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Role of tissue culture for elimination of viral disease
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By Allah Jurio Khaskheli
Issues/Analysis M.Sc. Agri. (Hons.)
Weather Department of Biotechnology
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Interactive Almighty Allah has created human being as supreme to all other livings,
Ashraf-ul-Makhlukat. He has bestowed human being with great
Pak APIN capabilities. The man has made immense progresses in biological and
Feed Back physical science, due to this supreme quality.
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Tissue culture is the most effective and important discovery of human
Site Info being in the field / world of biological sciences. It is an establish fact
Search that plant provides about 80% requirement of our daily life. It is also fact
Ads that man and animals also depend on plants. Unfortunately, nowadays,
our crop plants are highly affected by many dangerous and
Pakissan Panel 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

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4/5/2019 Pakissan.com; Role of tissue culture for elimination of viral disease
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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