Role of tissue
culture for elimination of viral disease
By Allah Jurio Khaskheli
M.Sc. Agri. (Hons.)
Department of Biotechnology
Sindh Agriculture University Tandojam
 Almighty
Allah has created human being as supreme to all other
livings, Ashraf-ul-Makhlukat. He has bestowed human
being with great capabilities. The man has made immense
progresses in biological and physical science, due to
this supreme quality.
Tissue culture is the most effective and important
discovery of human being in the field / world of
biological sciences. It is an establish fact that plant
provides about 80% requirement of our daily life. It is
also fact that man and animals also depend on plants.
Unfortunately, nowadays, our crop plants are highly
affected by many dangerous and damageable diseases such
as bacterial, viral, and fungal diseases. The most
virulent diseases, which cause great losses in our crop
plants, are the viral diseases. Unfortunately, there is
no any chemical control against viral diseases, to
eliminate the virus from our crops; but in the light if
modern world, the method of tissue culture is going to
solve this problem.
VIRAL PLANT DISEASES
Viral plant diseases are very common, which causes great
economic losses in our cultivated crops, in the form of
reduction in quantity and quality of yield, all over the
world. The plants, which are infected by viral diseases,
show many symptoms, which depend on the types of disease
attack. Some of these symptoms are leaf distortion e.g.
Curling of leaf, leaf yellowing (either of the yellowing
of whole leaf or in a pattern of strips or blotches), or
other growing distortions, which include the stunting of
whole plant, and abnormalities in flower or fruit
formation.
The most of the viruses are generally inhabits the
process of photosynthesis, which results in blank of
chlorophyll in the plants and as a result, the plants
become disable to prepare their food. The example of
common viral diseases, which causes tremendous losses in
our crop plants are; CLCV (cotton leaf curl virus),
sugarcane leaf virus, BBTV (banana bunchy top virus),
TMC (tobacco mosaic virus) etc. The most of the growers,
even general scientists/experts, just gave an idea that
viral dieses can be controlled by controlling the viral
vectors, through restricting the spread of an organism,
from infected one to another host. The few examples, of
the generally viral vectors are insect pests of plants,
which are fed on different plant parts, such as aphid,
white fly etc.
Now, in these days, in the modern world, the method of
tissue culture is considered as the most effective
method for the control of viral diseases. The plantlets
are grown in the Plant Tissue Culture Laboratories (PTCL),
for the production of disease free plants through tissue
culture techniques, under intensive care and control
conditions. The baby plants are nourished with
artificial medium, containing all the nutrients required
for the growth of plants, in laboratory condition.
ARTIFICIAL NUTRIENT MEDIUM
Food is the basic requirement of every organism.
Similarly, the first thing for the elimination of viral
diseases from the plants through tissue culture is to
provide proper food, which is called medium, an
artificial food. It is prepared in the laboratory.
Mostly MS medium is used in
tissue culture laboratory. In 1962, two scientists,
Murashige and Skooge used first time MS medium, contains
all the necessary components, which are needed for the
growth of plants in vitro conditions (in laboratory
conditions) i.e. micronutrient elements, macro nutrients
elements, vitamins, growth regulators, glucose, proteins
and also the agar (a type of sea weed for solidifying
the media). Carbon is the structural backbone of the
organic compounds in the make up the living cells. The
carbon source includes glucose, sucrose and lactose. The
nitrogen is also basic need, which increases ratio of
chlorophyll pigments in the plants, which is essential
for the processes of photosynthesis nitrogen. It is
needed for the synthesis of such molecules e.g. amino
acid, DNA, RNA and ATP. The trace elements like zinc,
iron ,copper ,manganese ,etc are also required in very
minute amounts, for the proper growth of plants. The
artificial nutrient medium can also be changed according
to the nature of plants.
PROCEDURE OF TISSUE CULTURE TO PRODUCE VIRUS FREE
SEEDLINGS
First of all, healthy and young portion of the target
plants are collected from the field. The collected
portion should be free from virus. Amputated healthy
portion of the plant is called ex-plant. It is washed
thoroughly to remove dust particles and disease
organisms. After washing, the ex-plant is sterilized
with 70% sodium hypo chloride or normal bleach solution
for 20 to 25 minutes. Sterilized exp-plant and all other
necessary equipments /materials are transferred in
laminar air flow cabinet.
Under laminar air flow
cabinet, again wash and sterilize the ex-plant with
double distilled and sterilized water, to remove sodium
hypo chloride from the plant, to avoid the chance of
contamination (attack of microorganism). The required
portion of exp-plant is cut (i.e. growing part) with the
help of sterilized knife and scissor. The cutting
portion of explants is transferred in to MS medium,
which is already prepared in the tissue culture
laboratory. After the process of culturing, the culture
bottle is transferred into the growth room, where the
temperature, humidity and light are already under
control, artificially. The plants are kept in dark room
for 3 to 4 weeks until the process of differentiation
start. In the process of differentiation, the roots and
shoots are formed. After shooting, the newly plants are
sub-cultured. Sub-culture is process, in which cultured
plants are again cultured in a new medium under aseptic
condition under laminar air flow cabinet, near about
similar to as described for the first culture
preparation. The process of sub-culturing is repeated 3
to 5 time for complete elimination of virus from plants,
after which, the plant are transferred into the glass
house. This process of transferring plants in to glass
houses is called hardening and weaning process. This
process is very necessary for the survival of tissue
cultured plants, because, when the plants are coming
from tissue culture laboratory, they are more sensitive
to natural light, humidity and temperature. Therefore,
the process of hardening and weaning makes these plants
stable/ string, to survive in natural conditions.
Finally, these plants are transferred from green house
to the field.
The plants required 3 to 6 month for development into
complete virus free plants, through tissue culture
techniques.
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