CHINMAYA VIDYALAYA

PALLAVUR

ACADEMIC YEAR: 2023-24

INVESTIGATORY PROJECT REPORT
Topic: PLANT TISSUE CULTURE AND ITS
APPLICATIONS
ROLL NO : 17
NAME : RESHMI K
CLASS : XII A
SUBJECT : BIOLOGY
SUB CODE : 044

PROJECT GUIDE: SAPNA P

 CHINMAYA VIDYALAYA PALLAVUR

CERTIFICATE
This is to certify that RESHMI K Roll No:17 has successfully
completed the Investigatory Project work entitled PLANT TISSUE
CULTURE AND ITS APPLICATIONS in the subject BIOLOGY-
044 for the academic year 2023-24, laid down in the regulations of
CBSE for the purpose of Practical Examination in Class XII
held in Chinmaya Vidyalaya Pallavur.

Signature of INTERNAL EXAMINER:

Signature of PRINCIPAL:
Signature of EXTERNAL EXAMINER:

ACKNOWLEDGEMENT

I would specially thank Mrs SAPNA P my BIOLOGY teacher

for providing her valuable guidance, support, suggestions,
and comments throughout the course of project.

I would also like to express my sincere gratitude to our

Principal Mrs. SHINI T S for providing an opportunity to
work on the Investigatory Project titled PLANT TISSUE
CULTURE AND ITS APPLICATIONS. The completion of
the project would not have been possible without their help
and insights.

I would like to extend my appreciation to all, especially my

group members who contributed their cooperation and inputs
to the success of this project.

Signature:
Name: RESHMI K
ROLL NO: 17

PLANT TISSUE CULTURE

AND ITS APPLICATIONS
INDEX
TABLE OF CONTENTS

SER DESCRIPTION PAGE NO

01 Introduction 1
 02 Background of plant tissue culture 2

03 Stages of tissue culture process 3

04 Processes of plant tissue culture 5

05 Types of plant tissue culture 6

Commercially propagated plants through micro-

06 7
propagation in India

Why is plant tissue culture better than other conventional

07 7
methods?

08 Advantages of micro-propagation technology 8

09 Applications 9

10 Risks of plant tissue culture 9

11 Environmental conditions 10

12 Conclusion 11

13 Bibliography 12

INTRODUCTION
A whole plant can be regenerated from a small tissue or plant cells in
a suitable culture medium under controlled environment. The plantlets
so produced are called tissue-culture raised plants. These plantlets are
a true copy of the mother plant and show characteristics identical to
the mother plant. For example, if the mother plant is a high yielding
plant the plantlets will also be high yielding. Many plant species are
presently being propagated through tissue culture successfully.
This capacity of a single cell to grow into a complete plant is termed
as Totipotency, which was first put forward by a German Botanist
Haberlandt in 1902. Tissue culture is the propagation of plants
wherein a part/tissue of the plant is placed in nutrient media that
favours the production of shoots, roots following which they are
hardened and transferred to soil. Quality planting material of
economically important species can be produced in a large
scale/desired quantity through tissue culture.
Tissue culture is the term used for “the process of growing cells
artificially in the laboratory”. Plant tissue culture can be initiated from
almost any part of a plant however, for micropropagation or direct
shoot regeneration, meristematic tissue such as shoot tip is ideal. The
physiological state of the plant does have an influence on its response
to tissue culture. The mother plant must be healthy and free from
obvious signs of disease or pest. The shoot tip explants being juvenile
contain a higher proportion of actively dividing cells. It is important
to use quality mother plant stock to initiate cultures.
The cultural conditions required to initiate and sustain plant cells in
culture, or to regenerate intact plants from cultured cells, are different
for each plant species. Each variety or clone of a species often have a
particular set of cultural requirements.

BACKGROUND OF PLANT TISSUE CULTURE

 Plant tissue culture, or the aseptic culture of cells, tissues,
organs, and their components under defined physical and
chemical conditions in vitro, is an important tool in both basic
 and applied studies as well as in commercial application. It owes
its origin to the ideas of the German scientist, Haberlandt, at the
beginning of the 20th century.
 The early studies led to root cultures, embryo cultures, and the
first true callus/tissue cultures. The period between the 1940s
and the 1960s was marked by the development of new
techniques and the improvement of those that were already in
use. It was the availability of these techniques that led to the
application of tissue culture to five broad areas, namely, cell
behaviour (including cytology, nutrition, metabolism,
morphogenesis, embryogenesis, and pathology), plant
modification and improvement, pathogen-free plants and
germplasm storage, clonal propagation, and product (mainly
secondary metabolite) formation, starting in the mid-1960s.
 The 1990s saw continued expansion in the application of the In
vitro technologies to an increasing number of plant species. Cell
cultures have remained an important tool in the study of basic
areas of plant biology and biochemistry and have assumed
major significance in studies in molecular biology and
agricultural biotechnology.

STAGES OF TISSUE CULTURE PROCESS

1. Preparation of nutrient medium
A semi- sealed solid medium is prepared in double distilled water
containing macro elements, micro elements, amino acids, vitamins,
iron source, carbon source like sucrose and Phyto-hormones. The
medium is heated for dissolving the agar and 25 to 50 ml is dispensed
into each wide mouth bottles. The vessels containing culture media
are then and sterilized by autoclaving.
2. Inoculation
Inoculation is carried out under aseptic conditions. In this process
explants or micro shoots are transferred on to the sterilized
nutrient medium.
3. Development of plants in growth room
After the inoculation of the plant tissue, the bottles are sealed and
transferred into growth room to trigger developmental process under
diffused light (fluorescent light of 1000-2000 lux) at 25 ± 2oC and 50
to 60% relative humidity. The cultures are observed daily for growth
and any signs of infection/ contamination. Cultures, that do not show
good growth or infected, are discarded. Healthy cultures develop into
small shoot buds, which are sub-cultured on fresh medium after 4
weeks. The number of subcultures depends on the plant species and is
standardized. After 4 weeks, shoots develop and are transferred to
rooting mediums. Roots form within 2 to 4 weeks, and plants require
careful handling due to their delicate nature.
4.Hardening of micro plants
High humidity and artificial conditions in the culture vessel cause
plantlets to be tender. After being washed and maintained under mist

or transparent plastic, they are transferred to a greenhouse for another

4 to 6 weeks before being ready for transfer to the net house or field.
Tissue culture plants are typically sold as ex-agar or hardened plants.
  Ex-agar plants
Ex-agar plants for sale can be in vitro rooted or shoots and are washed
in sterilized water to remove the agar medium. Sorted into 2-3 grades,
packed in corrugated plastic boxes, and treated with specific
fungicides and antibiotics for export. These plants are preferred for
export or hardening facilities and should be transplanted within 72
hours after being removed from nutrient media. The number of plants
per box depends on the customer's requirements.
 Hardened plants
Plants are acclimatized in net pots after developing shoots and roots in
bottles. Rooted plantlets are then placed in pots filled with substrate
and watered. After 4 to 6 weeks in a greenhouse, plants are treated
with fertilizers and acclimatized. After acclimatization, plants are
transferred to polybags, which are hardened and ready for field
cultivation. Hardening units can be set up in different locations.
PROCESSES OF PLANT TISSUE CULTURE
TYPES OF PLANT TISSUE CULTURE
Seed Culture
In this culture, the explants are obtained from an in-vitro derived
plant and introduced into a laboratory where they proliferate.
The explant should be sterilized to prevent it from tissue
damage.
Embryo Culture
This involves the in-vitro development of an embryo. For this,
an embryo is isolated from a living organism. Both, a mature or
an immature embryo can be used in the process. Mature
embryos can be obtained from ripe seeds. The immature
embryos are obtained from the seeds that failed to germinate.
The ovule, seed or fruit is already sterilized therefore, it does not
need to be sterilized again.
Callus Culture
A callus is an unorganized, dividing mass of cells. When the
explants are cultured in a proper medium, the callus is obtained.
The growth of callus is followed by organ differentiation. The
culture is grown on a gel-like medium composed of agar and
specific nutrients required for the growth of the cells.
Organ Culture
In this, any organ of the plant such as shoot, leaf, can be used as
an explant. Several methods can be used for the organ culture,
such as plasma clot method, raft method, grid method, and agar
gel method. This method is used to preserve the structure and
functions of an organism.
Protoplast Culture
It is a cell without a cell wall. A protoplast can be cultured using
the hanging-drop method, or micro-culture chambers. In
protoplast culture, several phases can be observed: development
of cell wall, cell division, regeneration of a whole plant.
COMMERCIALLY PROPOGATED PLANTS
THROUGH MICRO-PROPOGATION IN INDIA
Plant category Plants
Fruits Banana, Pineapple, Strawberry
Cash crops Sugarcane, Potato
Spices Turmeric, Ginger, Vanilla,
Cardamom
Medicinal plants Aloe vera, Neem, Geranium,
Stevia, Patchouli
Ornamentals Gerbera, Carnation, Lily,
Syngonium, Anthurium,
Cymbidium
Woody plants Teak, Bamboo, Eucalyptus
Populus
Biofuel Jatropha, Pongamia

WHY IS PLANT TISSUE CULTURE BETTER

THAN OTHER CONVENTIONAL METHODS?
Tissue culture is a method that produces thousands of exact copies of
plants, unlike conventional breeding which requires seeds or
propagative parts. This process is faster and requires a small explant,
making it more efficient. The offspring are clones of parent plants,
making pure line production less time-consuming and requiring less
than 4 to 5 generations.
ADVANTAGES OF MICRO-PROPAGATION
TECHNOLOGY
Micro-propagation has several advantages over conventional methods
of propagation such as:

 Rapid multiplication
Micro-propagation offers rapid multiplication of desired plant species.

 Requirement of only limited number of explants

Small pieces of plant (explants)/tissue can be used to produce many
plants in a relatively.
small space.

 Uniform or true to type plants.

Micro-propagation offers high uniformity in phenotypic/physical
traits due to controlled production, allowing for market demand
planning and scheduling, compared to traditional propagated plants.

 Germplasm storage
Plants can be stored in vitro in a small space and less labour is
required for maintenance of stock plants.

 Disease free planting material

Tissue culture produces disease-free plantlets, which can be
eliminated through proper diagnosis and treatments, and indexed
using serological and molecular techniques for mass multiplication.

 Growth manipulation
Nutrient levels, light, temperature, and other factors can be more
effectively controlled to manipulate the growth.
APPLICATIONS
•Micropropagation
•Somatic embryogenesis
•Application of plant biotechnology
•Genetics
•Haploid development via tissue culture
•Plant transformation
•Endophytes and secondary metabolite

RISKS OF PLANT TISSUE CULTURE

The utilization of plant tissue culture for commercial production is
limited by two major risks viz., spread of diseases especially those
caused by viruses, and variations. The movement of plants also
involves accidental risk of introducing plant disease. Pathogens that
are often symptom less, such as viruses, pose a risk. The risk of
distribution of inferior micro propagated plants has posed a major
threat to the ever-increasing agribusiness industry. To prevent these
risks, effective testing (indexing) procedures are required prior to
bulking up culture for commercial propagation. Standard procedure
should be adopted such as:

 Carefully selection of mother plants.

 Ensuring establishment of virus free culture through indexing of
100% explants.
 Proper package and practices to be adopted such as limited
number of cycles of multiplication, grading of cultures as well
as plants, insect, pest monitoring in hardening area etc.
ENVIRONMENTAL CONDITIONS
There are three important aspects:
(i) Nutrient medium
(ii) Aseptic conditions
(iii) Aeration of the tissue

1)Nutrient Medium:
The composition of plant tissue culture medium can vary depending
upon the type of plant tissues or cell that are used for culture. Plant
hormones play important role in growth and differentiation of
cultured cells and tissues. An optimum pH (usually 5.7) is also very
important.
2)Aseptic conditions (Sterilization):
Nutrient medium contains ample sugar which increases growth of
microorganisms such as bacteria and fungi. These microbes compete
with growing tissue and finally kill it. It is essential to maintain
aseptic conditions of tissue culture. Thus, sterilization means
complete destruction or killing of microorganisms so that complete
aseptic conditions are created for in vitro culturing.
3)Aeration of the Tissue:
Proper aeration of the cultured tissue is also an important aspect of
culture technique. It is achieved by occasionally stirring the medium
by sterring or by automatic shaker.
CONCLUSION
 Plant tissue culture as an important tool for the continuous
production of active compounds including secondary
metabolites and engineered molecules.
 Novel methods (gene editing, abiotic stress) can improve the
technique.
 Humans have a long history of reliance on plants for a supply of
food, shelter and, most importantly, medicine. Current-day
pharmaceuticals are typically based on plant-derived
metabolites, with new products being discovered constantly.
 Nevertheless, the consistent and uniform supply of plant
pharmaceuticals has often been compromised.
 One alternative to produce important plant active compounds is
in vitro plant tissue culture, as it assures independence from
geographical conditions by eliminating the need to rely on wild
plants. Plant transformation also allows the further use of plants
to produce engineered compounds, such as vaccines and
multiple pharmaceuticals.
 This review summarizes the important bioactive compounds
currently produced by plant tissue culture and the fundamental
methods and plants employed for their production.
BIBLIOGRAPHY
 https://byjus.com/biology/plant-tissue-culture/
 https://mitsmegafoodpark.com/mobile/documents/project_report/
Plant_Tissue_Culture_laboratory.pdf
 https://www.linkedin.com/pulse/plant-tissue-culture-environmental-
condition-methods-types-dey--1
 https://pubmed.ncbi.nlm.nih.gov/17914178/#:~:text=Plant%20tissue
%20culture%2C%20or%20the,well%20as%20in%20commercial
%20application
 https://www.agriclinics.net/SampleProjects/14.pdf