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ERT 101 Biochemistry
Assignment 4
1. The following is a code for a strand of DNA
DNA
3’ A T A T T
1
2
3
4
5
A C T T T
6
7
8
9
G
C
A
C
G
t
L y
G
U
tRNA
M e
G
A
C
T
10 11 12 13 14 15 16 17 18 19
mRNA
Amino
T
s
a
r
g
U
G
G
t
h
r
U
G
A
e
n
d
Acid
a) Using the genetic code provided (appendix), fill in the blanks to complete the
segment of DNA shown.
b) Fill in the blanks to complete the sequence of amino acids coded for the
complementary strand of DNA
c) Write the code for the complementary strand of DNA completed in part a.
5’ TATA ATG AAA CGT ACC TGA 3’
d) What would be the effect if C were substituted for T at base 10?
No change. Coded for the same amino acid lysine. Silent mutation
e) What would be the effect if A were substitute for G at base 11?
Cysteine substituted for arginine. Coded for different amino acid. Missense
mutation.
f) What would be the effect if G were substituted for T at base 14?
Proline substituted for threonine. Coded for different amino acid. missense
mutation.
g) What would be the effect if C were inserted between base 9 and 10?
Major differences in amino acid. Frameshift mutation.
h) How would UV radiation affect this strand of DNA?
Adjacent thymines might polymerize
i) Identify a nonsense sequence in this strand of DNA.
ACT – stop codon
2. Match the following examples of mutagens.
______b_______ A mutagen that is incorporated into DNA in place of a normal base
______d_______ A mutagen that causes the formation of highly reactive ions
______c_______ A mutagen that alters adenine so that it base-pairs with cytosine
______a_______ A mutagen that causes insertions
______e_______ A mutagen that causes the formation of pyrimidine dimmers
a. frameshift mutagen
b. nucleoside analog
c. base-pair mutagen
d. ionizing radiation
e. nonionizing radiation
3. Identify three ways of preventing mistakes in DNA.
Light repair, dark repair, proofreading by DNA polymerase OR
Photoreactivation repair (or repair of pyrimidine dimers)
Excision repair (base excision repair)
Recombinational repair (mis-match repair)
4. Why are mutation and recombination important in the process of natural selection
and the evolution of organisms?
Mutation and recombination provide genetic diversity. Environmental factors
select for the survival of organisms through natural selection. Genetic diversity is
necessary for the survival of some organisms through the processes of natural
selection. Organisms that survive may undergo further genetic change, resulting
in the evolution of the species.
5. PCR replication of DNA is similar to bacterial population growth. If scientist,
starting PCR with 15DNA helices, runs the reaction for 15 cycles, how many
DNA molecules will be present at the end? Show your calculations.
The formula for growth of a bacterial population is: original number of cells 2n
where n is the number of generations. In PCR n is the number of cycles;
therefore, in this case, there will be 491,520 DNA molecules (15 x215)
N = No2n
Where N = final cell number, No = initial cell number, n = the number of
generation that have occurred during the period of exponential growth
N = 15 x 215
6. The polymerase chain reaction (PCR) is a technique to produce a large number of
identical molecules of DNA in vitro. Describe in detail the PCR technique.
Principles of PCR:
 Denaturation.
 Exposure to heat (about 94°C)
 separates the two strands of the target DNA by breaking the hydrogen
bonds between base pairs but otherwise leaves the two strands unaltered.
 Priming.
 A mixture containing an excess of
 DNA primers (synthesized such that they are complementary to
nucleotide sequences near the ends of the target DNA),
 DNA polymerase, and
 an abundance of the four deoxyribonucleotide triphosphates (A, T,
G, and C)
is added to the target DNA
 This mixture is then cooled to about 65°C, enabling double-stranded DNA
to reform.
 Because there is an excess of primers, single strands are more likely to
bind to a primer than to one another.
 The primers provide DNA polymerase with the 3' hydroxyl group it
requires for DNA synthesis.
 Extension.
 Raising the temperature to about 72°C increases the rate at which DNA
polymerase replicates each strand to produce more DNA
These steps are repeated over and over, so the number of DNA molecules increases
exponentially
7. By referring to the Genetic Code,
(i)
What DNA nucleotides triplet codes for codon UGU?
The DNA triplet for this codon is ACA.
(ii)
Identify a base-pair substitution that would produce a silent mutation at
this codon.
A change in the DNA to ACG (mRNA, UGC) produces a silent mutation, that is, it
codes for the same amino acid;
(iii)
Identify a base-pair substitution that would result in a missense mutation
at this codon.
a change in the
DNA to ACA (mRNA, UGU)
ACT (mRNA, UGA)
ACC (mRNA, UCU)
ACG (mRNA,UGC)
AGA (mRNA, UCU)
AAA (mRNA, UUU)
ATA (mRNA, UAU)
ACA (mRNA, UGU)
TCA (mRNA, AGU)
CCA (mRNA,GCU )
GCA (mRNA,CGU )
results in a missense mutation, that is, a different amino acid;
(iv)
Identify a base-pair substitution that would produce a nonsense mutation
at this codon.
and a change in the DNA to ACT (mRNA UGA) produces a nonsense mutation,
that is, a stop in transcription.
8. List and describe the steps in prokaryotic DNA replication. How does this process
appear to differ from eukaryotic DNA?
Briefly, prokaryotic DNA replication consists of
DNA unwinding,
RNA primer formation (Primer synthesis),
DNA synthesis catalyzed by DNA polymerase, and
the joining of Okazaki fragments by DNA ligase.
Prokaryotic DNA replication differs from the eukaryotic process in that
prokaryotic replication is faster, and
in prokaryotes the Okazaki fragments are longer
9. Define and describe the roles of the following in replication.
a. DNA ligase
b. DNA polimerase III
c. primase
d. helicase
e. Okazaki fragments
a. DNA ligase– an enzyme that links DNA segments; responsible for linkage
of Okazaki fragment during DNA synthesis
b. DNA polimerase III– the major DNA synthesizing enzyme in prokaryotes
c. primase– an RNA polymerase that synthesizes short RNA segments, called
primers, that are required in DNA synthesis
d. helicase– an ATP-requiring enzyme that catalyzes the unwinding of duplex
DNA during DNA synthesis
e. Okazaki fragments – short DNA strands that are synthesized during
discontinuous replication of the lagging DNA strand as the leading strand
is continuously replicated