Germplasm Conservation
1. Germplasm conservation refers to the conservation of living genetic resources like pollen, seeds or tissue of plant material maintained for the purpose of selective plant breeding, preservation in live condition and used for many research works.
2. Germplasm conservation resources is a part of collection of seeds and pollen that are stored in seed or pollen banks, so as to maintain their viability and fertility for any later use such as hybridization and crop improvement.
3. Germplasm conservation may also involve a gene bank, DNA bank of elite breeding lines of plant resources for the maintenance of biological diversity and also for food security.

Somatic embryogenesis is the formation of embryos from the callus tissue directly and these embryos are called Embryoids or from the in vitro cells directly form pre-embryonic cells which differentiate into embryoids
Applications:
i. Somatic embryogenesis provides potential plantlets which after hardening period can establish into plants.
ii. Somatic embryoids can be used for the production of synthetic seeds.
iii. Somatic embryogenesis is now reported in many plants such as Allium sativum, Hordeum vulgare, Oryza sativa, Zea mays and this is possible in any plant.

1. The growing of cells including the culture of single cells or small aggregates of cells in vitro in liquid medium is known as cell suspension culture.
2. The cell suspension is prepared by transferring a portion of callus to the liquid medium and agitated using rotary shaker instrument.
3. The cells are separated from the callus tissue and used for cell suspension culture.

Sterilization of Nutrient Media:
1. Culture media are dispensed in glass containers, plugged with non-absorbent cotton or sealed with plastic closures and then sterilized using autoclave at 15 psi (121^o(C)  for 15 to 30 minutes.
2. The plant extracts, vitamins, amino acids and hormones are sterilized by passing through Millipore filter with 0.2 mm pore diameter.
3. Then added to sterilized culture medium inside Laminar Airflow Chamber under sterile condition.
Other techniques used to remove the microbes
(a) Maintenance of aseptic environment
(b) Sterilization of culture room
(c) Sterilization of explants

Pineapple, banana, strawberry and potato.

Artificial Seed
1. Artificial seeds or synthetic seeds (synseeds) are produced by using embryoids (somatic embryos) obtained through in vitro culture.
2. They may even be derived from single cells from any part of the plant that later divide to form cell mass containing dense cyptoplasm, large nucleus, starch grains, proteins, and oils etc.
3. To prepare the artificial seeds different inert materials are used for coating the somatic embryoids like agrose and sodium alginate.

Types of Plant tissue cultures
Based on the explants some other plant tissue culture types are
(a) Organ culture
(b) Meristem culture
(c) Protoplast culture
(d) Cell culture

(a) Organ culture
The culture of embryos, anthers, ovaries roots, shoots or other organs of plants on culture media.
(b) Meristem Culture:
The culture of any plant meristematic tissue on culture media.

(c) Protoplast Culture:
1. Protoplasts are cells without a cell wall, but bounded by a cell membrane or plasma membrane.
2. Using protoplasts, it is possible to regenerate whole plants from single cells and also develop somatic hybrids.
The steps involved in protoplast culture
(i) Isolation of protoplast:
1. Small bits of plant tissue like leaf tissue are used for isolation of protoplast.
2. The leaf tissue is immersed in 0.5% Macrozyme and 2% Onozuka cellulase enzymes dissolved in 13% sorbitol or mannitol at pH 5.4.
3. It is then incubated over-night at 25^oC.
4. After a gentle teasing of cells, protoplasts are obtained, and these are then transferred to 20% sucrose solution to retain their viability.
They are then centrifuged to get pure protoplasts as different from debris of cell walls.
(ii) Fusion of Protoplast
1. It is done through the use of a suitable fusogen. This is normally PEG (Polyethylene Glycol).
2. The isolated protoplast are incubated in 25 to 30% concentration of PEG with Ca⁺⁺ ions and the Protoplast shows agglutination (the formation of clumps of cells) and fusion.
(iii) Culture of Protoplast:
MS liquid medium is used with some modification in droplet, plating or micro drop array techniques. Protoplast viability is tested with fluorescein diacetate before the culture. The cultures are incubated in continuous light 1000-2000 lux at 25^oC. The cell wall formation occurs within 24-48 hours and the first division of new cells occurs between 2-7 days of culture.

(iv) Selection of somatic hybrid cells:
The fusion product of protoplasts without nucleus of different cells is called a cybrid. Following this nuclear fusion happen. This process is called somatic hybridization.
(d) Cell Suspension Culture
The growing of cells including the culture of single cells or small aggregates of cells in vitro in liquid medium is known as cell suspension culture. The cell suspension is prepared by transferring a portion of callus to the liquid medium and agitated using rotary shaker instrument. The cells are separated from the callus tissue and used for cell suspension culture.

Basic concepts of Tissue Culture
Basic concepts of plant tissue culture are totipotency, differentiation, dedifferentiation and redifferentiation.
Totipotency
The property of live plant cells that they have the genetic potential when cultured in nutrient medium to give rise to a complete individual plant.
Differentiation
The process of biochemical and structural changes by which cells become specialized in form and function.

Redifferentiation
The further differentiation of already differentiated cell into another type of cell. For example, when the component cells of callus have the ability to form a whole plant in a nutrient medium, the phenomenon is called redifferentiation.
Dedifferentiation
The phenomenon of the reversion of mature cells to the meristematic state leading to the formation of callus is called dedifferentiation. These two phenomena of redifferentiation and dedifferentiation are the inherent capacities of living plant cells or tissue. This is described as totipotency.