Breeding Techniques & Genetic Engineering – Comprehensive Study Notes
National News & Societal Context
Vitamin A Deficiency (VAD)
DOST-FNRI 2021 report: 1 out of 6 Filipino children are “most affected” by VAD.
Golden Rice (bio-fortified with provitamin A) is promoted as a counter-measure (article dated 30 Aug 2022).
Regulatory & Legal Updates on GM Crops in the Philippines
07 Feb 2020, 13:50 GMT: Golden Rice declared safe, yet farmer adoption remains uncertain.
19 Apr 2023 & 20 Apr 2023: Philippine Supreme Court issued a Writ of Kalikasan stopping commercial release of GM Golden Rice and Bt eggplant; petitions cite environmental and health concerns.
Protest motifs: “NO TO GMO / GOLDEN RICE!” banners illustrate persistent public resistance.
Philippine Agriculture Snapshot
Characterised as an agricultural country; crop & livestock production are pillars of food security and economic stability.
Fundamentals of Breeding
Definition: Breeding = controlled mating of two parents to obtain offspring with desired traits.
Applies to plants, animals, and conservation programs (captive breeding).
Objectives of Plant Breeding
Stress resistance: salinity, drought, frost.
Yield enhancement: vegetative parts & grains.
Pest/Insect resistance.
Disease resistance: bacterial, viral, fungal.
Processing quality improvement.
Nutrient fortification.
Objectives of Animal Breeding
Increased production: milk, eggs, meat, wool.
Enhanced meat quality.
Disease resistance.
Behavioural traits: improved docility, reduced aggression, good mothering ability.
Captive-Breeding Examples (Philippines)
Philippine crocodile (Crocodylus mindorensis)
Critically endangered; program at Silliman University, Negros Oriental.
Philippine eagle (Pithecophaga jefferyi)
Critically endangered due to habitat loss; breeding centre in Davao City.
Classical Plant Breeding Techniques
Selective Breeding
Repeatedly choosing individuals with superior traits within the same population.
Historical example: domestication of maize from wild grass teosinte; multi-generation selection for bigger kernels.
Crossbreeding / Hybridisation
Combines two superior traits from different breeds/species.
Resulting hybrids (e.g., modern corn, sunflower, cauliflower) display heterosis (hybrid vigour).
Process steps: parent choice, emasculation, pollen transfer, bagging & tagging, seed harvest.
Classical Animal Breeding Techniques
Inbreeding
Mating related individuals to reinforce specific traits.
Risks: reduced genetic diversity; accumulation of deleterious alleles.
Example: high-milk-yield cow mated to its sons.
Crossbreeding
Combines desirable traits of distinct breeds; offspring must be reproductively compatible.
Example: American Saddlebred × Arabian → National Show Horse.
Domestication & Artificial Selection Showcases
Gray wolf → domestic dog: long-term selective breeding for behaviour & morphology.
Wild mustard (Brassica oleracea) diversified into broccoli, cauliflower, cabbage, kale, kohlrabi, turnips via selection for different plant parts.
Limitations of Classical Breeding
Some traits (e.g., wheat stem-rust resistance) are hard or slow to incorporate.
Requires sexual compatibility; limited gene pool.
Modern Breeding (Post-Mendel) Techniques
Mutation Breeding
Induced mutagenesis (gamma rays, chemicals) to generate novel alleles; e.g., soybean.
Somatic Hybridisation
Fusion of protoplasts from different plants to combine genomes.
Artificial Reproductive Technologies (ART)
Artificial insemination; example: superior bull semen for cattle.
Genetic Engineering (GE)
Direct manipulation of DNA; transcends species barriers.
Genetic Engineering Overview
Central Question: “What if we directly introduce foreign genes to an organism so it can produce proteins it never naturally made?”
Capabilities:
Add, delete, over-express, or under-express genes.
Applicable when donor & recipient are reproductively incompatible.
Core technique: Recombinant DNA (rDNA) technology.
Historical Milestones in rDNA Technology
1972 Cohen & Boyer: transferred a plasmid segment between bacteria; created first transgenic E. coli.
1974 Rudolf Jaenisch: produced first transgenic animal (mouse).
1977 Genentech: synthesised human somatostatin in GM E. coli.
1983 Bevan, Flavell, Dell-Chilton: produced first transgenic crop (tobacco).
Terminology:
Plasmid = small, circular, extrachromosomal DNA capable of independent replication.
Transgenic organism / GMO = carries foreign DNA from another species ("trans = from the other").
rDNA Technology: Guiding Principles
DNA encodes proteins → proteins determine phenotype.
Steps (general):
Identify gene of interest (GOI).
Isolate GOI with restriction enzymes.
Insert GOI into a vector (plasmid).
Introduce recombinant vector into host organism.
Select & regenerate transformed cells/organisms.
Case Study: Bt Corn
Problem: corn borers (Ostrinia spp.) cause severe yield loss.
Solution: introduce gene from Bacillus thuringiensis (Bt) into corn genome.
Bt toxin forms pores in insect gut, leading to larval death.
Outcome: transgenic Bt corn resistant to borers; reduced pesticide use.
Molecular Tools & Enzymes
Restriction Enzymes
Werner Arber discovered their presence in bacteria (natural defence).
Hamilton Smith & Daniel Nathans mapped sequence specificity.
Example: EcoRI recognises / and cuts to generate sticky ends.
Key property: sequence-specific cleavage, enabling precise DNA manipulation.
DNA Ligase: seals sugar-phosphate backbone, joins GOI & vector.
Gene-Delivery Methods
Microprojectile Bombardment (Biolistics)
DNA-coated gold particles shot into cells.
Electroporation
Electric pulse creates transient pores in cell membrane.
Agrobacterium tumefaciens–mediated Transformation
Utilises Ti (tumour-inducing) plasmid; natural ability to insert DNA causes crown-gall tumours.
Engineered Ti plasmid carries GOI (e.g., ) into plant genome.
Additional GE Applications
Transgenic Mice
Microinjection of DNA into zygote nucleus; implanted embryo develops; offspring screened for insert.
Pharmaceutical E. coli
Human insulin gene ligated into plasmid → inserted into bacteria → large-scale fermentation → insulin purification.
Demonstrates GE’s pharmaceutical significance: affordable recombinant proteins (insulin, HGH, vaccines).
Standard Workflow for Creating Transgenic Crops (Detailed 7-Step Memory Aid)
Determine GOI.
Isolate GOI using restriction enzymes & gel electrophoresis.
Locate GOI with a labeled probe.
Combine GOI & plasmid via restriction enzymes + DNA ligase.
Reintroduce recombinant DNA into bacterial cell (vector amplification).
Allow bacteria (e.g., A. tumefaciens) to transform plant cells.
Regenerate whole plants from cultured cells.
Advantages of rDNA Over Classical Breeding
Bypasses sexual compatibility barriers.
Incorporates single, well-characterised genes without hitch-hiking undesirable alleles.
Faster trait deployment; precision edits (
Example rhetorical prompt from slides: “What makes recombinant DNA technology more powerful?” → answers above.)
Ethical, Environmental & Practical Considerations
Public resistance (e.g., Golden Rice protests) underscores biosafety, corporate control, ecological impact, and socio-economic debates.
Legal instruments (e.g., Writ of Kalikasan) highlight need for risk assessment, transparency, and inclusive stakeholder consultation.
Concept-Check / Discussion Prompts
Distinguish classical vs modern breeding (sexual recombination vs direct DNA manipulation).
Human application? ART & gene therapy exist, but germline editing raises ethical objections.
Potential problems in classical methods: linkage drag, limited gene pool, time-consuming.
Challenge scenario: Without knowledge of A. tumefaciens infection mechanism, plant GE would lack an efficient natural vector; scientists would rely more on biolistics or protoplast fusion.
True / False Self-Test (Slide 68 Key)
T — GE manipulates genomes for desired traits.
T — rDNA can up- or down-regulate genes.
F — Cohen & Boyer created first transgenic bacterium, not Werner Arber.
T — first GMO was E. coli carrying foreign plasmid.
T — bacteria engineered to produce human proteins (insulin, HGH).
Take-Home Message
rDNA technology is an evolving toolkit for agriculture, medicine, and industry.
Continuous improvements focus on gene identification, isolation, and delivery, pursuing sustainable solutions to global challenges such as food security, malnutrition (e.g., Vitamin A deficiency), and therapeutic protein supply.