Chapters 16-21:

Molecular Genetics

I. Structure/Function of DNA and

RNA

Genetic information is stored in and passed to subsequent

generations through DNA and in some cases, RNA.
In eukaryotes, multiple, linear chromosomes = thread-like
structures found in the nucleus and made-up of one, long
DNA molecule, contain the information that is passed from
parent to offspring genes.
Prokaryotes and viruses can contain plasmids = small
extra-chromosomal double-stranded DNA molecules, in
addition to circular chromosomes.
DNA CODES FOR PROTEINS/ENZYMES WHICH
REGULATE AND DIRECT METABOLIC FUNCTIONS
FOR CELLS.
DNA and RNA are both POLYMERS OF NUCLEOTIDES.

Nucleotides are monomers made-up of three parts:

1. Sugar
2. Phosphate
3. Nitrogen base/nitrogenous base
The nitrogen base is the variable part
of each nucleotide.
There are five nitrogen bases:
1.
Adenine (A) DNA/RNA
Purines
2.
Guanine (G) DNA/RNA
3.
Cytosine (C) - DNA/RNA
4.
Uracil (U) RNA only
Pyrimidines
5.
Thymine (T) DNA only
Purines = double carbon-ring nitrogen bases
Pyrimidines = single carbon-ring nitrogen bases

Nitrogenous Bases:

The DNA double-helix structure is defined by

complementary base pairing:
1. adenine (A) always pairs with thymine (T)
2. cytosine (C) always pairs with guanine (G).
Complementary base pairs bond via weak hydrogen
bonds allows for the bonds to be broken easily for
replication and transcription.
1. 2H bonds between A-T.
2. 3H bonds between C-G.

 Two strands of the DNA double helix are antiparallel =

oriented in the opposite directions ONE STRAND

RUNS 5 TO 3, THE OTHER RUNS 3 TO 5.
Phosphates attach to the 5 and 3 carbons of the
molecules backbone.
The end of a strand refers to the free carbon the end
where no further phosphates attach.

# of types in
an organism

DNA

RNA

4:
mRNA
(messenger),
tRNA
(transfer),
rRNA
(ribosomal),
RNAi
(interference)

Sugar

deoxyribose

Nitrogen
bases

A, T, C, G

Functions

Contains the heredity

antiparallel,
information/instruction double-helix with
s for making proteins
complementary
(genes) of the cell.
base pairing
mRNA: copies the DNA
code (instructions for
making proteins) in the
nucleus during
transcription and
carries said information
to the ribosome.

ribose

A, U, C, G

Structure

tRNA: carries the

correct amino acids to
the ribosome during
translation.

single-stranded
and linear

single-stranded
and clover-leaf

rRNA: combines with

proteins to make up the
ribosomes.

single-stranded
and globular

RNAi: regulate gene

expression at the level
of mRNA transcription.

double- stranded

II. DNA Replication

During the S phase of the cell cycle, DNA is

replicated (a 2nd chromatid is made and attached via
the centromere) to ensure continuity of hereditary
information = DNA replication.
Both strands of the DNA molecule will serve as a
template for a new strand DNA must be unzipped
at the weak hydrogen bonds located between
nitrogen bases done by the enzyme HELICASE.
Semiconservative replication results in two
double-stranded molecules of DNA each consisting of
one old strand of DNA (template) and one new
strand (complementary strand).

Replication occurs in two directions resulting in a leading

strand and a lagging strand.

The steps of DNA replication are as follows:

1. Helicase unzips the DNA double helix forming a Y-shaped
replication fork.
2. Single-stranded binding proteins attach to each strand of
the uncoiled helix to keep them separated.

3.

3.

As helicase moves down the DNA unzipping it, the

double helix still in front of the enzyme is forced to twist
up and forms knots topoisomerase breaks and
rejoins the double helix in front of helicase to prevent
these knots.

The unzipped DNA has two templates: 3 5 and 5

3. On BOTH templates RNA primase adds an RNA
primer to start the new strand.

5.

Once the RNA primer is in place (short sequence of RNA

nucleotides), the enzyme DNA polymerase begins adding
complementary DNA nucleotides down the remaining length
of the molecule it can only move in the 3 5 direction
(3 to 5, youre alive!)
*(NOTE: DNA polymerase can ONLY move DOWN THE
TEMPLATE in the 3 5 direction and can ONLY build off of
an already existing nucleotide why a primer is needed)*

6.

7.

8.

9.

For the 3 5 template strand, DNA polymerase

can add complementary nucleotides without
interruption as it follows directly behind helicase =
LEADING STRAND.
For the 5 3 template strand, however, DNA
polymerase MUST add complementary
nucleotides in fragments as it goes in the opposite
direction of helicase/replication fork = LAGGING
STRAND. The fragments are called OKAZAKI
FRAGMENTS.
Once an Okazaki fragment is formed, DNA
polymerase replaces the RNA nucleotides of the
RNA primer with DNA nucleotides.
The enzyme DNA ligase connects each Okazaki
fragment together producing a single
complementary strand.

DNA Replication:

Because DNA polymerase can only add nucleotides off

of the template in the 3 5 direction, when the RNA
primer is removed from the end of the leading and
lagging strands, the gaps cannot be filled.
With each successive round of DNA replication, the length of
DNA shortens.
To prevent the deletion of genes with subsequent
replications, the ends of a length of DNA contain multiple
repetitions of one short nucleotide sequence (TTAGGG) =
TELOMERES (now believed to be the basis of aging).
The number of repetitions in a telomere ranges from 100 to
1,000.
Telomerase = an enzyme that catalyzes the lengthening of
telomeres absent from most cells of multicellular
organisms including humans.
Telomeres are believed to be the limiting factor in the life
span of tissues and organisms is telomerase the fountain
of youth?

Telomere
s

III. Protein Synthesis

The DNA of chromosomes contains the information that

determines which proteins, and how many of each, a cell will
produce.
DNA codes for proteins (polypeptides) MOST of which are
enzymes that regulate chemical reactions influences the
final characteristics of a cell.
One gene-one polypeptide hypothesis = every DNA
segment or gene codes for one specific protein.
Protein synthesis = the process that describes how these
proteins are made from DNA.
There are three steps to protein synthesis:
1. Transcription = DNA, which cannot leave the nucleus, is
made into mRNA which can leave the nucleus.
2. RNA Processing = mRNA undergoes a series of enzymeregulated modifications.
3. Translation = tRNA constructs a polypeptide at the
ribosome off of the mRNA.

Overview of
Protein
Synthesis

1.

Transcription

DNA is found in the nucleus of a cell and CANNOT leave the

nucleus.
DNA carries the code to make proteins.
Proteins are made at the ribosomes located in the cytoplasm
and OUTSIDE of the nucleus.
How, then, does the DNA code get out of the nucleus and to
the ribosome? a messenger is needed = mRNA.
mRNA is made off of the DNA template IN THE NUCLEUS
during transcription.
There are three steps of transcription:
1. Initiation = the cell recognizing the promoter region of the
DNA (the starting line).
2. Elongation = the addition of complementary RNA
nucleotides by RNA polymerase building a single-stranded
mRNA molecule.
3.
Termination = the cell recognizing the termination region of
the DNA (the finish line).

Overview of
Transcription:

1.

INITIATION = the cell recognizing the promoter region of

the DNA (the starting line of the gene).
i.
Helicase unzips the DNA double helix forming a Yshaped replication fork.
ii. Single-stranded binding proteins attach to each strand
of the uncoiled helix to keep them separated.
iii. Topoisomerase breaks and rejoins the double helix in
front of helicase to prevent knots.
iv. Transcription factors look for the promoter region =
a sequence of DNA nucleotides, and once found,
mediate the binding of RNA polymerase by flagging
down the enzyme and showing it where to bind.
v. The promoter region that indicates where transcription
is to begin is the sequence TATA and is called the
TATA Box.
vi. Once RNA polymerase binds to the TATA Box,
initiation is complete.

2.

ELONGATION = the addition of complementary RNA

nucleotides by RNA polymerase building a single-stranded
mRNA molecule.
i.
Once bound to the TATA Box, RNA polymerase begins
adding the complementary RNA nucleotides off of the
DNA template.
ii.
RNA polymerase moves off the template in the 3 5
direction (much like the DNA polymerase during DNA
replication).
iii. Unlike DNA replication, however, ONLY ONE SIDE OF
THE DNA IS TRANSCRIBED because mRNA is singlestranded LEADING STRAND.
iv. Remember, when an adenine DNA nucleotide is
transcribed, RNA polymerase places a uracil nucleotide
as its mRNA complement.

AU

TA

CG

GC

3.

TERMINATION = the cell recognizing the

termination region of the DNA (the finish line).
i.
RNA polymerase reaches a termination
sequence = a special sequence of mRNA
nucleotides that indicate the end of
transcription.
ii.
The termination sequence is AAUAAA once
RNA polymerase places these nucleotides,
transcription will stop some 10-35 nucleotides
later.
iii.
The termination sequence acts like the brakes
on a train once hit, it takes a bit longer for the
process to stop.
iv.
The full sequence of RNA nucleotides on the
mRNA molecule directly determines the order of
amino acids in the pending polypeptide.

2. RNA Processing

Before the mRNA is allowed to leave the nucleus it MUST

undergo a series of enzyme-regulated modifications.

There are three modifications made:

1. 5 GTP cap = a GTP is added to the 5 end of the mRNA.

GTP = guanine triphosphate guanine nucleotide

with 2 additional phophates attached (like ATP).

Provides stability to the mRNA and a point of

attachment of the molecule to the ribosome.
2. Poly-A tail = 150 to 200 adenine nucleotides added to
the 3 end of the mRNA.

Provides stability and controls the movement of

the mRNA across the nuclear envelope.
3. Cleaving of Introns = the removal of NON-CODING
segments from the mRNA.

mRNA contains two types of sequences: exons =

sequences of nucleotides that code for a polypeptide

and introns = intervening sequences that are NON-CODING.

The original unprocessed mRNA has exons and introns and is

called heterogeneous nuclear mRNA.

Small nuclear ribonucleoproteins (snRNPs) = biological

scissors, found in larger spliceosomes cut out the non-coding
introns and splice together the exons.
Once RNA processing is completed, the mRNA can leave the
nucleus and head to the ribosome in the cytoplasm.

3.

Translation

Occurs in the CYTOPLASM of the cell and on the ribosome.

The nucleotide sequence of the processed mRNA is read in triplets
called codons.
Each codon represents a specific amino acid (1 codon = 1 amino acid
BUT 1 amino acid DOES NOT EQUAL 1 codon).
The correct amino acids are brought to the ribosome by tRNA =
transfer RNA that contains the complementary anticodon for each
mRNA codon amino acids are connected to the correct tRNAs by
the enzyme aminoacyl-tRNA synthetase.
Amino acids are transferred to a growing polypeptide and attached via
peptide bonds.
There are three steps of transcription:
1.
Initiation = the ribosomal subunits come together and a start
codon (AUG) is read.
2.
Elongation = each successive tRNA brings the corresponding
amino acid to the growing polypeptide.
3.
Termination = one of three stop codons is reached and the
polypeptide is released from the ribosome.

Overview of Translation:

How each tRNA is connected

to the correct amino acid at
the 3 end:

1.

INITIATION = the ribosomal subunits come together

and a start codon (AUG) is read.
i.
The 5 GTP cap of the processed mRNA attaches to
the small ribosomal subunit.
ii.
The 1st tRNA with the anticodon UAC carrying the
amino acid methionine attaches to the mRNA start
codon, AUG, at the P site.

E = exit
P = polypeptide
A = arrival

i.

The large subunit attaches to the complex

completing the ribosome and ending initiation.

Initiation of translation:

2.

ELONGATION = each successive tRNA brings the

corresponding amino acid to the growing polypeptide.
i.
The 2nd tRNA arrives at the A site carrying the next
amino acid.
ii. Methionine is removed from the tRNA at the P site and
attaches via a peptide bond to the new amino acid.
iii. The whole complex shifts the empty amino acid is
now sitting at the E site and the tRNA with both amino
acids is now at the P site.
iv. The empty tRNA exits can again bind to a
methionine allowing it to repeat deliveries to other
ribosomes in the cell.
v. A 3rd tRNA arrives at the now empty A site steps ii.
through iv. are repeated until termination is reached.
vi. The hydrolysis of GTP into GDP and Pi drives
elongation.

Elongation

3.

TERMINATION = one of three stop codons is

reached and the polypeptide is released from the
ribosome.
i.
One of three stop codons is reached on the
mRNA (UAA, UGA, UAG) and a release factor (not
an amino acid) is brought in.
ii. The completed polypeptide, the last tRNA, and the
two ribosomal subunits are released.
iii. All the players can now attach elsewhere in the
cell and continue translation at other locations (all
materials are recycled).
iv. Once the primary structure of the polypeptide is
complete, interactions among the amino acids give it
its 2o, 3o, and 4o structure subsequent processing
by the E.R. and Golgi give final modifications to the
protein allowing it to function as a structural element
or enzyme.

Termination of translation:

IV. Mutations

Changes in the DNA of a cell.

Point mutations = changes in just one single base pair of a
gene.

Can have an adverse effect on the phenotype of an

organism and can be passed on to future generations.

E.g., sickle cell anemia change in a single amino acid in

the polypeptide hemoglobin which carries O2 in R.B.C.s

There are two types of point mutations:

1. Substitutions = the replacement of one nucleotide with
another three different types.
i.
Silent
ii. Missense
iii. Nonsense
2.
Frameshifts = insertions/deletions a single nucleotide
is either added or removed from a gene.

1.

Substitutions = the replacement of one nucleotide with

another three different types.
i.
SILENT = the change from one nucleotide to another has
NO effect on the amino acid inserted wobble = 61
coding codons and only 20 amino acids more than one
codon codes for a single amino acid. (e.g., if DNA
mutated from CCG to CCA, the mRNA codon GGC would
become GGU, but glycine would still be the amino acid
coded for).

ii.

MISSENSE = the change from one nucleotide to

another CHANGES the amino acid inserted
deleterious = changes shape/function of polypeptide.
(e.g., if DNA mutated from CCG to TCG, the mRNA
codon GGC would become AGC, and glycine would
be replaced with serine).

iii.

NONSENSE = the change from one nucleotide to another

results in a STOP codon deleterious = nonfunctional
polypeptide. (e.g., if DNA mutated from TTC to ATC, the
mRNA codon AAG would become UAG, and lycine would
be replaced with stop).

2.

Frameshifts = insertions/deletions a single

nucleotide is either added or removed from a gene
more disastrous effects on resulting proteins
than substitutions.
i.
INSERTIONS = a nucleotide is added to the
gene codon reading frame shifts and all the
nucleotides downstream of the mutation are
regrouped into new codons = all different amino
acids coded for results in a nonfunctional
protein.
ii.
DELETIONS = a nucleotide is removed from the
gene codon reading frame shifts and all the
nucleotides downstream of the mutation are
regrouped into new codons = all different amino
acids coded for results in a nonfunctional
protein.

V. DNA Organization
In eukaryotes, DNA is wrapped around proteins called
histones to form chromatin.
DNA histone complexes are called nucleosomes
appear like beads on a string through the electron
microscope.

In a non-dividing cell, DNA is arranged as two types of

chromatin:
1.
Euchromatin = regions where DNA is loosely attached
to nucleosomes DNA can be transcribed.
i.
Acetylated
Open palm
ii.
(demethylated)
2.
Heterochromatin = regions where the nucleosomes
are tightly compacted (more dense, stains darker)
DNA is inactivated and CANNOT be transcribed.
i.
Methylated
Closed fist
ii.
(deacetylated)

*Mechanisms for gene expression in eukaryotes gene

expression = an organisms ability to turn on genes ONLY
when proteins are needed.*
Turning on a gene = transcribing the DNA into mRNA.

Acetyl groups =
COCH3

O
Euchromatin
CH3 C

Methyl groups =
CH3

CH3 C

Heterochromatin

Some segments of
nucleotides within a DNA
molecule can move to new
locations = transposable
genes
transposons/jumping
genes (more common in
prokaryotes).
Transposons can move
within a chromosome or to
new chromosomes.
They act as mutations
may be inserted within
genes changing nucleotide
sequences and gene
expression.
They can turn on and
off expression or have no
effect at all.

VI. Molecular Genetics of Viruses

Viruses are cellular parasites penetrate a host = a cell

infected by a virus - take over all metabolic processes,
reproduce assembling hundreds of new viruses, and leave
the cell to infect others, destroying the host in its wake.
Not considered alive cannot reproduce on their own.
Specific for the types of hosts they infect/parasitize (e.g.,
bacteriophages/phages = viruses that ONLY infect
bacteria).
Consist only of nucleic acid (DNA or RNA) surrounded by a
protein = capsid, and sometimes viral envelopes =
membranes (phospholipids/proteins) surrounding a viral
capsid that assist in penetrating the host cell.
Viral envelopes have identifying glycoproteins (e.g., H1N1
virus refers to the specific glycoprotein on the outside of the
Swine Flus envelope. H5N1 represents a different type of
influenza avian).

Viruses reproduce by injecting their nucleic acid into the

genome of a host cell and using the host cells machinery
to replicate and transcribe their nucleic acid.

There are two basic replication cycles that viruses follow:

1. Lytic Cycle = the virus penetrates the cell membrane of
the host and uses the host cells enzymes to transcribe,
translate, and assemble new viruses, before lysing the
host and going on to infect other cells.
2. Lysogenic Cycle = the virus penetrates the cell
membrane of the host cell and temporarily incorporates
its nucleic acid directly into the host cells genome.
The virus remains dormant or inactive =
provirus/prophage, until a stimulus signals the virus to
transition into the lytic cycle and destroy the host cell.

Lytic vs. Lysogenic Cycles:

Some viruses have RNA as their nucleic acid rather than

DNA.
RNA viruses can use their nucleic acid directly as mRNA.
If an RNA virus infects a host cell with DNA, an alternative
method is needed to incorporate the viral nucleic acid into
the hosts genome = retroviruses.
HIV is a retrovirus.
Retroviruses need to transcribe DNA from their RNA
templates done by the enzyme reverse transcriptase.

VII. Molecular Genetics of Bacteria

Bacteria do NOT contain a nucleus or membrane-bound

organelles.
The primary genetic material is a naked chromosome NO
histones.
The naked chromosome is a single, circular molecule of DNA.
Bacteria reproduce by binary fission = simple mitosis the
chromosome replicates and the cell splits in two (asexual
clones).
Bacteria may also contain plasmids = short, circular DNA
molecule OUTSIDE of the chromosome genes of a plasmid
are beneficial but not necessary (e.g., antibiotic resistance = R
plasmid and sex pili = F plasmid).
Plasmids replicate INDEPENDENTLY of the naked
chromosome.
Some plasmids = episomes, CAN become incorporated into
the naked chromosomes.

Genetic variation occurs in bacteria in three ways:

1. Conjugation = DNA exchanged between two bacteria.
2. Transduction = new DNA is introduced via viral infection.
3. Transformation = DNA is absorbed from surroundings.

Conjugation

Transduction:

Transformation:

VIII. Prokaryotic Regulation of

Gene Expression

Best understood in the bacterium E. coli lives in the

digestive tracts of humans.
Genes are only transcribed and translated when a
feedback mechanism indicates that specific proteins are
needed
Genes are inducible = able to be turned on, or
repressible = able to be turned off.
Many proteins are made via a group of structural
genes that code for several enzymes which function in
the production of a single end product.
Structural genes are part of DNA operons = segments
of DNA that control gene expression.

Operons have four components:

1. Regulatory gene = produces the repressor which can
bind to the operon and block RNA polymerase from
moving down the DNA.
2. Promoter = sequence of DNA to which RNA polymerase
binds to begin transcription (TATA Box).
3. Operator = the sequence of DNA to which the repressor
binds to block RNA polymerase the genes on/off
switch.
4. Structural genes = the DNA segments to be transcribed.

There are two E. coli operons, one inducible and one

repressible:
1. Trp operon= produces enzymes for the synthesis of the
amino acid tryptophan.
i.
REPRESSIBLE (on, can be turned off).
ii. E. coli needs to produce tryptophan via the trp operon
the gene is on as RNA polymerase transcribes the
structural genes the regulartory gene is producing
an INACTIVE repressor = cannot bind to the operator
and block RNA polymerase.
iii. If tryptophan becomes available to the bacterium from
the environment, it no longer needs to produce the
amino acid on its own the gene needs to be turned
off or repressed tryptophan = corepressor - binds
to the inactive repressor CHANGES its SHAPE
making it ACTIVE can now bind to the operator and
block RNA polymerase transcription cannot occur.

trp Operon

2.

Lac operon = produces enzymes that breakdown

lactose.
i.
INDUCIBLE (off, can be turned on).
ii.
E. coli needs to breakdown/metabolize lactose
WHEN/IF it is present normally, the genes are
off as the regulatory gene produces an ACTIVE
repressor binds to the operator and blocks
RNA polymerase transcription of structural
genes that code for lactase enzymes CANNOT
occur.
iii.
If lactose becomes available to the bacterium, it
binds to the active repressor, CHANGES its
SHAPE making it INACTIVE it falls off of the
operator, inducing transcription as the RNA
polymerase can now move down the structural
genes transcribing the enzymes to breakdown the
lactose in the cell.

lac Operon

IX. Genetic Engineering

Genetic engineering = DNA/RNA technology used to

directly manipulate genes for practical
purposes/experimental study: genetically modified foods,
cloned animals, pharmaceuticals (human insulin), etc.
DNA technology results in recombinant DNA = DNA in
which genes from different sources (often different species)
are combined in vitro to make one molecule.
In vitro = in the lab.
In vivo = in life.
Recombinant DNA is made using restriction enzymes =
protein enzymes used to cut DNA each restriction enzyme
cuts DNA at a SPECIFIC location = recognition sequence.
Restriction enzymes cut the double helix leaving sticky ends
= staggered cuts in DNA that leave one strand longer than
the other.

restriction enzymes

sticky ends

(Recombinant DNA)
(Note: sticky ends are connected via DNA ligase
just as in DNA replication).

Various techniques are used in genetic engineering:

1. Plasmid Transformation = fragments cut from DNA by
restriction enzymes are inserted into bacterial plasmids
cut with the same restriction enzymes sticky ends
connect via DNA ligase the recombinant plasmid is
then introduced into a bacterium via transformation
the bacterium will produce the proteins for the inserted
gene (e.g., the human gene for the production of insulin).
2. Gel Electrophoresis = the separation of DNA cut with
restriction enzymes through a gel DNA is separated by
length (smaller segments move faster and will travel
further through the gel) separation occurs due to
charge DNA is negatively charged (PO4-) and therefore
travels to the positive pole used to identify the origin of
unknown DNA samples (e.g., crime scenes, paternity
tests, etc.)

Recombinant Plasmids

Plasmid Transformation

Gel
Electrophoresis

Analyzing an Electrophoresis Gel:

Which samples came from the same source AND WHY?
A
B
C
D
E
F
G

3.

Polymerase Chain
Reaction (P.C.R.) = a
technique used to
replicate billions of copies
of a specific DNA segment
without using bacteria
DNA polymerase,
synthetic primers, and
nucleotides are added to a
test tube with the DNA
segment and allowed to
replicate with automation
= heating and cooling
billions of copies of DNA
are made in hours not
days as it takes with
plasmids (e.g., amplify a
tiny DNA sample from a
crime scene for numerous
tests).