed imme
diately suggests a possible copying mechanism for the gen
etic material. ”
A Variety of DNA Structures
• Computer graphic models of A-, B-,
and Z-DNA. (a) A-DNA. Note the b
ase pairs (blue), whose tilt up from r
ight to left is especially apparent in t
he major grooves at the top and near
the bottom. Note also the right-hand
ed helix traced by the sugar –phosph
ate backbone (red). (b) B-DNA. Not
e the familiar right handed helix, wit
h roughly horizontal base pairs. (c)
Z-DNA. Note the left-handed helix.
All these DNAs are depicted with th
e same number of base pairs, empha
sizing the differences in compactne
ss of the three DNA forms.
• The structure for DNA proposed by Watson and Crick repr
esents the sodium salt of DNA in a fiber produced at very
high relative humidity (92%). This is called the B form of
DNA. Although it is probably close to the conformation of
most DNA in the cell, it is not the only conformation avail
able to double-stranded nucleic acids. If we reduce the rela
tive humidity surrounding the DNA fi ber to 75%, the sodi
um salt of DNA assumes the A form ( Figure 2.16a ). This
differs from the B form in several respects. Most obviously
, the plane of a base pair is no longer roughly perpendicula
r to the helical axis, but tilts 20 degrees away from horizon
tal.
• Both the A and B form DNA structures are righthanded: The heli
x turns clockwise away from you whether you look at it from the
top or the bottom. Alexander Rich and his colleagues discovered
in 1979 that DNA does not always have to be right-handed. They
showed that double stranded DNA containing strands of alternat
ing purines and pyrimidines (e.g., poly[dG-dC] poly[dG-dC]):
• —GCGCGCGC —
• —CGCGCGCG —
• can exist in an extended left-handed helical form. Because of the
zigzag look of this DNA ’s backbone when viewed from the side
, it is often called Z-DNA.
Separating the Two Strands
of a DNA Double Helix
• Although the ratios of G to C and A to T in an organism ’s DNA are fi
xed, the GC content (percentage of G 1 C) can vary considerably f
rom one DNA to another.

• The values range from 22 –73%, and these differences are refl ected i
n differences in the physical properties of DNA.
 DNA denaturation (DNA
melting)

• When a DNA solution is heated enough, the noncovalent forces that

hold the two strands together weaken and finally break. When this h
appens, the two strands come apart in a process known as DNA den
aturation, or DNA melting. The temperature at which the DNA stra
nds are half denatured is called the melting temperature, or Tm .
• The amount of strand separation, or melting, is measured by the absorbance of th
e DNA solution at 260 nm. Nucleic acids absorb light at this wavelength because
of the electronic structure in their bases, but when two strands of DNA come toge
ther, the close proximity of the bases in the two strands quenches some of this ab
sorbance. When the two strands separate, this quenching disappears and the abso
rbance rises 30 –40%. This is called the hyperchromic shift. The precipitous
rise in the curve shows that the strands hold fast until the temperature approaches
the Tm and then rapidly let go.

• Melting curve of Streptococcus pneumoniae DNA. The DNA was heated, and its melting
was measured by the increase in absorbance at 260 nm. The point at which the melting is
half complete is the melting temperature, or T m . The T m for this DNA under these
conditions is about 85 8C.
• The GC content of a DNA has a significan
t effect on its T m . In fact, as Figure 2.18 s
hows, the higher a DNA ’s GC content, the
higher its T m . Why should this be? Recal
l that one of the forces holding the two stra
nds of DNA together is hydrogen bonding.
Remember also that G –C pairs form three
hydrogen bonds, whereas A –T pairs have
only two. It stands to reason, then, that two
strands of DNA rich in G and C will hold t
o each other more tightly than those of AT-
rich DNA. Consider two pairs of embracin
g centipedes. One pair has 200 legs each, t • Relationship between DNA melting
he other 300. Naturally the latter pair will temperature and GC content. AT-DNA
be harder to separate. refers to synthetic DNAs composed
exclusively of A and T (GC content 5 0).
• Heating is not the only way to denature DNA. Organic solvents suc
h as dimethyl sulfoxide and formamide, or high pH, disrupt the hyd
rogen bonding between DNA strands and promote denaturation. Lo
wering the salt concentration of the DNA solution also aids denatur
ation by removing the ions that shield the negative charges on the t
wo strands from each other. At very low ionic strength, the mutuall
y repulsive forces of these negative charges are strong enough to de
nature the DNA at a relatively low temperature.
Reuniting the Separated
DNA Strands
• Once the two strands of DNA separate, they can, under the proper con
ditions, come back together again. This is called annealing or renatura
tion. Several factors contribute to renaturation efficiency. Here are thr
ee of the most important:

• 1. Temperature The best temperature for renaturation of a DNA is ab

out 25 8C below its T m . This temperature is low enough that it does
not promote denaturation, but high enough to allow rapid diffusion of
DNA molecules and to weaken the transient bonding between mismat
ched sequences and short intrastrand base-paired regions. This sugges
ts that rapid cooling following denaturation would prevent renaturatio
n. Indeed, a common procedure to ensure that denatured DNA stays d
enatured is to plunge the hot DNA solution into ice. This is called que
nching.
• 2. DNA Concentration The concentration of DNA in the solution is
also important. Within reasonable limits, the higher the concentration,
the more likely it is that two complementary strands will encounter ea
ch other within a given time. In other words, the higher the concentra
tion, the faster the annealing.
• 3. Renaturation Time Obviously, the longer the time allowed for ann
ealing, the more will occur.
DNAs of Various Sizes an
d Shapes
• The sizes are expressed three ways: molecular weight, number of base pairs, and l
ength.

• About 10.4 bp occur per helical turn, which is 33.2 Å long. To convert base pairs t
o molecular weight, we simply need to multiply by 660, which is the approximate
molecular weight of one average nucleotide pair.
DNA Content and the
C-Value Paradox
• C-values: DNA content per haploid cell.

• You would probably predict that complex organisms such as verteb

rates need more genes than simple organisms like yeast.
• In general, your prediction would be right; mouse and human haplo
id cells contain more than 100 times more DNA than yeast haploid
cells. Furthermore, yeast cells have about fi ve times more DNA th
an E. coli cells, which are even simpler. However, this corresponde
nce between an organism ’s physical complexity and the DNA cont
ent of its cells is not perfect. Consider, for example, the frog. Intuiti
vely, you would not suspect that an amphibian would have a higher
C-value than a human, yet the frog has seven times more DNA per
cell. Even more dramatic is the fact that the lily has 100 times more
DNA per cell than a human.
• This perplexing situation is called the C-value paradox. It becomes ev
en more difficult to explain when we look at organisms within a group
. For example, some amphibian species have C-values 100 times high
er than those of others, and the C-values of flowering plants vary even
more widely. Does this mean that one kind of higher plant has 100 tim
es more genes than another? That is simply unbelievable. It would rais
e questions about what all those extra genes are good for and why we
do not notice tremendous differences in physical complexity among th
ese organisms. The more plausible explanation of the C-value paradox
is that organisms with extraordinarily high C-values simply have a gre
at deal of extra, noncoding DNA. The function, if any, of this extra D
NA is still mysterious.
• In fact, even mammals have much more DNA than they nee
d for genes. Applying our simple rule (dividing the number
of base pairs by 1090) to the human genome yields an estim
ate of about 3 million for the maximum number of genes, w
hich is far too high. In fact, the finished version of the huma
n genome suggests that there are only about 20 –25,000 gen
es. This means that human cells contain more than 100 time
s more DNA than they apparently need. Much of this extra
DNA is found in intervening sequences within eukaryotic ge
nes. The rest is in noncoding regions outside of genes.
 REVIEW QUESTIONS
• 1. Compare and contrast the experimental approaches used by Avery and colleagues, a
nd by Hershey and Chase, to demonstrate that DNA is the genetic material.
• 2. Draw the general structure of a deoxynucleoside monophosphate. Show the sugar st
ructure in detail and indicate the positions of attachment of the base and the phosphate
. Also indicate the deoxy position.
• 3. Draw the structure of a phosphodiester bond linking two nucleotides. Show enough
of the two sugars that the sugar positions involved in the phosphodiester bond are clea
r.
• 4. Which DNA purine forms three H bonds with its partner in the other DNA strand?
Which forms two H bonds? Which DNA pyrimidine forms three H bonds with its part
ner? Which forms two H bonds?
• 5 The following drawings are the outlines of two DNA base pairs, with the bases ident
ified as a, b, c, and d. What are the real identities of these bases?
• 6. Draw a typical DNA melting curve. Label the axes and point out the melting tempe
rature.
• 7. Use a graph to illustrate the relationship between the GC content of a DNA and its
melting temperature. What is the explanation for this relationship?
• 8. Use a drawing to illustrate the principle of nucleic acid hybridization.