GENETICS

OBJECTIVES
• At the end of the lesson, the students shall be able to:
1. Identify the mode of inheritance of a particular trait given the
pedigree;
2. predict the genotypes os the parents; and
3. compute the probability of occurrence of an affected offspring in
a given cross
DEFINITION OF TERMS
• PEDIGREE - making use of diagrams showing the ancestral
relationships and transmission of genetic traits over several
generation in a family
• PROBAND - the individual in the pedigree that led to the construction
of a pedigree
• LAW OF SEGREGATION (1ST MENDELIAN LAW) - for every trait
governed by a pair of alleles, these alleles segregate or separate
during gamete formation in meiosis
DEFINITION OF TERMS
• LAW OF INDEPENDENT ASSORTMENT (2ND MENDELIAN LAW) - A pair
of alleles for one trait will segregate or separate independently of
another pair of alleles for another trait during meiosis
• AUTOSOMAL TRAIT - a trait whose alleles that control it are found in
the autosomes (body chromosomes/ non-sex chromosomes)
• GENOTYPE - the gene pair an individual carries for a particular trait
symbolized by letters
• dominant allele - uppercase letters
• recessive allele - lowercase letters
DEFINITION OF TERMS
• HOMOZYGOUS DOMINANT TRAIT - with two dominant alleles
• HETEROZYGOUS TRAIT - with a dominant and recessive allele, the
individual showing the dominant phenotype
• HOMOZYGOUS RECESSIVE TRAIT - with two recessive alleles
• PHENOTYPE - the observable trait of an individual based on its
genotype
• Dominant - a trait that requires at least one dominant allele for the trait to be
expressed
• Recessive - a trait that requires two recessive alleles for the trait to be
expressed
DEFINITION OF TERMS
• PHENOCOPY - a trait that is expressed due to specific environmental
condition and is not due to genotype
• IDENTICAL TWINS - monozygotic twins; derived from a single
fertilization event and blastomores separate and become
independent mass of cells implanted in the mother's womb
• FRATERNAL TWINS - dizygotic twins; twins that arederived from
separate fertilization events resulting to two separate zygotes
MENDELIAN LAWS OF INHERITANCE
• LAW OF SEGREGATION
The law of segregation is based on four main concepts:
• Genes exist in more than one form or allele.
• Organisms inherit two alleles (one from each parent) during sexual
reproduction.
• These alleles separate during meiosis, leaving each gamete with
one allele for a single trait.
• Heterozygous alleles exhibit complete dominance as one allele is
dominant and the other recessive.
MENDELIAN LAWS OF INHERITANCE
• LAW OF INDEPENDENT
ASSORTMENT
- allele pairs separate
independently during the
formation of gametes.
Therefore, traits are
transmitted to offspring
independently of one
another.
PEDIGREE ANALYSIS
 WHAT IS A PEDIGREE ANALYSIS CHART?

• A pedigree chart displays a family tree, and shows the members of

the family who are affected by a genetic trait.
 USES OF A PEDIGREE ANALYSIS

• Describe the mode of inheritance of a trait

• Calculate the probability of occurrence an affected offspring in a given
cross
SYMBOLS IN PEDIGREE ANALYSIS
WHAT TO EXPECT IN A HUMAN PEDIGREE
•For autosomal dominant trait: Two
affected individuals can have a normal
offspring
•For autosomal recessive trait: Two
affected individuals can NEVER have a
normal offspring
• A. Look at the family of IV-9 and IV-10. If the trait is dominant, is it
possible for them to have an affected offspring?
• B. If the trait is recessive, is it also possible for IV-9 and IV-10 to have
an unaffected offspring?
• C. Based on your answers for a) and b), is the trait dominant or
recessive?
• D. Give the genotypes of the following:
• i. IV-9
• ii. IV-10
• iii. V-1
• iv. I-1
• v. I-2
• E. If IV-9 and IV-10 were to have another child, what is the probability
that they will have an affected offspring?
•A. Is the trait dominant or recessive?
•B. What are the most probable genotypes
of I-2 and I-3?
•C. What is the probability that II-2 is Dd?
•D. What is the probability that II-1 and II-2
will have another normal offspring?
ACTIVITY
Make a pedigree of the following:

• I. With (dominant) or without finger hair (recessive)

• II. Left-handed (dominant) or right-handed (recessive)
• III. Widow’s peak (dominant) or straight hairline (recessive)
• IV. Free (dominant) or attached earlobe (recessive)
• V. Curly (dominant), wavy (heterozygous) or straight
(recessive) hair
• VI. Can tongue-roll (dominant), cannot tongue-roll
(recessive)
QUIZ
• Determine the genotypes of all the individuals in the pedigree. (14pts)
• Is the trait dominant or recessive? (2pts)
• What is the probability that II-2 is Dd? (1pt)
• If III-2 will marry a heterozygote, what is the probability that they will have a normal
offspring? (1pt) What will be the probability that they will have an affected offspring?
(1pt)
• Give an example of a dominant Mendelian trait from the activity yesterday. (1pt).
Make a pedigree of the following:

• I. With (dominant) or without finger hair (recessive)

• II. Left-handed (dominant) or right-handed (recessive)
• III. Widow’s peak (dominant) or straight hairline (recessive)
• IV. Free (dominant) or attached earlobe (recessive)
• V. Curly (dominant), wavy (heterozygous) or straight
(recessive) hair
• VI. Can tongue-roll (dominant), cannot tongue-roll
(recessive)
SEX LINKAGE AND RECOMBINATION
OBJECTIVES
At the end of the lesson, the students shall be able to:
1. illustrate the transmission of an X-linked and a Y-linked
character;
2. compute the probability of the occurrence of a sex-linked trait;
and
3. give examples of other sex-related traits.
DEFINITION OF TERMS
• SEX LINKED TRAIT - the gene pair that determines a
character is located on the sex chromosome
• X-LINKED TRAIT - a sex-linked trait is where the gene or
allele for the trait is found on the X chromosome
• COLOR BLINDNESS - an X-linked recessive trait where an
affected individual could not distinguish red from green
• HEMOPHILIA - an X-linked recessive trait where an individual
suffers delayed blood clotting during injuries because of the
absence of certain blood clotting factors
DEFINITION OF TERMS
• Y-LINKED TRAIT - a sex-linked trait where the gene or allele
for the trait is found on the Y chromosome
• HYPERTRICHOSIS PINNAE AURIS - a Y-linked trait where the
affected males have hair growing from the external ears
• SEX-INFLUENCED TRAIT - any trait in a diploid organism
whose expression is affected by an individual's biological sex;
a trait that occurs at a higher frequency in one sex over the
other
DEFINITION OF TERMS
• SEX-LIMITED TRAIT - any trait in a diploid organism
whose expression is limited to just one biological sex
 SEX-LINKED TRAITS
• X-linked traits are sex-linked traits where the gene or allele
for the trait is found on the X chromosome
• In humans, males and females are represented by different
sex chromosomes
• Females have two X chromosomes in the nucleus of their
cells while males have one X chromosome and one Y
chromosome in the nucleus of their cells
• Depending whether the trait is dominant or recessive, the
expression pattern of the trait differs in males and females
It is said that sex-linked traits occur more
frequently in men. Why?
What happens to the expression patterns
if the trait is X-linked and dominant?
 SEX-LINKED TRAITS
• Y-linked traits are sex-linked traits where the gene or allele
for the trait is found on the Y chromosome
• Only males will express the trait since females do not have Y
chromosomes
 SEX-INFLUENCED TRAITS
• Sex influenced traits are traits in a diploid organism whose
expression is affected by an individual's biological sex; a trait
that occurs at a higher frequency in one sex over the other
• Hormonal or physiological differences between the sexes
cause differences of expression of certain genes
• Baldness in humans is an example of a sex-influenced trait
 SEX-LIMITED TRAITS
• Sex-limited traits are traits in a diploid organism whose expression is
limited to just one biological sex
• Hormonal or physiological differences between sexes may limit the
expression of some genes to one biological sex only
• Only females can express functional mammary glands that produce
milk immediately after giving birth
In a cross between a white-eyed female fruit fly and
red-eyed male,
what percent of the female offspring will have white
eyes? (White eyes are X-linked, recessive)

A. 100%
B. 25%
C. 50%
D. 75%
E. 0%
In a cross between a pure bred, red-eyed female fruit
fly and a white-eyed male, what percent of the male
offspring will have white eyes? (white eyes are X-linked,
recessive)

A. 100%
B. 75%
C. 50%
D. 25%
E. 0%
Hemophilia in humans is due to an X-
chromosome mutation. What will be the
results of mating between a normal (non-
carrier) female and a hemophilac male?
A human female "carrier" who is
heterozygous for the recessive, sex-linked
trait red color blindness, marries a
normal male.
What proportion of their female progeny
will show the trait?
• The last Emperor of Russia, Nicolas II, was married to
Empress Alexandra, and they had five children, Olga, Tatiana,
Maria, Anastasia, and Alexis. Alexis was the only one who
was afflicted with hemophilia or the royal bleeding disease;
all other members were normal.
• If only Prince Alexis was afflicted with the disease,
determine his genotype.
• What could be the genotypes of the Emperor and Empress?
• Is it possible that each daughter could have been a carrier?
How?
MODIFICATION TO MENDEL'S CLASSIC RATIOS
OBJECTIVES
At the end of the lesson, the students shall be able to:
1. distinguish Mendelian from non-Mendelian modes of
inheritance
2. describe some cases of non-Mendelian genetic traits
DEFINITION OF TERMS
• Co-dominance - When two contrasting alleles are present in
the same locus or trait (heterozygote genotype), then the
phenotype expressed is a “blend” of the two extreme
phenotypes. The two genes interact and the offspring shows
the effects of both alleles.
• Incomplete dominance - When two contrasting alleles are
present in the same locus or trait (heterozygote genotype),
then both alleles are expressed in the same phenotype
DEFINITION OF TERMS
• Multiple alleles - When there are more than two types
of alleles for a given locus or trait, this will result in
more than two kinds of phenotypes that may be
expressed for that trait.
RECALL: MENDELIAN RATIOS
• LAW OF SEGREGATION
(monohybrid cross of F1
generation)
• GENOTYPIC RATIOS:
1:2:1
(homozygous dominant : heterozygous :
homozygous recessive)
• PHENOTYPIC RATIOS: 3:1
(dominant : recessive)
RECALL: MENDELIAN RATIOS
• LAW OF INDEPENDENT
ASSORTMENT (dihybrid
cross of F1 generations
• GENOTYPIC RATIOS :
1:2:1:2:4:2:1:2:1
• PHENOTYPIC RATIOS:
9:3:3:1
INCOMPLETE DOMINANCE

• When two contrasting

alleles are present in the
same locus or trait
(heterozygote genotype),
then both alleles are
expressed in the same
phenotype
• phenotypic ratio is the same
as the genotypic ratio
 CODOMINANCE
• When two contrasting alleles are
present in the same locus or
trait (heterozygote genotype),
then the phenotype expressed is
a “blend” of the two extreme
phenotypes. The two genes
interact and the offspring shows
the effects of both alleles.
• the phenotypic ratio is the same
as the genotypic ratio
INCOMPLETE DOMINANCE VS. CODOMINANCE
 MULTIPLE ALLELES
• There are more than
two types of alleles,
and the relationship of
each allele with respect
to others will
determine the number
of phenotypes that may
be expressed.
MULTIPLE ALLELES

• in the ABO system, the O

allele is recessive to both A
and B alleles while the A and
B alleles are co-dominants of
one another
TRY THIS:
A local hospital has sent word to a family of a possible mix up of
some of the children with other families when they were born.
To rule out any possible mix up, the hospital obtained the blood
types of every individual in the family, including the surviving
maternal grandfather and paternal grandmother. The results
were as follows:
Father: Type O 3rd child: Type B
Mother: Type A Maternal grandfather: Type AB
1st child: Type O Paternal grandmother: Type B
2nd child: Type A
TRY THIS:
Based on the results, is there a possibility that any one of the
children is not a biological offspring of the couple?
TRY THIS:
In cattle, coat color is inherited in a co-dominant fashion. Homozygous
B1B1 produces black coat, homozygous B2B2 produces white coat, and
heterozygous B1B2 produces roan coat. Give the phenotypic ratio of the
following crosses:
A. B1B1 x B1B1
B. B1B1 x B2B2
C. B1B2 x B1B2
D. B1B1 x B1B2
E. B1B2 x B2B2
TRY THIS:
In a hypothetical plant, a serrated leaf margined plant, when crossed
with a smooth leaf margined plant, produces offsprings with wavy leaf
margin.
A. Identify the mode of inheritance.
B. Two serrated plants, when crossed, will give what type of offspring?
C. Two wavy plants will produce what possible kinds of offspring? Give
their ratios?
TRY THIS:
A man who is blood type B is married to a woman who is
blood type A. None of the man’s parents is blood type O.
This couple has 4 children with the following blood types: B,
AB, AB and O. Give the genotypes of the parents.
TRY THIS:
In guinea pigs, coat color is governed by four alleles that
constitute a multiple allelic series, C (black), cS (sepia), cC
(cream), and c (albino) with the following dominance
hierarchy: C>cS>cC>c. Determine the phenotypic ratios of
the progeny from the following crosses:
A. Cc x CcS
B. CcS x cCc
EDMODO CLASS CODE
•CARET: awkgfb
•TILDE: mvjk2c
•OBELUS: 3e4csq
1. Amparo’s Farmville is the most prominent rice supplier in the
country. They produce different kinds of rice that fit the Filipinos’
tastes. The dominant traits produce tall and flood resistant rice
plants. What will be the phenotypes of the following crosses?
A. AaBb X AaBb B. AaBb X Aabb
2. Muscular dystrophy is an X-linked recessive trait. Determine the
phenotypic ratios of a cross of (a) a normal carrier female and a
normal male; and (b) a normal female and an affected male.

3. Given: RR = red, Rr = pink, rr = white

Determine the phenotypic ratios of the cross of the following:
a. RR x Rr b. Rr x Rr
4. Given: AA = black Aa = roan aa = white
Determine the phenotypic ratio of the following:
a. Aa x Aa b. Aa x aa

5. In rabbits, coat color is governed by four alleles that

constitute a multiple allelic series, C (wild type), cch
(chinchilla), ch
(Himalayan), and c (albino) with the following dominance
hierarchy: C>cch>ch>c. Determine the phenotypic ratios of
the progeny from the following crosses:
A. cchch x cchc B. Ccch x chc
QUIZ
1. Mang Juan’s Garden is well-known for supplying blue
roses all over the country. He experimented on breeding
different colors of roses to produce different hues of
blue roses. Given that the dominant trait produces a
dark blue rose and the recessive, a white rose, what will
be the phenotypes of the following crosses? Note:
Heterozygous roses have light blue color.

A. BB X bb
B. Bb X Bb
2. Bantay, your 3-year old male cat with beautiful yellow
fur, is close with your crush’s female cat which has
yellow and black coat. In cats, the gene for coat color is
sex-linked: YY gives yellow coat color and yy gives black,
but the heterozygote has a peculiar combination of
yellow and black, a condition known as tortoise-shell.
Show the types of offspring expected from a cross
between Bantay and your crush’s cat (genotypes and
phenotypes). NOTE: Your cat’s love life is not always
directly proportional to yours.
In man, red-color blindness, depends on a X-
linked recessive allele. Bernadette, whose
father is color blind, but who is not herself a
“color blind” marries Phillip, a normal man.
What are the genotypes of their children?
What is the chance of red-color blindness to
appear in their children?
George, who has Type A blood, marries
Jennifer with Type B blood. Their child, Faye
has Type O blood. What are the genotypes of
these three individuals? What phenotypic
ratio, would you expect in future offspring
from this marriage?
Amparo’s Farmville is the most prominent rice
supplier in the country. They produce
different kinds of rice that fit the Filipinos’
tastes. The dominant traits produce tall and
flood resistant rice plants. What will be the
phenotypes of the following crosses?

A. AaBb X AaBb
B. AABb X Aabb
TEST YOUR KNOWLEDGE
1. making use of diagrams showing the A. Proband
ancestral relationships and transmission of
genetic traits over several generation in a family B. Sex-limited trait
2. the individual in the (1) that led to the
construction of (1) C. Pedigree
3. for every trait governed by a pair of alleles, D. Sex-influenced trait
these alleles segregate or separate during
gamete formation in meiosis E. Codominance
4. any trait in a diploid organism whose F. Incomplete dominance
expression is affected by an individual's
biological sex; a trait that occurs at a higher G. Law of Segregation
frequency in one sex over the other
5. The heterozygote genotype is expressed as a
H. Law of Independent
distinct phenotype (a “blend” of the two Assortment
extreme phenotypes).
TEST YOUR KNOWLEDGE

A man with hemophilia (a recessive, sex-linked condition) has a

daughter of normal phenotype. She marries a man who is normal for
the trait. What is the probability that a daughter of this mating will be
a hemophiliac? That a son will be a hemophiliac? If the couple has
four sons, what is the probability that all four will be born with
hemophilia?
TEST YOUR KNOWLEDGE

A man with hemophilia (a recessive, sex-linked condition) has a

daughter of normal phenotype. She marries a man who is normal for
the trait. What is the probability that a daughter of this mating will be
a hemophiliac? That a son will be a hemophiliac? If the couple has
four sons, what is the probability that all four will be born with
hemophilia?
TEST YOUR KNOWLEDGE

A man with Type A blood marries a woman with Type B blood. Their
child has Type O blood. What are the genotypes of these three
individuals? What genotypes, and in what percentage, would you
expect in future offspring from this marriage?
MOLECULAR STRUCTURE OF DNA, RNA,
and PROTEINS
DIAGNOSTIC QUIZ
1. What are the building blocks of DNA?
2. What are the different parts of #1? (Clue: There are three parts)
3. What does DNA stand for?
4. What does RNA stand for?
5. What is the function of DNA?
6. What is the function of RNA?
7. What is a protein?
8. Give four levels of protein synthesis.
9. What is the function of proteins?
OBJECTIVES
At the end of the lesson, the students shall be able to:
1. describe the building blocks of DNA, RNA and proteins;
2. identify the structural and functional differences between DNA
and RNA;
3. explain the different levels of protein structure; and
4. explain how protein structural features may influence their
functions
NUCLEIC ACIDS
NUCLEOTIDES
NUCLEOSIDES
NITROGENOUS BASES
• Contain carbon and
nitrogen
• “bases” – contain an
amino group that can
bind to an extra
hydrogen
• Can be classified as
purines or pyrimidines
• Adenine and Guanine
• Two carbon-nitrogen
rings
• Cytosine, thymine and uracil
• Single carbon – nitrogen ring
DEOXYRIBOSE VS. RIBOSE
SPECIFIC BASE PAIRING
CHARGAFF’S RULE
COMPUTATION OF DIFFERENT BASE
PAIRS
1 TURN OF THE MAJOR GROOVE
= 10 base pairs
10 bp= 34 Angstroms
1 Angstrom = 1 nm
PROBLEM SOLVING

Assume 102 base pairs of DNA to be

responsible for transcription of particular
mRNA molecule. What is the length of the
mRNA in (a) Angstroms (b) in micrometers?
Conversion units: 34 Angstroms = 10bp
1nm = 10Angstroms
1um = 1 x 10^3 nm
• Made up of three bases
in the mRNA
• Each codes for a single
amino acid
• Single stranded
• Contains Uracil instead of
Thymine
• Carries the message from
DNA which controls all pf
the cellular activities in a
cell
• Synthesized in the nucleus
and then transported in
the cytoplasm
• Made up of three bases
in the mRNA
• Each codes for a single
amino acid
CODON CHART
 PROBLEM SOLVING

The chain of hemoglobin in man is 144

amino acids long. What would be the
length (in nucleotides) of the mRNA
molecule required to direct the synthesis
of this polypeptide chain?
• Major constituent of
the ribosomes
• Has enzymatic activity
(peptidyl transferase)
and catalyzes formation
of the peptide bonds
among amino acid
chains
• One of the smallest of
the four types of RNA
• Carries the correct type
of amino acid to the
site of protein synthesis
• Most abundant organic molecules in living
systems
• Most diverse range of functions of all
macromolecules
• Structural, regulatory, contractile, or protective
• Serve in transport, storage, or membrane
components
• Toxins or enzymes
• Catalysts of
chemical reactions
• Catabolic –
enzymes for
breakdown
• Anabolic –
enzymes for
building complex
molecules
• Enzymatic reactions increase
as temperature increases
until it reaches optimum
temperature
• If the temperature continues
to increase, the enzymes get
denatured
• Most enzymes in the human
body have optimum
temperature at 37 degrees
Celsius.
• Optimum pH is unique
for every enzyme.
• A decrease or increase
in pH depending on its
affinity may change its
shape and lose its
function
• If enzyme concentration
remains constant,
enzyme activity
increases as substrate
concentration increases
(if active sites are
available)
• monomers that make up
proteins
• 20 amino acids (10
essential amino acids)
• Can be acidic, basic,
polar or nonpolar
• Amino acid linkages
• Bonds formed : PEPTIDE BOND
• Chains of amino acids - polypeptide
FOUR STRUCTURAL LEVELS OF PROTEINS
• Chains of amino acids from translation process
• Local folding of the polypeptide. Most common
: alpha-helix and beta-pleated sheets
• Three-dimensional structure of a polypeptide
• Protein subunits forms interactions with other
subunits
• DENATURATION
• If the protein is subject to changes in temperature,
pH, chemicals, the protein may change its shape and
lose its functions.
• Can be reversible
SUMMARY OF IMPORTANT PHYSICAL PROPERTIES
 QUIZ
1. All of the statements are true, except
I. The building blocks of any nucleic acid are the nucleosides.
II. DNA containing high A-T content are more stable.
III. The primary structural level of proteins consist of sequence of amino
acids in the polypeptide chain.

A. I and III B. II and III C. I and II D. I, II, III are false.

 QUIZ
2. Which of the following composition ratio confirms Chargaff’s
Rule?

A. G x T/ A = C
B. pyrimidines/purines = 1
C. A+T/G+C = 1
D. None of the above
3. A DNA fragment produced 315 amino
acids then encountered a stop codon.
How long (in nucleotides) is the mRNA?

4-5. 13,540 base pairs were present in a

DNA fragment. How long is the DNA in
a) Angstroms
b) micrometers
Conversion units: 34 A=10 bp; 1nm = 10A; 1000nm= 1um
 QUIZ
6-10. Complete the table below.
Molecule Feature Function
6. Double-stranded helix 7.
RNA 8. Encoded DNA,
transcribes mRNA and
translates mRNA to
protein
9. Folded chains of amino 10.
acids
DNA REPLICATION AND
PROTEIN SYNTHESIS
OBJECTIVES
At the end of the lesson, the students shall be able to:
1. describe the requirements, proteins and enzymes in
DNA replication, transcription and translation;
2. diagram the steps in replication, transcription and
translation; and
3. explain what happens to a gene sequence that
undergoes transcription and eventual translation into
protein
CENTRAL DOGMA OF MOLECULAR
BIOLOGY
DNA REPLICATION
DNA REPLICATION
• DNA synthesis
• transfer of information from cell to cell or from generation to
generation
• DNA is replicated in a semi-conservative manner
• newly synthesized strand contains an old strand from the parent
DNA and a newly synthesized DNA strand
INITIATION OF REPLICATION
• HELIX-UNWINDING PROTEIN (HELICASE) uses ATP to unwind dsDNA
by breaking H-bonds between the complementary strands
• To keep the strands from reannealing, several SINGLE STRAND
BINDING PROTEINS (SSBPs) bind to a ssDNA
• The enzyme DNA GYRASE relaxes the twisting tension created on the
DNA during unwinding
• Once the templates are exposed, short stretches of segments of RNA
are synthesized by PRIMASE to serve as primers for the elongation of
the complementary strand
ELONGATION OF THE COMPLEMENTARY
STRAND
• Once the primers are set, DNA POLYMERASE III adds nucleotide
triphosphates complementary to the DNA template from the 3' OH
end of the primer
• The complementary strand that is synthesized using the 3' to 5'
template elongates in the same direction as the unwinding of the fork
is called the LEADING STRAND
• The complementary strand of the 5' to 3' direction but opposite the
direction of the replication fork is called as the LAGGING STRAND
ELONGATION OF THE COMPLEMENTARY
STRAND
• The replication of the 5' to 3' template requires multiple primers
producing multiple discontinuous segments of complementary DNA
called OKAZAKI FRAGMENTS
• The enzyme DNA POLYMERASE I proofreads the DNA molecule and
removes the primers by digestion by its exonuclease activity starting
from the 3' to 5' direction
• After its replacement of excised primers, it leaves breaks or nicks in
the replicated DNA segments which are sealed by an enzyme called
LIGASE
TERMINATION OF REPLICATION
• Replication ends when all the parent DNA nucleotides have
been complemented
• The two daughter molecules are exact copies of each other
and the parent DNA
SUMMARY
ACTIVITY ENZYMES INVOLVED
Helicase
Single-stranded Binding
Initiation Proteins
Gyrase
DNA REPLICATION Primase
DNA Polymerase III
Elongation DNA Polymerase I
Ligase
2 copies of semiconservative
Termination
DNA
DNA TRANSCRIPTION
DNA TRANSCRIPTION
• RNA synthesis
• In eukaryotes, transcription occurs within the nucleus, and
the RNA products are transported to the cytoplasm for
translation into proteins
• Prokaryotes synthesize their RNA and proteins in the
cytoplasm resulting in coupling of transcription and
translation
INITIATION OF TRANSCRIPTION
• RNA POLYMERASE attaches to a specific sequence in the
DNA called the promoter which is located upstream to the
INITIATION SITE where the transcription actually starts
• SIGMA FACTOR recognizes the promoter sequence so the
RNA polymerase can selectively bind to it. Once bound, the
sigma factor disassociates and is free for another round of
initiation
ELONGATION OF TRANSCRIPTION
• RNA POLYMERASE moves along strand of DNA strands to
produce single stranded messenger RNA.
TERMINATION OF TRANSCRIPTION
• When RNA polymerase reaches termination point, protein
rho factor binds to the enzyme and dissociates it from the
DNA to stop transcription.
SUMMARY
ACTIVITY ENZYMES INVOLVED
RNA polymerase

Initiation
Sigma factor
DNA TRANSCRIPTION
RNA polymerase
Elongation

Termination Rho Factor

DNA TRANSLATION
ACTIVATION OF AMINO ACIDS
• amino acids are attached to
their respective tRNAs via
the action of the enzyme
AMINO-ACYL SYNTHETASE
at the expense of ATP
DNA TRANSLATION (initiation)
INITIATION OF TRANSLATION
• INITIATION FACTOR 1 (IF1) stimulates the disassociation of the small
and large subunits of ribosomes
• INITIATION FACTOR 3 (IF3) binds to the 30S subunit to prevent
reassociation of the 30S and 50S subunits and binds to the 30S
subunit to the start codon of the mRNA
• INITIATION FACTOR 2 (IF2) + GTP attaches the first amino-acyl-tRNA
to the 30S subunit
• IF1 attaches to the 30S subunits then 50S subunit joins the initiation
complex to disassociate IF3 and IF1 form the 30S subunit
• GTP is hydrolyzed to GDP + Pi that disassociates IF2
DNA TRANSLATION (elongation)
ELONGATION OF THE PEPTIDE CHAIN
• Once the Fmet is set at the P-site with ELONGATION FACTOR Tu
(EFTu) + GTP, a new amino acid-tRNA complex enters the A site
complementary to the succeeding codon
• PEPTIDYL TRANSFERASE catalyzes the formation of peptide bonds
between the Fmet and the amino acid carried by the tRNA in the A
site
• The tRNA in the P-site is released to the cytoplasm for recycling which
is followed by the movement of the ribosome along the mRNA in the
5' to 3' direction through the action of ELONGATION FACTOR G (EFG)
+ GTP
DNA TRANSLATION (termination)
TERMINATION OF TRANSLATION
• RELEASE FACTOR 1 (RF1) would recognize UAA and UAG while
RELEASE FACTOR 2(RF2) would recognize UAA and UGA
• RELEASE FACTOR 3 (RF3) aids either RF1 or RF2 and binds to the
ribosome with GTP
• Once the polypeptide has been released, the ribosome disassociated
through the action of initiation factor I preparing the ribosomal units
for another round of translation
• The polypeptide chain may undergo folding to become functional or
may undergo further processing
SUMMARY
Activity Enzymes Involved
Initiation Factor 1
Initiation Initiation Factor 3
Initiation Factor 2
Elongation Factor Tu
RNA Translation Elongation Peptidyl Transferase
Elongation Factor G
Release Factor 1
Release Factor 2
Termination
Release Factor 3
Initiation Factor 1
 A. RNA POLYMERASE
B.LIGASE
a. REPLICATION 1. INITIATION
C. RELEASE FACTOR 1

D. INITIATION FACTOR 3
E. RHO FACTOR
F. DNA POLYMERASE I b. TRANSCRIPTION 2. ELONGATION
G. RELEASE FACTOR 3

H. ELONGATION FACTOR Tu
I. GYRASE c. TRANSLATION 3. TERMINATION
J. ELONGATION FACTOR G
GENETIC ENGINEERING
OBJECTIVES
At the end of the lesson, the students shall be able to:
1. compare classical breeding with modern genetic engineering
techniques;
2. enumerate the steps in molecular cloning;
3. describe methods to introduce DNA into cells; and
4. explain the selection and screening of transformants/ geneticall
modified organisms (GMOs)
CLASSICAL BREEDING
CLASSICAL BREEDING
• Classical breeding practices focus on the mating of
organisms with desirable qualities.
DNA RECOMBINATION
I. cutting or cleavage of DNA by restriction enzymes (REs)
II. selection of an appropriate vector or vehicle which would
propagate the recombinant
DNA ( eg. circular plasmid in bacteria with a foreign gene of interest)
III. ligation (join together) of the gene of interest (eg. from animal)
with the vector ( cut
bacterial plasmid)
DNA RECOMBINATION

IV. transfer of the recombinant plasmid into a host cell (that

would carry out replication to
make huge copies of the recombined plasmid)
V. selection process to screen which cells actually contain the
gene of interest
VI. sequencing of the gene to find out the primary structure of
the protein
INTRODUCTION OF RECOMBINANT DNA
Biolistics.
• “gene gun” is used to fire DNA-coated pellets on plant
tissues
• Cells that survive the bombardment, and are able to take
up the expression plasmid coated pellets and acquire the
ability to express the designed protein.
INTRODUCTION OF RECOMBINANT DNA
Plasmid insertion by Heat Shock Treatment.
• transfer plasmid DNA into bacteria.
• The target cells are pre-treated before the procedure to increase the
pore sizes of their plasma membranes. This pretreatment (usually
with CaCl2) is said to make the cells “competent” for accepting the
plasmid DNA.
• After the cells are made competent, they are incubated with the
desired plasmid at about 4°C for about 30min. The plasmids
concentrate near the cells during this time.
INTRODUCTION OF RECOMBINANT DNA
Plasmid insertion by Heat Shock Treatment.
• Afterwards, a “Heat Shock” is done on the plasmid-cell solution by
incubating it at 42°C for 1 minute then back to 4°C for 2 minutes.
• The rapid rise and drop of temperature is believed to increase and
decrease the pore sizes in the membrane.
• The plasmid DNA near the membrane surface are taken into the cells
by this process.
• The cells that took up the plasmids acquire new traits and are said to
be “transformed”.
INTRODUCTION OF RECOMBINANT DNA

Electroporation.

• This technique follows a similar methodology as Heat Shock

Treatment, but, the expansion of the membrane pores is done
through an electric “shock”.
• This method is commonly used for insertion of genes into mammalian
cells.
SCREENING RECOMBINANT CELLS
• Selection of plasmid DNA containing cells
• Selection of transformed cells with the desired gene
• PCR detection of plasmid DNA
 APPLICATION OF
RECOMBINANT DNA
OBJECTIVES
At the end of the lesson, the learners will be able to
1. give examples of products from recombinant DNA technology
2. illustrate the use of databases to search genes for desired traits
3. describe steps in PCR to amplify and detect the gene of interest
4. identify the parts of an expression vector
5. explain how genes may be cloned and expressed
“Once a desired trait is chosen,
information must be acquired for
either its detection or expression in
a given organism.”
DETECTION
• PCR AMPLIFICATION
PCR amplification is an in-vitro method that simulates DNA
replication in vivo. It utilizes a thermostable (heat-resistant)
DNA polymerase that builds single stranded DNA strands unto
unwound DNA templates.

NOTE: unlike DNA replication in vivo, PCR reactions do not use

too many helper enzymes such as helicases and gyrases to
help denature and stabilize the template DNA strands.
PCR AMPLIFICATION
PCR uses repeated cycles of incubation at different
temperatures to
promote:
1. the unwinding of the DNA template (~95°C);
2. the annealing of a primer (a ~20bp oligonucleotide
sequence (recall RNA primers in DNA replication) onto the
ssDNA template strand (~54 - 60°C); and
3. the extension of the generated ssDNA strand through the
binding of complementary bases to the template strand
(~72° C).
PCR RESULTS
• The expected product of PCR amplification will depend on the
sequences / position at which the primer sequences bind.
CLONING AND EXPRESSION
BASIC COMPONENTS OF AN EXPRESSION
PLASMID
• PROMOTER - Allows the controlled expression of the
desired gene in the presence of an inducing agent
(e.g. beta- galactosidase; heat treatment (~65°"C)
• MULTIPLE CLONING SITE - DNA sequence or portion
for the insertion of the desired gene.
• INSERTED GENE SEQUENCE - Successful insertion of
a gene allows the expression of its protein product.
This usually provides a specific trait to the
“transformed” bacteria.
BASIC COMPONENTS OF AN EXPRESSION
PLASMID

• ANTIBIOTIC RESISTANCE GENE - Provides a way to

screen a population of bacteria for those that took
up the plasmid.
TEST YOUR KNOWLEDGE. Correct or Wrong.
1. Classical breeding practices focus on the
mating of organisms with desirable qualities.
2. Heat-shock treatment is used to fire DNA-
coated pellets on plant tissues
3. Electroporation is commonly used for bacterial
cells.
TEST YOUR KNOWLEDGE. Correct or Wrong.
4. PCR reactions do not use too many helper
enzymes such as helicases and gyrases to help
denature and stabilize the template DNA strands.
5. The antibiotic resistance gene provides a way
to screen a population of bacteria for those that
took up the plasmid.