Biotechnology for Engineer

(SKKK4413)
Semester 2015/2016-11
April, 2016

Chapter 6
Biotechnology In Agricultural Industry

Muhammad Tahir (Ph.D)

Department of Chemical Engineering,
Faculty of Chemical and Energy Engineering,
University Technology Malaysia
 Course Outline
Weeks Chapters Topic Test/Assign/Quiz
1 1 Introduction to Biotechnology -
2 2 Tools in biotechnology -
3 3 Biotechnology in Food and Quiz #1
Beverages Industry

4 4 Microbial Biotechnology Assign # 1

5, 6, 7 6 Biotechnology in Agriculture Quiz#2, Test 1
(Plant, Animal and Marine)

8, 9 7 Medical Biotechnology Assign # 2

10-11 8 Environmental Biotechnology Quiz #3, Test 2
12-15 Bioethics forum Assign# 3,
Group presentation
16 Revision week -
17-19 Final Examination Final exam
 Topic Outcomes
Topic Outcomes
Week Topic
#
Biotechnology in Agriculture (Plant,
Animal and Marine)
 Plant Tissue Culture and
Applications
 Application of Plant genetic
Engineering
5,6,7  Genetic Engineering in Animals
 Transgenic Animals
 Animal Diseases
 Animal Propagation
 Conservation Biology
 Biotechnology in Aquaculture
 Products from Marine Sources
 Algal Cell Culture
 Transgenic Fish
 Conservation of Marine
Environment
 Biotechnology in Agriculture-Public
Concerns and The Future
Learning Outcome
It is expected that students will be able to:
 appreciate the contribution of
biotechnology in plant, animal and
marine life in order to increase product
quantity and quality.
 design techniques used to produce
genetically modified transgenic plants,
animals and marine life
 identify marine based products that can
be exploited by biotechnology
techniques
 Appreciate and identify the role of
biotechnology in the conservation of
certain endogenous species.
 Chapter 6: Biotechnology In
Agricultural Industry

6.1 Animal Biotechnology

6.2 Plant Biotechnology
6.3 Marine Biotechnology

Bongo Antelope

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 6.1: Animal biotechnology

Introduction
1. Started as selective breeding over 10,000 years
ago, slow and restricted.
2. Recomb DNA tech.; improve quality and quantity
of products.
3. Animals as products and as bioreactor for health or
food products
 Products and services of biotechnology in animals

1) Producing Transgenic Animals,

2) Control, reduced and eliminate animal diseases
3) Animal propagation
4) Animal conservation

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 Technology involves

1. Gene transfer methods

2. Artificial insemination
3. Selective breeding

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 1. Gene transfer methods

3 techniques
a) Microinjection
b) Embyogenic stem cell gene transfer
c) Retrovital gene transfer
 a) Microinjection
Technique:
cloned gene is transferred to fertilised eggs of donor animal
– Donor females are superovulated (more than 1 ova) and mated
– Fertilised eggs removed and male and female pronuclei
separated.
– Male pronucleus is injected with DNA by thin needle
– Implanted eggs are transferred to speudopregnant female.
– Transgenics are screen by southern blotting
– Success rate differ from one animal to another
– Low gene targeting efficiency
 Microinjection

PROBLEMS OF MICROINJECTION

• Few injected eggs survive

• Not all surviving eggs retain the
new DNA
• The new DNA might be integrated
at random and not at specific
sites
• Homologous recombination and
gene targeting efficiency are low
in animals
• Sometimes not all cells in the
transgenic animal are
transformed
• Sometimes expression of
incorporated gene is low or
nonexistent
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b) Embryonic stem cell gene transfer

1. Embryonic stem (ES) cell culture

transfected with specific gene.
2. Transformed ES are microinjected
into animal blastocysts so that it
will be established in somatic and
germ line tissues
3. Successful in mice (so far) due to
pluripotency
4. Transformed mice extensively used
for study of human genetic
disorder diseases

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 c) Retroviral gene transfer

• Retrovirus efficiently effect animal cells and integrated into

genomes
• Use disable virus (by gene deletion) as vector
 2. Transgenic animals
– Functions:
• Animals are used as human model for many diseases eg:
cancer studies
• Used as bioreactor-living factories eg: produce specific protein
in cows milk
• Use to produce quality products
• Use to produce products in higher quantity eg:
• Transgenic fish-bigger in size
• Cow: resistance to diseases/reduce fat
• Birds: resistance to virus/low cholesterol eggs
 2. Transgenic animals-cont
• Techniques:
• Use retrovirus vectors
• Use of blastoderm cell, transformed with liposomes
into subgerminal space of host embryo (see Fig 7.6)
 2. Transgenic animals-cont
– Mice
• Use as model study of human diseases
– Cows/ goats
• As living factories
– Production of IL-2, 1-antitrypsin, clotting factor
IX
– Secreted in milk and purified
• Transgenic cows
– Costly and time consuming
– Possible of producing
» high protein -casein (increase cheese in
milk)
» Viral, bacterial and other pathogen resistant
breed
Max-founder cow producing » rHBT(recombinant bovine somatotropin)
human Lactoferrin, or hLF in milk. produced in transformed E. Coli injected in
cows results in increase 25% milk
production (deemed safe by FDA)
 3. Treatment or prevention of
animal diseases

• Treatment/prevention:
• Animal threats and diseases:
• Production of vaccines,
a. Bacterial and viral infections
monoclonal antibodies and
b. Fungal and Insects (ticks
and lice) antibiotic
• Diagnostic kits and parasite
antigen detection kits
• Health supplements
• Growth hormones
 Common Diseases in animals and Hope of
Biotechnology Approaches
Diseases Causes Biotechnology Approaches
Trypanosomiasis Transmitted by tse tse fly Isolation of tryposome-resitance
gene for vaccine development

Theileriosis Caused by tick-transmitted Cattle dipped into chemical

(East coast fever) protozoans.Parasites formed sporozoites in
tick’s salivary glands and transferred to a
bitten cattle. Infected lymphocytes formed
leukemic and lympholysis occurs
immunisation
insecticide, resistance developed
Search for sporozoites vaccine for
cattle

Foot and mouth disease Died by myocarditis Vacinated by inactivated virus

(30% cloven-hoofed (Infected animals killed
animals infected world Milk and meat cannot be eaten)
wide)

Coccidiosis Parasitic protozoan (Eimeria) Coccidiostat Vaccine in trail

Infection of epithelial cell of digestive tract
and associated glands
Example of Treatment or prevention of animal diseases

Annie, born March 3, 2000, is

a clone of a pure-bred Jersey
calf whose cells were
modified with genes for
producing lysostaphin, a
protein that kills
Staphylococcus aureus
bacteria, a leading cause of
mastitis disease in dairy
cows. She is the first
transgenic cow clone
engineered to resist mastitis,
which costs the U.S. dairy
industry $1.7 billion annually.
Photo by Scott Bauer
 4. Animal Propagation

– Artificial insemination
• A diluted sperm sample from 1 bull can inseminate
500 to1000 females
• Methods available >40 years ago
 4. Animal Propagation-cont
Animal clones
• Embryo twinning, splitting or cloning
• Could create more eggs in human in vitro
fertilisation
• Nuclear transfer methods
– Pluoripotent nuclei (capable of forming any cell type in an
individual) are taken out from embryonic cells and
transferred into donor oocytes and eggs (lacking a
functional nucleus or pronucleus), which are
subsequently transferred into surrogate female to
produce animals that are genetic clones of one another
and the nuclei donor (Fig 7.7)
– Dolly sheep (1/277 cloning attempts)
• Producing identical twins
– Mice (1 manipulated and 1 as control)
 5. Animal Conservation
1. Function:
1. Preserve habitats,
2. Re-established wild
population for later use
2. Techniques:
Bongo Antelope i) In vitro fertilisation
ii) embryonic transfer,
iii) oocytes maturation,
Malaysian Ox-Gaur iv) control of ovarian cycle and
transgenic livestocks,
v) cryopreservation of semen,
gametes and embryos in
germplasm banks
Elands
Preservation of Bongo Entelope (Kenya)
(Example: Embryonic transfer in Animal Conservation)
Method:
1. Female is superovulated and artificially inseminated with selected
bull sperm.
2. Fertilised eggs or embryos retrieved from the female and frozen in
liquid Nitrogen tank (-196 C)
3. Embryos can be implanted into surrogate mothers that are made
speudopregnant with hormones.
4. Easy transport of ET around the world
a) Holstein cow as SM for rare Malaysian ox (Gaur) in 1984 and
bongo antelope
b) Elands antelope SM of Kenya rare bongo antelope
6.2: Plant Biotechnology

The uses of plants.

1. Food / medicinal/energy sources
2. Furniture
3. Building structures etc
4. Environmental and ecosystem control
The need for biotechnology:
1. To improve quantity and quality
2. To reduce harvest time
3. To prevent and control diseases
4. To assist mass propagation of plantlets
5. To breed and produce new varieties of plants
1. Production of haploids by anther and ovary culture
(gametoclonal variants), useful for cereals.
6. To produce biopharmaceutical products (bioreactor)
7. Storage of essential germ plasm instead of conventional
seeds
8. Embryo rescue
 General techniques in plant biotechnology:
– Micropropagation (rapid propagation or mass production of identical
clones
• Oil palm plantlets (1960’s in Malaysia) with 30% improve in oil yield.
• Orchids farms in Thailand, Malaysia, SEA
– Protoplast fusion
– Plants from callus
– Somatic embryogenesis
– Somaclonal variation
– Gametoclonal variation
– Valuable germ plasm
– Chemicals from plants
• Genetic engineering of plants
– The Chilton Experiment
– Sanford experiment
– Two methods for gene transfer into plants (Fig 14.5)
 Biotech Application In Plant Farming
• Insect Resistance
• Virus Resistance
• Herbicide Resistance
• Plant As Bioreactors
• Enhanced Nutrition
 Insect Resistance
What is Bt Corn?
• Bacillus thuringiensis (Bt), a bacterium that occurs
naturally in soil, used to kill certain insects
• Bt contains a gene taken from Bt bacteria. Gene produce
protein that kills European corn borer
• Target species for Bt are: European corn borer, Southwest
corn borer, black cutworm, armyworm and corn
rootworm.
• No effect on humans, animals or other non-target species
• Corn that has been modified to include Bt helps farmers
and the environment by reducing the need to spray
pesticides on the crop
•Scientist find a type of Bt that
makes a protein to kill the insect.
•The gene that produces the
protein is taken from the bacteria
and inserted into the plant.
•The corn borer eats come of the
Bt corn plant.
•The Bt protein in the corn plant
dissolves the corn borer’s stomach
wall.
•The cells in the corn borer’s
stomach burst. It stops eating and
dies in a few days.
 Virus Resistance
• Many crops are lost to viral diseases. Infection leads to
reduced growth rate, poor crop yield, and low crop
quality.
• Genetic engineering provides a desirable alternative to
chemicals.
• Focus is on isolating genes involved in resistance to
diseases caused by viruses, bacteria and fungi. (Examples
of virus –resistance crops : melons, tomatoes, potatoes
• One option is to inject the plant with vaccines. Vaccines
contain dead or weakened strains of the plant virus. The
vaccine turns on the plant’s version of immune system,
making it invulnerable to the real virus.
• The coat protein genes of a number of viruses are being
used to genetically engineer resistance in crops.
• Among the viruses are cucumber mosaic virus, alfalfa
mosaic virus, tobacco streak virus and have been used
in crops such as tomato, tobacco, papaya, and potato.
 •Potato
• Papaya –The "NewLeaf" line of
– Outbreak of Papaya Ring Spot Virus Bt-potatoes include
(PRSV) swept through the papaya two additional virus-
plantations of Hawaii papaya resistant varieties,
production dropped 40% in the NewLeaf Plus and
course of 5 years. NewLeaf Y.
– Control of the disease was limited
–These include the Bt
to careful observation to remove
gene plus genetically
infected plants.
engineered resistance
– Developed two varieties of GE to potato viral diseases
papaya resistant to PRSV using coat (caused by the Potato
protein-mediated resistance. Leaf Roll Virus and
– Seed of the two varieties, named Potato Virus Y,
"SunUp" and "Rainbow," have been respectively).
freely distributed to papaya growers
since May of 1998.
 Herbicide Resistance
• Frequently used in agriculture to control weeds.
• Weeds reduce crop yields because they compete
for soil nutrients, water, and sunlight with crops.
• Broad-spectrum herbicides kill all (or most) plants
• A a crop plant is genetically engineered to be
resistant to a broad-spectrum herbicide, weed
management could be simplified to an application
of a single herbicide without concern of damaging
the crop plant itself.
• How does genetically engineered herbicide resistance
work?
– the genetically engineered plant produces a new
protein which detoxifies the herbicide, or
– the protein in the plant which is normally the target of
the herbicide's action is replaced by a new protein
which is unaffected by the herbicide.
• Examples of herbicide
– Glyphosate (corn, soybeans, cotton, canola,
sugarbeets)
– Glufosinate (corn, soybeans, cotton, canola, rice,
sugarbeets)
– Bromoxynil (cotton)
– Sulfonylurea (cotton,flax)
 Glyphosate Herbicide
• Affects an important metabolic pathway in plants by
blocking an enzyme called "3-enolpyruvylshikimate-5-
phosphate synthate“ (EPSPS)  plant dies
• Glyphosate is a broad-spectrum herbicide-- that is, all
green plants are killed by glyphosate
• Resistance to glyphosate is engineered into a plant by
adding a gene from a soil bacterium (Agrobacterium sp)
that encodes a glyphosate-resistant version of the EPSPS.
• Glyphosate-resistant plants have an additional gene
genetically engineered into them that also aids glyphosate
resistance.
• Glyphosate resistant crops that are now on the market,
includes corn, soybeans and cotton.
 Plant As Bioreactors
• Foreign gene is inserted in plant to produce large scale of
proteins of therapeutic and industrial value. (same
principles in microbes)
• An inexpensive, readily available complex metabolites
could be used as a nutrient source and cells could convert
this compound into a valuable product, which
accumulates until the cells are harvested.
• Whole plant is harvested and product will be extracted.
• Instead of placing the selected gene in a bacteria or yeast,
or producing antibodies in a laboratory mouse, it is
inserted into a plant, which is cheaper, cleaner and purer,
as plant organisms do not possess viruses that are
transferable to humans
• The cost of producing foreign compounds in plants is
much lower than producing them in bacteria and
mammalian cells.
• Soybean plants have been used as bioreactors to
produce a variety of monoclonal antibodies with
therapeutic value.
• Genistein is an important nutraceutical molecule found
in soybean.
• Potential health effects include: prevention of breast,
colon, and prostate cancers, cardiovascular disease and
post-menopausal ailments.
 Enhanced Nutrition
• Biotech gives the opportunity
to save millions of people from
effects of malnutrition
• Golden Rice – genetically
modified to produce large
amounts of beta carotene, a
provitamin that the body
converts to vitamin A.
• Because traditional rice
varieties do not produce
provitamin-A, transgenic
technologies were required.
1. Rice plants were developed
containing two daffodil genes and one
bacterial gene that carry out the four
steps required for the production of
beta-carotene in rice endosperm.

2. Endosperm is the nutritive tissue

surrounding the embryo of a seed and
makes up the majority of the rice grain
that we eat.

3. The resulting plants appear normal

except that after milling (to remove the
brown bran), their grain is a golden
yellow color due to the presence of
provitamin-A.
How does Golden Rice produced provitamin-A?

• Two genes from daffodil and one from the bacterium Erwinia
uredovora were inserted in the rice genome.
• These three genes produce the enzymes necessary to
convert GGDP to provitamin-A.
• The inserted genes are controlled by specific promoters such
that the enzymes and the provitamin-A are only produced in
the rice endosperm
• When golden rice is ingested, the human body splits the
provitamin-A to make vitamin A.
 Plant
1. Glowing Dendrobium Orchid Flower
- Population of firefly become
less
- transforming the orchid cells
with the firefly luciferase gene
- Young generation can see the
glowing live organism
 Conservation
• Valuable germplasm is becoming extinct
because of environmental degradation and
others.
• Germplasm and plant species must be
conserved.
• Purpose to conserve the germplasm and plant
species is to prevent extinction.
• Cross-breeding, ex-situ conservation and in-
vitro conservation are some options in
preventing extinction.
i) Cross-breeding
• Using ancient germplasm to introduce desirable traits
(such as insects resistance) into modern crops by
crossing susceptible plants with more resistant varieties.
ii) Ex-situ conservation
• Seed storage at low temperatures is the usual ex situ
conservation option, however this is not suitable for
vegetatively propagated plants or species that produce
recalcitrant seeds.
• Field Genebanks provide an important means of ex-
situ conservation but they do risk damage caused by
adverse weather, pest and pathogen attack
iii) In-vitro conservation
• Comprises tissue culture, cryopreservation
techniques and others.
• Cryopreservation is the method of choice for
conserving the germplasm of crops that are
vegetatively propagated in order to maintain
their unique genetic characteristics.
• Cryopreservation is the storage of living cells at
ultra-low temperatures ( −196°C) usually in
liquid nitrogen.
• Safe and cost-effective option for long-term conservation
and increasingly used in the management of crop plant
genetic resources and it is also an important component
of many plant biotechnology programmes.
• Cryogenic storage is currently used to conserve
germplasm derived from wild relatives, ancient and
modern cultivars, and biotechnologically-derived
genotypes.
• Successful temperate crop plant cryoconservation
methods have been enhanced by using cold hardening
pretreatments.
• The potential of plant cryopreservation can only be fully
exploited by effective technology transfer to genebanks
and culture collections.
 KWIAN-THONG (Chao Thui) Organic Fertilizer

 Developed by ASIA AGROTECH Co., Ltd. in Thailand.

 an excellent grade granular type organic fertilizer
 contents major key components to enhance plant growth
 suitable for most crop cultivation and bring hard soil back to its
balance and fertility. carry plenty of essential nutrients and plant
growth regulators to improve plant fertilization and enhance
growth as well as soil amendment.
 It is a special blend of selective raw materials and well suited to
most commercial crops to meet their nutrition level for growth
and lead to healthier crops, better fruit yield, and higher plant
growth
 Nutrient consist in KWIAN-THONG organic fertilizer

• Nitrogen (N) Organic

• Phosphorus (P2O5) Matter(O.M.)
Sulphur(S)
• Manganese (Mn) Hormone
• Potassium (K2O) Boron (B)
EnzymeCopper (Cu)
• Molybdenum (Mo) Humic Acid
• Calcium (Ca) Microbes
Iron (Fe)
• Zinc (Zn)
• Magnesium (Mg)
Process flow for Kwian Tong organic fertilizer
Raw Materials are supply constantly and mixed in silos

liquid plant hormone and enzyme sprayed on the granules.

organic fertilizer kilning process to reduce moisture

sorting process

quality check

packing
 Raw Materials are supply constantly and mixed in silos

Raw material constant flows are

guaranteed by four 60 MT steel type
silos. Raw material will be mixed
according to formula under close
supervision by qualified staffs.

Raw Material Silos

 liquid plant hormone and enzyme sprayed on the granules

• Thoroughly mixed raw

materials will be molded
into granular type organic
fertilizer on giant grilling
dishes. During this stage,
liquid plant hormone and
enzyme will be sprayed on
the granules .

Grilling Disc
organic fertilizer kilning process to reduce moisture

• Bunker oil and firewood are

used by most organic fertilizer
processors during kilning
process to reduce moisture.
heat temperature can be
controlled to meet specific
temperature level by an unique
gas burners so essential micro
organism which are beneficial
to plants and soil are preserved.

Kilning Tube
 sorting process

• Two tiers sorting process,

bigger and smaller size
granule will be sorted off,
guarantees that only
standard size will be passed
through to packing area.
Both under and over size
granules will be filtered out.
Through the process, cracked
and crushed granules will
also be minimal.
Sorter
 Quality check

• Periodic quality check is

another reason why this
products have been proven
as higher than standard.
Periodically before being
packed, Chao-Thui granule
organic fertilizer are
randomly picked and sent to
state university lab to test
their quality and integrity. Granular Organic Fertilizer
 Packing

• Before Chao-Thui organic

fertilizer pellets are to be
packed in bags, they are
manually checked by
packing staffs to make
sure that only high
standard product will be
put in the double layer
bags. KWIAN-THONG Organic Fertilizer
 Benefits by using Kwian-Thong organic fertilizer
• Reproduction of soil microbes stimulated by Kwian-Thong transforms soil structure
from fungus type to bacterial. This process results in minimization of plant
diseases. Hence, plant growth rate and nutrient absorption system are maximized.

• Kwian-Thong help decomposition of organic matters from remnants of farm

manure as well as compost then transform them to nutrients. Kwian-Thong initiates
better soil crumbing structure, water absorption, air ventilation, and prevents soil
acidifies.

• Hormone and enzyme from using Kwian-Thong stimulate plants' growth and
improve their nutrition absorption system.

• Natural growth stimulant by using Kwian-Thong organic fertilizer promotes further

plant growth and strength.

• Increase produce, more and healthier fruits, reduce flagging of fruits and flowers.

• Accelerate blossoming and fruiting.

• Increase of produces' quality, higher weight, better taste, more vivid color.

• Reduce needs of chemical fertilizer utilization and so lower production cost.

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 6.3. Marine Biotechnology
1. What are the aquatic living things
that benefits human?
Creatures of water: Aquatic
animals, plants, corals, protists and
lower organisms.
2. Need manipulate the aquatic
resources for the source of foods
and wealth.
3. Need for aquatic life conservation
for ecosystem balance
 Aquatics for food sources

1. Mariculture (sea water) or aquaculture (fresh

/terrestrial)
a. Marine life breeding (micropropagation)
b. Transgenic marine life (fish, crustaceans etc)
c. Algal products
 Basic Knowledge to aquaculture
• Chemical composition of soil where pond is
situated
• Amount & type of water used
• Type & quantity of animal feed – to achieve
maximum production
HOWEVER
Impose pollution by waste – destroying the
delicate habitat
-Eg: mangroves to make way for shrimp
pond
 Established Achievements of Marine
Biotechnology
• Increase yield and quality of finfish,
crustaceans, algae and various bivalves
– Example:
(i) Japanese maniculture : Produce transgenic
organisms which contributes to 92% of Japan’s total
aquaculture yield
a. Japanese oyster, Red Algae called Nori - largest seaweed
harvest
(ii) Improved hatchery methods : Increase production of
threatened and endangered species, thus enhancing
marine fisheries for both recreation and commercial
use
 Cross application of aquaculture and
agriculture
• Pond humus can be used to fertilize fields
• Livestock waste can serve as pond fertilizer –
stimulate growth of plankton
• Crop by-products used as fish feed
• Anaerobic digester used to produce methane
gas from aquaculture and agriculture waste
 EXAMPLES OF AQUACROPS

Alligator Flesh used for food and skin used for

leather
Bullfrog Cultured for food and laboratory use

Goldfish Popular ornamental fish

Watercress Grown in temperate and subtropical

climates, such as Hawaii
 Category Examples Biotechnology Applications

Gastropod Abalone Add hydrogen peroxide to seawater to

induce synthesis of protaglandin, triggers
spawning
Bivalves Oysters, clams BEFORE : Normal diploid oysters easily
lose flavor because massive amount of
reproductive tissue are formed
AFTER: Triploid Pacific oyster- Obtained
by treating eggs with cytochalasin B which
doubles amount of chromosome during cell
division
-do not form reproductive tissues, therefore
more flavorful and meatier
 Gastropode, bivalve and crustaceans
production
 Genetic manipulation is to increase quality and quantity of production.
 Faster growth
 Increased disease resistance
 Triploidy(3 sets of homologous chromosomes)
 Eg: GE in oyster.
 Diploid oyster spawn in summer and lose their flavour because they form a massive
reproductive tissue.
 Pacific oyster is triploid (2 from female and 1 from male).

Diploid oyster Triploid oyster eggs

eggs Cytochalasin B
( a inhibitor of normal
cell division doubles
the chromosomes
 When eggs are fertilised with normal sperm, zygotes hsve 3 sets of chromosomes
 Triploid oysters are sterile and do not form reproductive organs, more flavourful and
meatier all year round, larger and grow faster.
 Abalone treated with hydrogen peroxide to induce sysnthesis of prostaglandin, a hormone
that trigger spawning.
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 TRANSGENIC FISH
• 1st recombinant DNA application-increase growth & weight
of cultured finfish.
Growth hormone (GH) isolate & inject
Rainbow trout E.coli Young trout
• Injecting fish-labor intensive & poses technical problems
• Fish- excellent for gene transfer (eggs are large &
transparent visible)
• Gene transfer method:
a) Microinjection into nucleus of fertilized egg- method of
choice
b) Electroporation
c) Injection through microphyle (opening in the egg where
sperm enters)

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d) Direct pronuclear injection of unfertilized eggs (must be
isolated, cultured & fertilized in vitro)
• Antifreeze protein can extend fish range of survival into colder
water.
• For salmonid eggs (with hard chorion):
a) injection through the microphyle with very fine glass needles
(2-3 μm in OD)inserts the transgene very near the male
pronucleus (just under microphyle)
b) use sturdy glass needles (≤15 μm) to penetrate the chorion &
insert the DNA into the blastodisc just after fertilization but
before the chorion completely hardens.
c) cut a minute hole in the egg after the chorion hardens for
insertion of a glass needle into the blastodisc just before the 1st
cell division.
Sources of Energy and Medicine and
Conservation
Medicine Marine Environmental
– Algal cell Culture control
– Vaccine -Probing the Marine
– Antidotes Environment
Aquatic life
Fuel conservation
– Fuels from algae Breeding of
endangered species
 Algal Products
• Photosynthetic eukaryotes, 2 types (microalgae and
macroalgae). 3 group of macroalgae: green, red and
browm.
• Collected for food and medicine
• Harvested from wild and produced in culture (Japan,
China, Korea, USA, Canada, Chile UK, Indonesia,
Philippine).
• Kelp (Macrocystis) for fertiliser and source of potash
and acetone. (USA)
• Kelco-company producing algin products as feeds and
food supplements. (Table 8.2,8.3)
• Nori;red algae, Wakame:brown algae, Kombu/Japan,
 Spirulina
Spirulina-Cynobacterium
Spirulina farms:
Lake Texcoco in Mexico,
Lake Chad in West Africa.
70% protein dry weight
Largest in the world-Lake Texcoco (dry lake of Aztec
civilisation). Fig 8.6.
GM by mutation and selection.
GM for faster growth, disease resistant, increase the
synthesis of metabolites, new and unique products from
marine.
 Fuel from algae

• A potential alternative to fossil fuel.

• Bioconversion:
Biomass methane

Catalytic hydrogenation
Thermal gasification
Biological gasification (microbial conversion)

• Research stage
• Expensive technology
 Step 1: Acid forming
Products:
Complex CO2, H2, H2S,
BIOMASS macromolecules
NH3, acids
•Source of Non-methanogens
Bacteria: bacteria
–Agricultural
wastes
Methanogens
–High saline bacteria
environment
s
Step 2: Methane
–Estuaries producing
Methane
CO2
 Medical application of marine products

Marine animals as models to study human diseases

Sharks do not form cancerous cells
Icefish lacks of red blood and hemoglobin use to study anemia
Marine natural products and their medical potential
1. Researchers screen for marine natural products
2. >20,000 chemicals have been characterised.
3. Example of chemicals from marine natural products
Dedemnin B (effective against leukemia, melanoma)
Dolastatin 10 (antitumor activity)
Luffarialin A (anti-inflammatory, analgestic, and anti-fungal)
Halicondrins etc.
 Anticancer & Antiviral

• Studies marine extracts that show promise as

anticancer drugs.
• Dimentin B from Caribbean tunicate effective
against leukemia.
• Bryozoans or false corals effective for cancers
such as ovarian carcinima.
• Antiviral agent such as eudistomins extracted
from tunicated.
 Marine toxins

• Marines organism used it as predation, and

predators.
• Extremely toxics and may cause dead to
human.
• Studies for anti tumor – extracts from sea
hares (mollusk) for lymphocytic leukimia &
melanoma.
 Algal culture for medicine and other uses
To generate biomass for rare
metabolites isolation.
• Use large pond or raceway (Fig:
8.7) or in bioreactors.
• Eg: proline (aa) extracted from
Chlorella cell culture,
octopamine gene transferred
from bacteria to green algae.
• Expensive technique for
specialty chemicals
(dihydroxyacetone, gluconic
acid, acetic acid, medical
phycobiliprotein
 Marine Animal Health
• Marine animal are susceptible to protozoan,
bacterial, viral, fungal and marine pollution that
promote pathogens (fungus:Saprolegia,
bacterial:Vibrio).
• To cultivate disease-resistant marine animals
(vaccination or GM marine animals)
• Marine bacterial control to date-antibacterial agents
such as disinfectants and antibiotics (tetracyclin,
chloramphenicol, penicillin etc)
• Salmon commonly infected with viruses causing
hematopoitic necrosis and pancreatic necrosis-
vaccine underway.
 Marine Animal Health

• Marine pollution promotes patogens that

infect marine organisms
• Important incidents:
– (I)Tiger Shrimp – Virus that decimated shrimps
causing tremendous drop in shrimp production
• No vaccines to protect marine lives
– (ii)Salmon- threatened by two deadly viral diseases:
• Infectous Hematopoietic necrosis
• Infectous Pancreatic Necrosis
 Probing The Marine
Environment/Conservation

 To conserve and protect Recent Research on

the marine environment
 Use of PCR, DNA probes,
Vaccine for Hematopoietic
create diagnostic kits for necrosis:
pathogens, contaminants, A recombinant vaccine-produced by
pollutants etc isolating and expressing genes encoding
 Increase understanding of viral proteins, induces formation of
abundance marine antibodies within fish
products
 Screening of foods using
protected endangered
species