Chapter No. 2 Introduction to Plant Tissue Culture

INTRODUCTION TO PLANT TISSUE CULTURE Contents

2.1

Introduction and Definition

2.2

Advantages of Plant Tissue Culture

2.3

Objective of Tissue Culture Project

2.4

Conceptual Foundation of Plant Tissue Culture Technology

2.5

History of Plant Tissue Culture

2.6

Applications of Plant Tissue Culture

2.7

Flow Chart of Plant Propagation by Tissue Culture IVIethod

2.8

Important Steps in Plant Tissue Culture

2.9

Commercialization of Plant Tissue Culture Technique

2.10 Indian Scenario 2.11 Assistance for Plant Tissue Culture Business from Different Sources 2.11.1

Assistance from Government Agencies

2.11.2

Assistance from Micro Propagation Technology Parks

2.11.3

Assistance from Financial Institutes

2.11.4

Assistance for Marketing

2.11.5

Assistance from Biotech Consortium India Limited

2.12

Conclusion

INTRODUCTION TO PLANT TISSUE CULTURE 2.1 Introduction and Definition Agriculture today is on the verge of a technological revolution, in a manner it has never been seen before. As we enter the new millennium, one key development that comes to our mind is the emergence of biotechnology, which offers some of the best opportunities and solutions to some of the uncontrollable problems faced by us.

Biotechnology is a group of technologies that share two things in common; they manipulate living cells and their molecules and have a wide range of uses that can improve our lives. The major techniques of biotechnology are genetic engineering, cell culture, tissue culture, bioprocessing, protein engineering etc.

Plant Tissue Culture, Cell Culture or Micropropagation is the technique of producing selected plants of known desirable agriculture qualities, in large numbers of plants from small pieces of plant in relatively short periods of time. It is a method of rapid propagation under controlled disease free conditions. Entire crop population with premium qualities can be created from a single elite specimen plant. Depending on the species in question, the original tissue piece may be taken from shoot tip, leaf, lateral bud, stem or root tissue of the mother plant. Ex-plants from selected mother plant are established and multiplied under 'In-Vitro' conditions, providing the optimum pre-requisite for plant growth. These ex-plants go through the initiation, multiplication and rooting methods for producing a cell into a full-fledged plant. These ready plantlets are then hardened in climatically controlled green houses or poly houses. Depending on the species, the plants become ready for plantation in the field. This technique of plant propagation greatly reduces the labour and space

requirement, for producing new varieties and can also markedly enhance propagation rates.

Mettiods of plant propagation and establishment are of particular interest to our country and work on a wide range of vegetables, fruit crops and trees are in progress. Several scientists have been experimenting to extend the application of tissue culture to make plant species commercially important. For example experiments on crops like coconut, date palm. Cashew, Mango, Orange etc. are being made in the different research laboratories.

With the advent of plant tissue technology, it is now possible to propagate fine varieties of flowers, forest, and fruit trees by tiny plantlets. Commercialization of these crops has already taken place. In Ornamental crops, Orchids, Carnation, Gladiolus, Gerbera, Anthurium etc. has been commercially grown. In Fruit crops, Banana, Sugarcane etc. has been commercially grown. In Forest trees. Teak, Eucalyptus etc. has been commercially grown. Medicinal plants are also being experimented by tissue culture method and soon would be ready for commercial plantation.

Biotechnology is an area with a tremendous potential in solving basic problems of food, fiber, fuel and medicine particularly in developing Asian countries.

2.2 Advantages of Plant Tissue Culture are as follows •

Mass multiplication of elite clones: Micropropagation allows the production of large numbers of plants from small pieces of the mother plant. The production requires relatively short periods of time to grow plants. Depending on the species under production, a single ex-plant can be multiplied into several thousand plants in less than one year.

True to Type production: Large number of true to the type plants could be propagated within a short time and space and that too throughout the year. For example, it may be possible to propagate Two to Four lakhs of tissue cultured plants from a single bush or rose against 10 to 15 plants by conventional means. Also, it may take about Two to Four months to produce a healthy planting material by tissue culture means, whereas a minimum of Six to Eight months is required for most species by the latest method of plant propagation. •> Higher Yields: Tissue Culture Plants may have increased branching and flowering, greater vigour and higher yield, mainly due to possibility of elimination of diseases. Flexible method: The flexibility of nurseries can be improved. As the capital investment on mother plant is reduced to almost zero, it may be easier to adapt to changing conditions. Additionally, a better programme of production is possible, because of the greater plant uniformity and the availability in the mass at any time. •

Innovation of new varieties: Tissue culture can be utilized for breeding new varieties.

2.3 Objective of Tissue Culture Project

The primary objective of tissue culture project could be propagation of large quantity of good quality planting material from elite mother plants within short time, space with minimum cost per plant.

2.4 Conceptual Foundation of Plant Tissue Culture Technology i

Plant tissue culture refers to the cultivation "In-Vitro" (Vitro-Glass) of all plant parts, whether a single cell, a tissue or organ, under disease-free conditions on nutrient medium.

In the life cycle of any organism, two gamates of opposite sex fuse to form a single cell-zygote. From this single-celled zygote originates the entire multicellular and multiorganed body of a higher organism. In a flowering plant, for example, structures as functionally diverse as underground roots, green leaves and flowers all arise from the single-celled zygote through millions of divisions of cells. 1. Plant Tissue Culture Course Material, Modern College

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Theoretically, therefore, all the cells in a plant body, whether residing in the flowers, conducting tissues or root tips, should have received the same genetical material as originally present in the zygote. There must be some other factors superimposed on the genetic characteristics of cells, which bring about this vast variation expressed by the genetically identical cells. The process of variations is called differentiation. This differentiation is actually preceded by certain cellular and sub cellular changes. A question that arises at this stage is whether the cellular changes underlying differentiation of various types of cells are permanent and irreversible or whether there is merely an adaptive change to suit the functional need of organism in general.

During the normal life cycle of a plant, it is believed, that the events leading to differentiations are of permanent nature. However, the experiments of Vochting on polarity in cuttings (1878) suggested otherwise. He had observed that all cells along the length are capable of forming roots as well as shoots but their density is decided by their relative positioning of the cutting.

The best way to answer this question and understand more about the interrelationship between different cells of an organ and different organs of an organism would however be to remove them from the influence of their neighbouring cells and tissue and grow them in isolation on nutrient media. This has led to the foundation of a new branch of biology as 'Cell and Tissue Culture'. It is applicable to both plant and animal cells. Plant tissue culture has acquired many practical applications in agriculture, horticulture and forestry. It is increasingly becoming popular as a part of recent field of Biotechnology.

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2.5 History of Plant Tissue Culture 2

The German Botanist Guttlieb Haberlandt first proposed the importance of plant tissue and cell culture in isolation, in 1902. He is regarded as the father of plant tissue culture. He used tissue of Lamium Puroureum and Eichhornia crassipes, the epidemis of Ornithooalium and epidermal hairs of Pulmonaria Mollissima. He grew them on a particular salt solution with sucrose and observed obvious growth in the cells. The cells remained alive for up to 1 month. They grew in size, changed shape; thickening of cell walls occurred and starch appeared in the chloroplasts, which initially lacked it. However, none of the cells divided. The failure was that he was handling highly differentiated cells and the present day growth hormones, necessary for inducing division in mature cells, were not available to him.

Hanning (1940) had initiated a new line of investigation, which later developed into an important applied area of in-vitro techniques. Hanning excised nearly mature embryos of some plants like Raphanus Sativus and successfully grew them to maturity on mineral salts and sugar solution. Van Overbeck (1941) and co-workers demonstrated for the first time the stimulatory effect of coconut milk, which was similar to embryo sac fluid, on embryo development and callus formation in Datura. This proved a turning point in the field of embryo culture, for it enabled the culture of young embryos which failed to grow on a mixture of mineral salts, vitamins, amino acids and sugar.

Subsequent detailed work by Raghavan and Torrey (1963), Norstog (1965) and others led to the development of Synthetic media for the culture of younger embryos. Laibach (1925, 1929) demonstrated the practical applicafion of

2. Plant Tissue Culture Course Material, Modern College

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embryo culture in the field of plant breeding. He isolated embryos from nonviable seeds of a particular plant and reared them to maturity on a nutrient medium.

In 1922, working independently Robbins (USA) and kotte (Germany) reported some success with growing isolated root tips. White made the first successful report of continuously growing tomato root tips in 1934.

During 1939 - 1950 extensive work on root culture was undertaken by Street to understand the role of vitamins implant growth and shoot-root relationship. Gautheret (1934), White (1939) and Nobecourt successfully cultured cells of Salix, Nicotiana-Hybrid and carrot on synthetic media. They, for the first time, demonstrated that growth regulators and vitamins if added to media enhanced the growth forming mess of cells called callus.

Skoog (1944), Tsui (1951), and Miller (1955) demonstrated the induction of divisions in isolated, mature and differentiated cells by using synthetic as well as natural compounds. Muir (1953) developed a technique of growing single cells into liquid medium in case of Tenetes Erecta and Nicotiana Tabacum. Vasil and Hildeprandt (1965) raised whole plants starting from single cells of tobacco. Skoog and Miller (1957) showed that changing the relative concentrations of the two substances in the medium could regulate the organ differentiation.

The first reports of some embryo formation from Carrot tissue appeared in 1958-59 by Reinert (Germany) and Steward (USA). Ball (1946) successfully raised whole plants of Lupinus and Tropaeolum by culturing shoot tips. Morel and Martin (1952), for the first time, recovered virus-free Dahlia plants from infected individuals by culturing their shoots.

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Murashige (USA) used this technique to muUiply plants in large number for several species ranging from ferns to foliage, flower and fruit plants. Guha and Maheshwari (1966) demonstrated the possibility of raising large numbers of plantlets from pollen grains of Dhatura. In 1972 Carlson and others produced the first somatic hybrid between two plants by fiising their protoplasts.

2.6 Applications of Plant Tissue Culture 3

In the early fifties it was observed that plant cells are amenable to chemical manipulations in the medium whereby they can be induced to form organized structures and complete plants. This discovery is considered to be very important for the application of cell and tissue culture methods to overcome several problems connected with agriculture, horticulture and plant breeding.

1) The technique provides a way for rapid multiplication of desirable and rare plants. 20,000 plants/year/bud in turmeric, 1,00,000 plants/year/bud in Eucalyptus were found. 2) As the experiments reveal, virus infected plants also contain some healthy stocks as such they can be obtained by separating shoot tips for their in-vitro propagation. This has given successfiil results in Strawberries and Sugarcane.

3) The development of haploids through the technique of anther culture has a potential significance in basic and applied genetics and plant breeding. During the past 20 years the technique has been successfully extended to about 20 plant species including some economic plants.

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4) The embryo culture has been useful in overcoming seed dormancy. It is also utilized for producing viable plants from crosses, which normally fail due to the death of immature embryos. Experiments were successftil in case of Jute and Rice. 5) The embryo tissue culture is also applied for the propagation of rare plants. In some experiments, coconuts developed soft, solid and fatty tissue in place of the liquid endosperm (Mohan Ram, 1976). These are rare and very expensive, served only at special banquets in Philippines. Under normal conditions the coconut seeds fail to germinate. Using the technique of in-vitro culture of excised embryos De Guzman (1969) succeeded in making plantlets from makapuno nuts. 6) Another important use of embryo culture is found in obtaining some rare hybrids. It is possible to raise complete hybrid plants through embryo culture. This method has been profitably used for many interspecific crosses of crops like Tomato, Papaya and Cotton. 7) It is possible to isolate and culture single cells of plants. This helps in mutant selection in relation to crop improvement, as done in Tobacco, Datura etc. The technique is also useful in the production of some chemical substances in the industry. In some cases cell cultures contained twenty times more chemical content than the roots. 8) Recently tissue culture is used in protoplast culture of different varieties of plants and these protoplasts are used for somatic hybridization. 9) A few high performance crop varieties have been widely adopted, resulting in the disappearance of a large number of older varieties. The forests, which house the wild races of most of the crops, are being cut on large scale. Hence tissue culture can be used to preserve germplasm i.e. tissue conservation of these plants can be done identically.

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2.7 Flow chart of plant propagation by tissue culture method PREPARATION OF MEDIUM

I

STERLISATION IN AUTOCLAVE

POURING IN BOTTLES

SEALING AND RESTERILISATION

SELECTION OF A MOTHER PLANT

I

CUTTING OUT THE EX-PLANT PART

WASHING EX-PLANT IN WATER, SOAP AND ANTISEPTIC SOLUTION

I

REWASHING AND SURFACE STERILISATION OF EXPLANTS IN CHEMICAL SOLUTION

COOLING AND SETTING

I

SHIFTING OF EXPLANT TO LAMINAR AIR FLOW STATION

READY FOR INOCULATION SURFACE STERILISATION IN SODIUM HYPOCHLORITE FOLLOWED BY WASING IN DISTILLED WATER FOR 3-4 TIMES

INOCULATION OF EXPLANT ON STERILISED MEDIUM OF KNOWN COMPOSITION

SHIFTING OF CULTURE TO GROWTH ROOM (I5-25''C, 3000 LUX) 3-6 WEEKS BUNCH OF IN-VITRO SHOOTS (GROWTH)

SUBCULTURING

SEPARATION OF IN-VITRO SHOOTS

TRANSFER TO ROOTING SOLUTION IN GROWTH ROOM -3 WEEKS TRANSFER TO GREEN HOUSE FOR HARDENING 17

2.8 Important Steps in Plant Tissue Culture

1. Preparation of medium and sterilization in Autoclave: In the commercial laboratory, the medium is prepared for initiation, subculture and rooting purposes. The medium comprises of the nutrients such as Micro Nutrients, Macro Nutrients, Vitamins, Irons and growth regulators required for the growth of the plant. The medium is sterilized in the electrically operated Autoclave at Fifteen pounds temperature. This sterilization is important to avoid the bacterial contamination. This medium is then poured into the culture bottles and these bottles are again sterilized in the autoclave. Then the medium is cooled and is ready for inoculation.

2. Selection of a mother plant and sterilization: The commercial laboratory decides the plant species, which are to be multiplied in the laboratory. Accordingly, the mother plants are selected from the virus free areas. Healthy, disease and virus free plants are selected as mother plants. The actual plant part, which is called as ex-plant is selected for inoculation. This plant part is washed with water, liquid soap and antiseptic solution. This ex-plant is washed again with the chemical solution to avoid any fungal contamination coming from the field environment. To remove the remnants of the chemicals it is thoroughly washed by distilled water.

3. Inoculation of explants on sterilized medium of known composition: Inside the laminar flow station, the ex-plant is treated again with a disinfectantChemical called Sodium Hypochloride. After this treatment the ex-plant is washed thrice with double distilled water to remove the chemical remnants. The explants are inoculated on the sterilized medium. The inoculation takes place in the Inoculation room, on the Laminar Air Flow station, which maintains a continuous air current, which keeps this bench without the risk of

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contamination. The expert technicians in the laboratory do the inoculation. (Figure 1, 2)

4. Shifting of cultures to the growth room: Inoculated cultures are then shifted to the growth room where a typical temperature of Fifteen to Twenty degree centigrade with the help of air conditioners and Class 1000 clean air is maintained. The artificial lighting arrangement is also made for the growth of the plant. (Figure 3)

5. Shifting of cultures for Subculture: After three to six weeks from inoculation, the inoculated ex-plant shows growth in multiple shoots. These shoots are transplanted on the subculture or multiplication medium for ftirther growth of the plants. For subculture, separate growth medium is prepared. Several subculture cycles are done in the laboratory for mass production from the ex-plant. (Figure 2)

6. Separation of in-vitro shoots and rooting: The shoots are separated in the laminar airflow stations and these fully grown shoots are transferred to the rooting medium for root generation. Depending upon the species, rooting requires One to Three weeks. (Figure 2)

7. Transfer to green house for hardening: Rooted shoots are removed from the laboratory and are transferred to the green house for hardening. In the hardening procedure the plants are first kept in the humidity chambers for acclimatization and then are transferred to the green house. (Figure 4)

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Figure 1. Closer look of ex-plant at inoculation

Figure 2. Laboratory operators at inoculation, subculture and rooting work.

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Figure 3. Cultures in Growth Room

XV

Figure 4. Plants transferred to green house for hardening

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2.9 Commercialization of Plant Tissue Culture technique

In Western Europe, commercial micro-propagation started as late as early 1980's and there were total two hundred and forty commercial plant tissue culture laboratories producing millions of plants per year by the end of 1988. Even Holland started commercial micro-propagation in 1980's and by the end of 1988 Holland was producing sixty two million plants with the help of sixtyseven tissue culture laboratories. Even Israel had five commercial tissue culture laboratories producing five million plants per year by the end of 1988.4

Also in other countries many commercial tissue culture laboratories were set up and produced millions of plants for their country. These countries included, Poland, Yugoslavia, and Soviet Union etc.

In America commercial micro-propagation started in 1965 and there were about hundred commercial tissue culture laboratories then.

2.10 Indian Scenario

In 1980's while all these countries were producing millions of plants, India had only four commercial tissue culture laboratories. Eventually the laboratories increased, but they were unable to produce the quantity that agriculture and horticulture market needed.

Many commercial plant tissue culture laboratories commenced operations in 1990's. Currently the plant tissue culture is well studies, experimented and

4. Biotechnologies for Agriculture and Aqua Culture. Chapter 27-Present capabilities in Commercial Tissue Culture and the Potential for Growth. Dr. Jitendra Prakash 195-199

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accepted in India. India has achieved a milestone in this technique by conducting research and development with well-equipped research laboratories like; Indian Council for Agriculture Research, Delhi; Indian Institute of Horticulture Research, Bangalore and National Chemical Laboratory, Pune. Now the need is to make this technique strong in the commercial area of production. Despite of the support of the national level Research Institutes, Agricultural Universities and Government Agriculture Department, commercial tissue culture is still facing multifarious problems.

2.11 Assistance for Plant Tissue Culture business from different sources It was found that different type of financial and technical assistance was available for commercial plant tissue culture laboratories. The commercial laboratories did not identify these sources. Following are the different assistance schemes available for commercial tissue culture laboratories. 2.11.1 Assistance from Government Agencies Various Central and State Government agencies have been trying hard to boost agriculture business and agriculture processing sector in the State. The lacking factor is, a dialogue between the farmers, agriculture industry and the Government development agencies. To fill up this gap. Government departments have come up with different schemes. Most of the commercial laboratories were not aware about these schemes. Commercial laboratories should study these schemes and make use of them in making their business cost-effective.

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Schemes implemented by Central and State Government for promotion of Agri Business and Agro Processing Sector:

1. Name of the scheme: To establish Tissue Culture Laboratory in private sector. Scheme implementing agency: Central Government Nature of Assistance: Assistance is available for one unit per year for private sector for establishing tissue culture laboratory. Subsidy available is Rupees Ten Lakhs.

2. Name of the scheme: Assistance for establishing Green Houses. Scheme implementing agency: Central Government Nature of Assistance: Under this scheme for establishing Green Houses for high tech agriculture following assistance is available. a) G.H.I: Green house frame and U.V. Film would be subsidized upto Fifty percent with a limit of rupees Thirty-one thousand two hundred and fifty only for one green house. b) G.H.2: For partially controlled green houses using Fan and Pad, subsidy at the rate of forty percent with the limit of rupees One lakh is available. Both the schemes of green house are provided with the assistance only for five hundred sq.mtr. of area for each beneficiary.

3. Name of the scheme: Assistance for Drip Irrigation for High Value Crops. Scheme implementing agency: Agriculture Department, Government of Maharashtra Nature of Assistance: Under Centrally sponsored scheme assistance is provided for drip irrigation system for fruits, flowers and vegetables. The rate of subsidy is ninety percent for B.C., S.T., small and marginal farmers

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and women. For other farmers the rate of subsidy is seventy percent. Subsidy is available for an area upto limits prescribed under Agricultural Land Ceiling Act. The rate of subsidy is rupees twenty five thousand per hector as the maximum limit.

4. Name of the scheme: Property tax on Green House Scheme implementing agency: Government of Maharashtra Nature of Assistance: Green houses and Poly houses build for high tech cultivators of Vegetables, Flowers and Nursery plants will not be charged property tax by village panchayats.

5. Name of the scheme: To provide electricity at a concessional rate for high tech. agriculture. Scheme implementing agency: Maharashtra State Electricity Board. Nature of Assistance: Electricity rate for high tech agriculture like tissue culture, green houses in Private and Public sector will have concessional rate as follows: a) Rupees two and twenty-five paise per unit for high tech agriculture, which would include tissue culture and mushroom cultivation.

To avail the benefit from Maharashtra State Electricity Board for a concession in electricity charges, the laboratory should be located out side the Industrial area.

2.11.2 Assistance from Micro propagation technology parks In order to promote tissue culture activities, the Department of Biotechnology, Ministry of Science and Technology, Government of India has developed Micropropagation

Technology

Parks.

At

present

there

are

two

Micropropagation Technology Parks in existence. One is located in

25

Maharashtra at the National Chemical Laboratory, Pune and the other is at Tata Energy Research Institute, New Delhi. The technology parks are developed to provide an effective platform for transfer of proven technologies to the entrepreneurs in the field of commercial plant tissue culture. The technology parks also act as an interface between the Research Institutes and the tissue culture business for accelerating commercialization of the tissue culture technology.

Micropropagation technology parks offer following services to the tissue culture business:

1. Technology transfer: Transfer of proven technologies to the users, training, implementation and adoption of the technology at the client's site.

2. Contract research: Development of process for the newer crops, refinement of existing protocol.

3. Technical assistance for production of plants: An indigenously designed highly sophisticated laboratory and green house exists to produce plants on large scale. A strong group of scientists and highly experienced staff are available with the unit and give technical assistance for production of plants for plantation programs or field trials.

4. Training of the personnel: Training programs are organized on general training for plant tissue culture and specialized training on specific plant of interest.

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5. Setting up and commissioning of tissue culture laboratory and green house: Designing of the laboratory and green house, equipping the laboratory and the green house.

6. Advisory consultancy for overall running of the laboratory: Consultancy provided for running a tissue culture unit, which includes all the aspects, right from washing of glassware till production and hardening of plants in the green house.

7. Turn-key project based on the client's need: All the above services can be taken singly or in combinations as per the needs. Turn-key project is also offered which includes all the above services together and any other related aspects or problems.

2.11.3 Assistance from Financial Institutes

Financial assistance is available to tissue culture business houses from the State Bank of India, Bank of Baroda, Central Bank of India, Dena Bank and NABARD. The terms and conditions of assistance differ from institution to institution.

2.11.4 Assistance for Marketing

National Horticulture Board, Ministry of Agriculture, Government of India was set up in 1984 for integrated development of horticulture in the country. National horticulture board encourages and promotes the development of horticulture industry in the country. The board helps in increasing production and marketing of horticulture produce.

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National Horticulture Board's developmental program in marketing:

Development of marketing of horticulture produces through participation of Soft Loan. Objective: To increase flow fresh and processed horticultural produce to targeted domestic and external markets. Commodities: Fruits, vegetables of commercial importance Assistance: The loan support will be made upto forty percent of the loan portion upto a limit of rupees hundred lakhs per project @ four percent surcharge per annum, repayable in five installments after a moratorium of three to five years.

2.11.5 Assistance from Biotech Consortium India Limited

Biotech Consortium India Limited was set up with the objective of providing the linkages to facilitate accelerated commercialization of biotechnology. This organization was incorporated as a public limited company in 1990 under the Indian Companies Act 1956. It is promoted by the Department of Biotechnology, Government of India.

Biotech Consortium India Limited has been engaged

in technology

development, technology transfer, project consultancy, fund syndication, information dissemination, manpower training and placement related to biotechnology.

It has assisted over One hundred and twenty clients including scientists, technologists, research institutions, universities, first generation entrepreneurs, the corporate sector, government, banks and financial institutions.

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2.12 Conclusion

Propagation by tissue culture offers good commercial prospects in ornamental plants, vegetables and fruit plants, where value of the product is high. In India the tissue culture technique has reportedly been successful in more than a hundred species of plants. It has been estimated that in India, more than Three hundred and fifty million tissue cultured plants are being produced annually through tissue culture method.

Plant Tissue Culture has come to stay as a tool in plant biology. Plant Tissue Culture has the potential to resolve the problems of experimental biology, which otherwise through conventional methods is difficult to tackle. In the near future this technique will play a very prominent role in genetic engineering, breeding and afforestation programs. Tissue culture technique is a boon for the agriculture and the horticulture industry because of its numerous advantages. Tissue culture can produce several number of healthy, virus free and true-totype plants. The advantage of this technique is such that these plants can be planted anytime during the year, which solves farmers' seasonal plantation problems.

Disease-free plants, multiplied through tissue culture, produce higher yields than infected ones. This biotechnology application can be used for both, traditional and new varieties. Carefully monitored production would ensure clean plantlets for distribution.

Tissue culture activity is taking a shape of an industry as many farmers are planting tissue culture grown plantlets; agro-traders are buying and selling tissue culture grown plantlets while some are exporting either the plantlets or the produces of plants grown by tissue culture, especially varieties of flowers

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like roses, anthuriums and gerberas are enjoying high profits. Tissue cukure is the greatest advancement in plant breeding. Agriculture and horticulture industry should take full advantage of this technique in reaching the greatest heights at national and international levels.

In India, tissue culture is rapidly becoming a commercial method for propagating new and rare species and difficult-to-propagate plants. From a few research laboratories several years ago, a whole new industry is emerging. Currently, the demand for micro propagated plants is greater than the supply with some plants. Some growers specialize in only the micro propagation of plantlets, leaving the growing-on i.e. hardening activity to others. Many growers are integrating a tissue culture laboratory into their overall operation.

While many plant tissue culture laboratories are coming up, some of the laboratories are being closed down due to various reasons and many of the existing laboratories are found to be complaining about the problems they are facing. Commercial use of plant tissue culture technique has vast business potential if tissue culture laboratories are freed of their constraints, particularly financial and marketing. They are the businesses that would enable India retain its self-sufficiency on agricultural production front. Finance being the heart of any business, finances will have to be controlled and regulated systemically in tissue culture business as well. A thorough study of the financial viability and commercial prospects of the tissue culture venture should be carried out by the new entrant. It is the need of the time with the backdrop of an era of globalization and the emergence of World Trade Organizafion.

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