Chapter 11:
DNA Biology
and
Technology
Chargaff’s Rules
DNA contains 4 different types of
nucleotides based on nitrogencontaining bases
•
•
Adenine (A) and Guanine (G) are
purines with a double ring
Thymine (T) and cytosine (C) are
pyrimidines with a single ring
o
The amount of A, T, G, and C in
DNA varies from species to species
o
In each species, the amount of A
always equals the amount of T; the
amount of G always equals the
amount of C
•
•
A can only par with T
G can only pair with C
Franklin X-Ray Diffraction
Rosalind Franklin was a
researcher at King’s College in
London
Studied the structure of DNA
using X-ray crystallography
X-ray diffraction of DNA
showed that DNA is a double
helix
•
•
Indicated by the X pattern
The dark bands at the top
and bottom indicate that
portions of the helix are
repeated
Watson and Crick Model
o
Structure of DNA:
1.
2.
3.
o
DNA is a polymer of 4 types of
nucleotides (A, T, C, G)
The amount of A=T, and the
amount of G=C
DNA is double helix with a
repeating pattern
Model:
•
•
The deoxyribose sugar-phosphate
molecules make up the sides of the
twisted ladder
The nitrogenous bases make up
the rungs of the ladder


A and T ( 2 hydrogen bonds)
G and C (3 hydrogen bonds)
DNA Replication
The process of copying a DNA molecule
o
DNA is found in the nucleus
Steps in replication:
1.
2.
3.
Helicase unwind the old strands
and each old strand serves as a
template for the new strand
New complementary nucleotides
are lay down by the process of
complementary base pairing
The complementary nucleotides
join to form new strands
•
•
Each daughter DNA contains an
old and a new strand
(semiconservative)
This step is carried out by DNA
polymerase and ligase
RNA Structure and Function
RNA is found in the nucleus and
cytoplasm of eukaryotic cells
It contains the sugar ribose and base A,
U, C and G
RNA is single-stranded
3 forms of RNA
1.
Messenger RNA (mRNA) is
produced in the nucleus
-
2.
Transfer RNA (tRNA) is produced in
the nucleus
-
3.
DNA serves as a template for its
formation during transcription
tRNA transfer amino acids to the
ribosomes
20 different tRNAs
Each type carries only one type of
amino acid
Ribosomal RNA (rRNA) is produced
in the nucleolus of the nucleus
-
rRNA joins with proteins made in the
cytoplasm to form the ribosomes
Proteins are synthesized at the
ribosomes
Comparison of DNA and
RNA
Gene Expression
The central dogma of molecular biology:
genetic information flows from DNA to
RNA to protein
Gene expression requires transcription
and translation
•
•
o
Transcription takes place in the
nucleus (DNAmRNA)
Translation takes place in the
cytoplasm (mRNA  amino acids in
a protein)
Gene expression occurs when a gene’s
product (the protein) is functioning in
the cell
•
•
Proteins differ from one another by
the sequence of their amino acids
Proteins determine the structure and
function of cells
Genetic Code
DNA and RNA are written in a
different language from protein
20 amino acids found in proteins
Triplet code: 43 = 64 different
triplets
Codon: three-letter (nucleotide)
unit of an mRNA codes for a
single amino acid
•
•
61 triplets correspond to a
particular amino acid
3 stop codon signal the end of
a polypeptide
Note: Each of the codons are composed of 3 letters.
EX: Find the rectangle where C is the first base and A is the
second base
U, C, A, or G can be the third base
CAU and CAC are codons for histidine; CAA and CAG are
codons for glutamine
Transcription
Resulted in a strand of RNA that is complementary to a portion
of DNA
All 3 classes of RNA are formed
Process of Transcription:
1.
RNA polymerase binds to a promoter
2.
RNA polymerase opens up the DNA helix and adds new
RNA nucleotides that are complementary to the template
DNA strand
3.
The newly synthesized primary-mRNA is processed by
•
The addition of a cap to one end
•
The addition of a poly-A tail to the other end
•
The removal of introns so that only exons remain
o
The mature mRNA now contains instructions for assembling a
protein
o
Alternative mRNA splicing where the cells use only certain
exons rather than all of them to form the mature RNA
transcript
•
Can increase the possible number of protein products that
can be made from a single gene
Translation: Overview
Translation requires mRNA, tRNA, and rRNA
Ribosomal RNA (rRNA) is made in the nucleolus within the
nucleus
tRNA brings amino acids to the ribosomes
rRNA joins with proteins manufactured in the cytoplasm to
form the large and small ribosomal subunits
tRNA is single-stranded in a cloverleaf-like shape
At least one tRNA molecule for each of the 20 amino
acids
•
The amino acid binds to one end of the
molecule
•
The opposite end of the tRNA contains an
anticodon (codon complementary to the codon
of mRNA)
•
The anticodon pairs with an mRNA codon when
the tRNA-amino acid complex arrives at the
ribosome
•
o
o
The subunits leave the nucleus and join together to form a
ribosome in the cytoplasm
A ribosome has a binding site for mRNA and 2 sites (P and A
sites) for two tRNAs.
•
These binding sites allow complementary pairing of the
mRNA codons with the tRNA anticodons
•
The amino acid carried by the tRNA is added to the growing
chain of polypeptides
Several ribosomes can attach to and translate the same mRNA,
allowing the cell to produce many copies of the same protein at
a time
Translation
Translation has 3 phases
1.
2.
3.
Initiation: the small ribosomal subunit, mRNA,
and the initiator tRNA, (bound to the amino
acid methionine), and the large ribosomal
subunit all come together
•
The small ribosomal subunit attaches to the
mRNA with the start codon (AUG)
•
The anticodon on the initiator tRNAmethionine complex pairs with this codon
•
The large ribosomal subunit joins to the small
subunit
Elongation: new amino acids are added to the
growing polypeptide
•
The tRNA at the P site bears a peptide
•
The tRNA at the P site transfers its peptide to
tRNA-amino acid at the A site and leaves the P
site
•
The tRNA-peptide translocates to the P site
and the codon at the A site is ready for the
next tRNA-amino acid
Termination: the end of translation
•
A release factor binds to the stop codon and
cleaves the polypeptide from the last tRNA.
•
The ribosome detaches from the ER
•
Ribosomal subunits and the mRNA dissociate
Transcription and Translation
Genes and Gene Mutations
Gene mutations can be caused by replication errors,
transposons, or environmental mutagens
Transposons are specific DNA sequences that can move within
and between chromosome
•
They are known as the the “jumping genes”
•
They often disrupt genes, making these genes nonfunctional
o
Mutagens are environmental influences (radiation, X-ray, UV
light, pesticides, and cigarette smoke) may cause mutations in
DNA
o
Types of mutations:
1.
Silent mutations are ones that go undetected because they
have no observable effect
•
2.
Point mutations are those that involve a change in a single
nucleotide
•
3.
The severity depends on whether it affects one or more
codons of a gene
The severity depends upon the particular base change that
occurs
Frameshift mutations are caused by extra or missing
nucleotides in a DNA sequence
•
Often make the protein nonfunctional because it no longer
makes sense
Recombinant DNA
Recombinant DNA (dDNA) contains
DNA from two or more different sources
A plasmid is commonly used as a vector
to act as a carrier for the foreign DNA
Restriction enzymes cleave DNA at
specific places both on the foreign DNA
and on the plasmid forming “sticky ends”
Foreign DNA is inserted into the the
plasmid and DNA ligase will seal the
foreign DNA into the plasmid
Gene cloning is achieved when a host cell
takes up the recombinant plasmid and as
the bacteria cell replicates, the plasmid is
replicated; multiple copies of the gene are
made
Transgenic Organisms
Recombinant DNA technology is used to
produce transgenic bacteria, plants, and animals
Gene products are collected from the growth
medium
•
•
o
Transgenic plants such as cotton, corn, and potato
are resistant to pest by producing an insect toxin
•
o
Insulin, human growth hormone, tPA
(tissue plasminogen activator), and hepatitis
B vaccine
Degrade substances such as oil and nuclear
wastes
Advantageous in increasing crop yields
Gene “pharming” is the use of transgenic farm
animals in producing pharmaceuticals
•
Genes are incorporated into an animal’s
DNA and the proteins appear in the
animal’s milk
Polymerase Chain Reaction
PCR can make millions of copies of a segment
of DNA very quickly without the use of a vector
or a host
•
o
PCR is very sensitive in amplifying a DNA
sequence in a very small sample
Steps of PCR
•
•
The mixture of DNA and primers are
heated to 95°C to separate the two strands
of the DNA
The temperature is lowered so that the
primers can anneal (bind) to the single
strands of DNA and DNA polymerase can
copy the DNA using the supplied
nucleotides
o
The amount of DNA doubles with each
replication cycle so that trillions of copies can be
made in just 20 to 30 PCR cycles
o
Uses: creating recombinant DNA, decipher
evolutionary history of the human populations,
criminal analysis
DNA Fingerprinting
DNA fingerprinting makes use repeated noncoding
segments of DNA
PCR is used to amplify the repeated region
People can be heterozygous for the number of repeats so
both homologues has to be amplified separately
After amplification, DNA is ran through a gel in
electrophoresis
•
•
The longer segments travel slower than the
shorter segment; thus, the shorter segment
migrates further
Uses:
1.
To identify the presence of a viral infection or a
mutated gene that could predispose someone to
cancer or genetic illnesses
2.
To identify a suspect in forensics
3.
To identify the parents of a child or identify the
remains of someone who died (9/11)
Caveat: bioethical implications that can lead to discrimination and
invasion of privacy