MARY HELP OF CHRISTIANS COLLEGE – SALESIAN, INC.

Acacia St., Canlubang, Laguna

SY. 2023-2024

GENERAL BIOLOGY 2
Mutations and Genetic Engineering

MUTATIONS
• errors in genetic sequence
• caused by mutagens or errors during DNA replication or recombination

I. POINT MUTATION

EXAMPLE:

Original DNA strand: TAC TTC AAA CCG

mRNA sequence: AUG AAG UUU GGC
Amino acids produced: met – lys – phe - gly

TYPE EFFECT TO AMINO ACID SEQUENCE EXAMPLE

Silent amino acid produced from mutated strand is Mutated strand: TAC TTC AAA CCA
the same as the amino acid produced from mRNA: AUG AAG UUU GGU
original strand Amino acids: met – lys – phe – gly

Missense amino acid produced from mutated strand is Mutated strand: TAA TTC AAA CCA
different from the amino acid produced from mRNA: AUU AAG UUU GGU
the original strand Amino acids: ile – lys – phe – gly

Nonsense amino acid produced from mutated strand Mutated strand: TAC ATC AAA CCG
results to a stop codon mRNA: AUG UAG UUU GGC
Amino acids: met – stop

A. PRACTICE DRILL
Original DNA strand: AGG CGA TAT ATT GCA AAA TTA
mRNA sequence: ____________________________________
amino acids produced: ______________________________________________

1. Mutation 1: second guanine base is mutated into cytosine

Mutated DNA strand: _____________________________________________________________
Mutated mRNA sequence ________________________________________________________
Amino acids produced: ___________________________________________________________
Type of point mutation: ___________________________________________________________

2. Mutation 2: third thymine base is mutated into adenine

Mutated DNA strand: _____________________________________________________________
Mutated mRNA sequence: ________________________________________________________
Amino acids produced: ___________________________________________________________
Type of point mutation: ____________________________________________________________

II. FRAMESHIFT MUTATION

TYPE EFFECT TO THE GENETIC SEQUENCE EXAMPLE

Insertion nucleotides are inserted, changing the set of Mutated strand: TAC ATT CAA ACC A
codons and how they’re read mRNA: AUG UAA GUU UGG U
Amino acids: met – stop

Mutation: adenine base is inserted before

second thymine

Deletion nucleotides are deleted, changing the set of Mutated strand: TAC TTC AAC CA
codons and how they’re read mRNA: AUG AAG UUG GU
Amino acids: ile – lys – leu

Mutation: fourth adenine base was deleted

 A. PRACTICE DRILLS
Original DNA strand: AGG CGA TAT ATT GCA AAA TTA
mRNA sequence: ____________________________________
amino acids produced: ______________________________________________

3. Mutation 1: second and third thymine bases were deleted

Mutated DNA strand: _____________________________________________________________
Mutated mRNA sequence ________________________________________________________
Amino acids produced: ___________________________________________________________
Type of frameshift mutation: _______________________________________________________

4. Mutation 2: two adenine bases were inserted before the first cytosine
Mutated DNA strand: _____________________________________________________________
Mutated mRNA sequence: ________________________________________________________
Amino acids produced: ___________________________________________________________
Type of point mutation: ____________________________________________________________

GENETIC ENGINEERING

CLASSICAL BREEDING VS. MODERN GENETIC ENGINEERING

TECHNIQUE PROCESS ADVANTAGES / DISADVANTAGES

Classical Breeding • mating of two species with desired • exchanges are limited between the
qualities same or very closely related species
• natural process • no guarantee that desired
combination will be obtained
• takes a long time to achieve desired
results

Moderm Genetic • desired genes are directly introduced • genes are transferred regardless of
Engineering • genetic material is altered, cut, or whether species are closely related or
inserted not
• examples: in vitro fertilization, cloning, • switching on or off of genes is made
recombinant DNA technology, DNA possible
fingerprinting

I. RECOMBINANT DNA TECHNOLOGY

• involves cutting the DNA of interest and using a vector to propagate it
• vector – carries the designed DNA to a host cell (e.g., yeast, plasmid)

STEPS:

1. Obtaining the desired gene

- desired gene is selected; appropriate vector must be selected

2. Isolation of the desired gene

- desired gene is cut from the library plasmid at recognition sites using a restriction enzyme
• restriction enzyme – enzyme that recognizes and cuts foreign DNA at specific sites
EXAMPLE: EcoRI – cuts DNA at the AATTC sequence

3. Preparation of the transformation plasmid parts

- the desired gene, a selection marker gene, and “empty” transformation plasmid are cut
for compatibility in ligation

4. Ligation of the transformation plasmid parts

- the gene of interest is joined to the chosen vector using the enzyme ligase

5. Addition of desired gene to bacteria

- the transformation plasmid containing the desired gene, and the selection marker gene
were added to bacteria (or the host cell) for the replication of the recombined plasmid
• selection marker gene – gene sequence that is added to differentiate a transformed
plasmid from the non-transformed plasmids
- ensures that there are multiple copies of the recombined plasmid
 6. Isolation of the bacteria with the desired gene
- bacteria will be plated into a selective medium, and only those with the desired gene and
the selection marker gene will survive
- results to a supply of bacteria with the desired gene for future use

7. Separation of the desired gene

- the transformation plasmid with the desired gene is separated from the bacterial cells and
purified for future use

8. Transference of the desired gene

- an appropriate insertion method is chosen to transfer or insert the desired gene into the
organism
- there are different insertion methods, such as the following:
• biolistics – uses a gene gun to fire DNA-coated pellets on plant tissues
• heat shock treatment – uses heat to manipulate cell pore size
• electroporation – uses an electric “shock”

9. Propagating the genetically engineered organisms

- plant or organismal cells are grown on selective media so that only the transformed cells
carrying the new genes will grow

10. Testing the genetically engineered organisms

- plants or other organisms are tested to determine if they have incorporated the desired
trait

II. CRISPR-Cas9
• “gene editing” in living cells
• Cas9 – bacterial protein that helps in defending the bacteria against bacteriophage
infections; cuts any sequence to which it is directed
• gRNA – guides the Cas9 protein to a target gene; complementary to a target gene
1. Cas9-gRNA complex is introduced to a cell to be altered
2. Cas9 cuts both strands of the target DNA
3. broken ends of the DNA trigger a DNA repair system
4. repair enzymes rejoin the ends of the DNA → nucleotides are introduced or removed

III. GENE THERAPY

• introduction of genes to an afflicted individual for therapeutic purposes
• goal is to insert a normal allele of a defective gene into the body cells of the tissue affected
by the disorder

REFERENCES:
National Agriculture in the Classroom. (n.d.). Genetic Engineering. Retrieved from agclassroom.org

Urry, L.A., Cain, M.L., Wasserman, S.A., Minorsky, P.V., & Reece, J.B. (2017). Campbell Biology. Pearson Education,
Inc. pp. 413-437