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Introduction:
Agriculture plays a significant role in the national economy of
Egypt, so it receives high priority from government. Agriculture
accounts for 20% of gross domestic product (GDP) and total
exports and 34% of the total labor force. The agriculture sector
contributes to the overall food needs of the country and
provides the domestic industry with agriculture raw materials.
The agriculture sector has taken major steps to reform its
economic policy program, such as the following:
Gradually removing government controls on farm output prices.
Increasing farm-gate prices to cope with international prices.
Removing farm input subsidies.
Removing government constraints on the private sector on
importing and exporting agriculture crops.
Imposing limitations on state ownership of land and sale of
new
land to the private sector.
Adjusting the land tenancy system.
Confining the role of the Ministry of Agriculture (MOA) to
agriculture research, extension and economic policies.
As the government moves towards privatization, transfer of
technology to the private sector is also occurring (for example,
in vitro micropropagation of virus-free potato). This
shows the capacity and interest of the private sector in
adopting new technology. Technology transfer is expected to grow
dramatically in the short term as the research program become
more product-orientated.
One of the major targets for biotechnology in Egypt is the
production of transgenic plants conferring resistance to biotic
stresses (resulting from pathogenic viruses, bacteria, fungi and
insect pests) and abiotic stresses, such as salinity, drought
and high temperature. These biotic and abiotic constraints are
major agriculture problems, leading to serious yield losses in
many economically important crops in Egypt.
The agriculture sector has the following strategic goals:
Optimizing crop returns per unit of land and water consumed.
Enhancing sustainability of resource use patterns and
protection of the environment.
Bridging the food gap and achieving self-reliance.
Expanding foreign-exchange earnings from agriculture exports.
Some of the opportunities for deploying modern biotechnology
approaches include:
Producing transgenic plants resistance to indigenous biotic
and abiotic stress.
Reducing the use of agrochemicals and pesticides and their
environmental risks.
Improving the nutritional quality of food crops.
Reducing the dependence on imported agricultural products
(seeds, crops).
The private sector has access to biotechnology and has invested
heavily in research and development (R& D) of technology and the
necessary expertise to bring a product to market. The
competitive edge of a private company depends on the proprietary
nature of its R & D and the protection offered by intellectual
property laws. A private company might engage in development of
a product in conjunction with a developing country because:(i)
it addresses a technical problem critical to its own product
development,(ii) it presents an opportunity to enhance its
public relations, and /or (iii) it provides a window on an
important market, technology or germ-plasm of interest.
Developing-country institutions may be interested in working
with private companies to gain access to important technology,
develop managerial and business expertise and build intellectual
capability of bringing a product to market.
Biotechnology applications in Egypt, From different Universities
& Institutes
Use of genetic engineering for producing
transgenic squash
plants resistant to ZYMV
The cultivated cucurbits can be completely threatened when
infected by ZYMV, CMV and/or WMV, which causing reduction in
their productivity. The squash cultivated area, around 78,000
feddans and producing about 568,000 tons, in Egypt was found to
be affected by ZYMV. Zucchini yellow mosaic potyvirus (ZYMV), is
considered to be the most important virus threat the cucurbit
crops (e.g., squash, watermelon, melon and cucumber) in Egypt.
The virus was firstly recorded as a series virus affecting
squash, muskmelon (Cucumis melon L.) and other cucurbits.
The control of ZYMV based on using insecticides, mineral oil,
and/or inspection and rouging was ineffective. The control of
such viruses based on using insecticides and/or inspection and
rouging was ineffective. Developing transgenic plants resistant
to RNA viruses, e.g. ZYMV affecting cucurbits via genetic
engineering approach based on the coat protein-mediated
protection would be the possible future options for control of
ZYMV. In cucurbits, transgenic squash with resistance to two and
three viruses have been obtained in USA. Therefore, the research
team at AGERI in collaboration with Horticultural Research
Institute (HRI), ARC produced transgenic squash plants tolerant
to ZYMV via the following steps:
1.
Establishment and optimization of regeneration and
transformation systems in a local Eskandarani squash cultivar.
2.
Isolation, cloning and sequencing of cp gene of ZYMV-Eg
isolate.
3.
Construction of transformation vector (plasmid pGA643)
containing gene of interest.
4.
Introducing the ZYMV-Eg-cp gene into the Egyptian
Eskandarani cultivar of squash, via Agrobacterium-mediated
gene transfer system.
5.
Serological (ELISA) and molecular (PCR and Southern blot
hybridization) analysis of transgenic plants for detecting the
expression and presence of the ZYMV-Rg-cp gene,
respectively.
6.
Evaluation of highly ZYMV-tolerant squash lines containing the
cp-genes under a controlled greenhouse conditions (three
generations). It worth to mention that the transgenic as well as
non-transgenic plants were subjected to twice artificial
mechanical inoculation with sap prepared from ZYMV-infected
tissues.
7.
Evaluation of highly ZYMV-tolerant squash lines containing the
cp-genes under open field conditions (two generations).
Results could be summarized as a delay in developing of the
characteristic symptoms of ZYMV was found up to 10-13 weeks post
cultivation. In addition, the highest percentage of virus
infection was ranged from 4-8 and 60-80% in case of transgenic
and non-transgenic squash plants, respectively. The
horticultural characters, i.e., fruits quality and their
marketable degrees were also determined.
** Agricultural Genetic engineering Research Institute (AGERI)
www.ageri.sci.eg
Development of
transgenic rice resistant
to stem borers and fungi
Faculty of Agriculture, Cairo University, is the oldest in Egypt
among Faculties of Agriculture, since it was founded in 1890.
The faculty provides staff and students with the most advanced
farm and laboratory facilities to enhance the quality of
teaching and research. Genetic Engineering Center (GEC) is a
well equipped research center built inside the faculty to
provide the opportunity for performing advanced biotechnology
researches. GEC had a number of ongoing research programs in
that field, among them is the development of transgenic rice
resistant to stem borers and fungi.
Rice is a major field crop and a major source of calories for
the Egyptians. It is the second cash-export crop after cotton.
Rice production is affected by both blast and borers. Efforts to
control the insects and disease based on the use of existing
resistant varieties have only been partially successful as the
resistance rapidly breaks down under heavy fungal and insect
infection. Development of a novel strategy for broad-spectrum
and durable control of blast and borers is needed. Genetic
Engineering Center is working on introducing the plant
chitinase, cholesterol oxidase and Bt genes into rice to
engineer quantitative resistance to the blast fungus and stem
borers in a non-race specific manner. To achieve this goal, they
developed and optimized reproducible protocols for Egyptian rice
tissue culture and gene delivery (Agrobacterium and
Biolistic). Extensive trails are going on for improvement of
rice regeneration, focusing onto some modification and changes
in the type and level of growth regulators and removal of
selective pressure during regeneration. Nowadays they have
putative transformed rice cultures harboring genes of interest
(i.e. Bt., cholesterol oxidase and chitinase) and the selectable
marker genes GUS and/or kanamycin resistance. The results were
confirmed using both bioassay and PCR. The work is running on at
two parallel topics, the first one involves the enhancement of
the rate of shoot recovery following transformation, while the
second topic involves the transformation and evaluation of
transformed rice plants. Also an addition research task was
performed, dealing with AFLP fingerprinting of different
Egyptian rice varieties. The obtained data will be used for
differentiating between varieties and link the obtained data to
economically important traits.
Development of
wheat plants Tolerant to drought stress through
genetic engineering
This collaborative project between Fac. of Agriculture, Ain
Shams University and AGERI, ARC aims to develop transgenic wheat
Tolerant to abiotic stress by transferring the bacterial
Fructan-accumulating gene (sacB) into wheat.
Lab Testing:
sacB / bar genes have been introduced into the Egyptian wheat
cultivar G164 using the gene gun. The bar gene for herbicide
(baste) resistant was used as a selectable marker gene. Putative
transgenic herbicide-resistant wheat plants were selected for
subsequent evaluation. Molecular analysis indicated the presence
and expression of the transgene using PCR, Southern and RT-PCR
for the (T0) and (T1) plants.
Greenhouse Testing:
This test was done to detect the drought Tolerance level of the
transgenic at the (T1) generation compared with that of the non
transgenic plants. Control plants were watered with 140 cm/pot,
while those under drought stress received 40% this amount. No
significant differences were observed between well-watered and
drought-stressed transgenic plants.
According to the successful results of the greenhouse, field
trails is taking place to test these transgenic plants.
Biosafety measures will evaluate the transgenic materials to be
cultivated in areas where amount of water available for
irrigation is limited.
Development of transgenic wheat
with improved tolerance to drought stress
Agricultural Genetic Engineering Research Institute (AGERI),
ARC, has developed a transgenic wheat tolerant to drought
stress.
Lab and greenhouse testing:
HVA1 gene, isolated from barley has been introduced into
cultivated bread wheat using gene gun to improve its tolerance
to drought stress. The gene encode one of the LEA (late
embryogenesis abundant) proteins that confer drought tolerance
to embryo cells at maturity stage during seed desiccation. The
herbicide resistance gene bar gene has been used as a
selectable marker gene during the course of transformation and
tissue culture and to test homogeny of the original transgene at
subsequent generations. Presence and expression of the transgene
have been proven in ten different transgenic events. Drought
stress experiments have been conducted in the greenhouse events
for subsequent field testing.
Field testing:
Six drought stress experiments have been conducted in three
consecutive seasons to compare the performances of transgenic
versus non transgenic plant. At the first two seasons, drought
stressed plants were irrigated only once one month after
germination, while eight times for control non-stressed plants.
At the third season, rain fed experiments at four locations,
differing in the amount of water received during the season,
were conducted.
Results at the first two seasons indicate no significant
differences in grain yield and another yield attributes between
stressed and non stressed transgenic plants. At the third
season, one of the transgenic events significantly differed from
the non transgenic plants in biological and grain yields.
Transgene is now introduced into a number of Egyptian wheat
cultivators (ex., Giza 164, G168, Seds 1, etc.) through
conventional breeding. Food safety issues will be handled by the
National Biosafety Committee that gives the approval for
genetically modified product to be commercialized in Egypt.
** Agricultural Genetic engineering Research Institute (AGERI)
www.ageri.sci.eg
Production of Transgenic
banana plants
resistant to some viruses
Banana is one of the major and economically important fruit
crops that play important roles in local diets and as export
crops in some African countries as well as in Egypt. In Egypt,
the cultivated banana areas (49294 Faddens, and producing 849293
tons) are infected with some common viruses, i.e., banana bunchy
top nanovirus (BBTV), banana-cucumber mosaic cucumovirus
(Banana-cMV), and banana bract mosaic potyvirus (BBrMV). BBTV
and Banana-CMV are the most serious viruses affecting banana,
causing reduction in yield and quality. The control of such
viruses using insecticides and/or inspection and rouging is
ineffective. Therefore, by production of transgenic banana
plants resistant to such viruses would be the most effective
means for its control beside the classical means.
Therefore, through a collaborative project between AGERI and
Faculty of Agricultural, Alexandria University, transgenic
banana plants expressing the coat protein (cp) genes of
BBTV and Banana-CMV were produced.
The steps which carried out could be summarized as
follows:
a)
Main goals:
Producing transgenic banana plants containing the cp
genes of BBTV and Banana-CMV the causal agents of banana bunchy
top disease (BBTD) and banana mosaic disease (BMD),
respectively, using the genetic engineering and tissue culture
approaches.
b)
Samples collection:
Seedlings of banana cv. Williams naturally infected with BBTV
and showing the typical symptoms of BBTD or BMD were collected
from a private farm in El-Kalubia governorate, Egypt. The
collected samples were tested for the presence of BBTV or
Banana-CMV using DAS-ELISA technique using an ELISA kit from
Sanofi, France.
c)
Gene isolation and construction:
The cp genes of BBTV and Banana-CMV under investigation
were isolated via PCR and RT-PCR technologies,
respectively. These genes were separately cloned into the pGEM-T
Easy vector and introduced in E. coli strain JM 109 strain. The
DNA plasmids were miniprepared and the nucleotide sequence of
the inserts were determined. After confirmation of the
expression of the applied genes into the bacterial cells, the
genes were separately subcloned into a plant expression vector.
d)
Establishment of a transformation system in banana:
In this experiment, the transformation system in cv. Williams
banana cultivar was established using the plasmid pAB6 carrying
the bar and gus genes and microprojectile
bombardment transformation system.
e)
Introducing the cp genes into banana cells:
The genes of interest were separately introduced in banana
plants via microprojectile bombardment transformation
system. On transformation the transformed plant materials were
acclimatized under a control greenhouse conditions.
f)
Genes detection:
The presence and expression of the cp genes were detected
in the transformed plant materials using PCR, ELISA as well as
western blot immunoassay.
g)
Evaluation of transgenic banana plants:
In a contained field trail, the virus resistance of the produced
banana plants is determining in the presence of virus-infected
banana plants, and source of banana-aphid. It is worth to
mention that no insecticides for insect control were used.
Improvement
the nutritional
quality of faba bean
using genetic engineering
The problem:
Seed storage proteins of faba bean constitute the food bases of
the diet of millions in Egypt. Unfortunately, these proteins are
deficient in some essential amino acids e-g. methionine and
cysteine. The deficiency of sulfur containing amino acid limits
its nutritional value. The physical and mental development of
children can be irreversibly retarded by the deficiency of
essential amino acids in their diet. Unfortunately, traditional
breeding programs were unable to solve this problem. Therefore,
improvement of nutritional quality of faba bean by genetic
engineering is a new promise to overcome this dilemma,
especially its proved to be effective in other crops such as
rice.
Achievements:
The gene coding for sulfur rich sunflower albumin (SFA8), under
the control of Vicia faba legumin B4 promoter, which
elicits seed-specific expression, was introduced into faba bean
genome. Transgenic faba bean plants were recovered and the
integration of the gene was confirmed by molecular analysis.
Amino acid analysis of transgenic faba bean seeds indicated the
expression of gene. The transgenic seeds contain methionine at
level of 0.76 % of total crude proteins of the seeds. This level
represents an increase of 15.1% of the total methionine found I
the seed as compared to the wild type. Moreover, the cyctine
level increased by 23% in comparison to the wild type.
Future
outlook:
Nowadays, the teamwork of this research in National Research
Center (NRC) is preparing to submit application for the national
biosafty committee to get its permission to grow the seeds ex
vitro (containment conditions), in order to monitor the
expression of sulfur rich proteins in successive generation.
Collaborators:
This work is carried out in collaboration with Hanover and
Berlin Universities, Germany, through a Ph.D. thesis of Dr.
Moemen S. Hanfy
** National Research Center (NRC)
www.nrc.sci.eg
Expression of hepatitis B antigen
(HBsAg)
in
transgenic
maize
(Zea mays L.)
The Hepatitis B virus (HBV) infection is one of the most
widespread viral infections of humans and causes acute and
chronic hepatitis and hepatocellular carcinoma. The world wide
problem of HBV infection has necessitated the development of an
effective vaccine. In many areas of the developing world, the
cost of immunization programs prohibits the use of the currently
available vaccines for large segments of the population. This
limitation led us to attempt the expression of the recombinant
Hepatits B surface antigen (rHBsAg) in plants with the hope of
developing low-cost production systems and effective delivery
systems for vaccines. In this study, we used Zea mays L. as
biological bioreactor for large scale production of HBsAg.
Immature embryos of maize inbred lines were bombarded using the
biolistic gene gun with the plasmid pBHsAg harboring the gene
encoding the HBsAg and the bar gene as a selectable marker.
Bombarded tissues were selected and regenerated on media
containing 3 mg/l Bialaphos. HBsAg gene was detected using PCR
analysis and the expression of the gene was tested via Western
blot immuno-assay using specific polyclonal antibodies directed
against human serum derived HBsAg. This study demonstrates the
feasibility of using products derived from transgenic corn as
vaccine delivery vehicles.
** Agricultural Genetic engineering Research Institute (AGERI)
www.ageri.sci.eg |
