What is plant tissue culture

Plant tissue culture is the propagation of plants throughcloning an asexual method of reproduction. A portion (explant) of a desired plant is cultured in vitro on a defined medium, which promotes rapid multiplication of cells. The new plants are removed from the culture and transferred to a standard potting medium. Tissue culture is based on the theory of totipotency; that is, the genetically based ability of a nonembryonic organ or cell to develop along a pathway similar to that of a zygote, leading to the formation of a new entire plant identical to the original.

Why PTC
Currently, tissue culture is being used in both research and commercial applications. Tissue culture not only provides a method of mass propagation, but also makes possible the production of disease-free plants, mutants, and secondary plant products. A new and important use is in the genetic engineering of plants. A single plant can be genetically modified and grown into a mature plant or plants having new characteristics.

Micropropagation
1.Clonal mass propagation. The important point here is that extremely large numbers of plants can be produced. Rather than getting 10000 plants per year from an initial cutting, one can obtain upwards of 1,000,000 plants per year from one initial explant. 2. Difficult or slow to propagate plants. Micropropagation enables growers to increase the production of plants that normally propagate very slowly such as narcissus and other bulbous crops. 3. Introduction of new cultivars eg. Dutch iris. Get 5 daughter bulbs annually. Takes 10 years for commercial quantities of new cultivars to be built up. Can get 100-1000 bulbs per stem section. 4. Vegetative propagation of sterile hybrids used as parent plants for seed production. Eg. cabbage. 5. Pathology - Eliminate viruses, bacteria, fungi etc. Use heat treatment and meristem culture. Used routinely for potatoes, carnation, mum, geranium, garlic, gypsophila 6. Storage of germplasm Generally the only successful method to date is keeping them in refrigerator. Slows down, but does not eliminate, alterations in genotype.

Stages of Micropropagation:Stage1Establishment of explant in culture. 1. Part of plant used 2. Disinfestation - or surface sterilization must clean plant tissues of all contaminating microorganisms

Contd:-

Stage -1(contd:)
3. Medium - Must contain all components necessary to nourish explant a. Inorganics Macroelements - N,P,K,Ca, Mg Microelements - B,Co, Cu, Mn, I, Fe, Zn b. Organics - carbon source - needed since plants do not seem to photosynthesize well in culture c. Vitamins Thiamine - essential Myoinisitol B vitamins folic acid biotin d. Growth regulators Cytokinins Auxins GA, ABA rarely used e. Complex organics - natural orange juice, coconut milk, bananas f. Inert supports Agar, foam rubber, filter paper bridge, liquid

Stage -1(contd:)
4. Growth regulators Basic research findings of Miller and Skoog have been born out by many investigators In general: cytokinins induce shoot bud formation auxins induce root formation 1. Absolute amount 2. Ratio important cyt/aux >1 = shoots cyt/aux < 1 = roots cyt/aux = 1 = callus or either

Stage -1(contd:)
5. Environmental conditions Light - light intensity , photoperiod, quality important. Temperature - there are usually high and low cutoffs. In some cases a specific temperature is needed.

Stage 2:Multiplication
1. Axillary shoot formation Axillary meristem - meristem located in the axil of a leaf and giving rise to an axillary bud. Shoot tips and meristems. Already have organization of shoot. Can use lower amounts of hormones than for other structures where shoot morpholgy is not present. Often desirable because of maintenance of diploidy and chimerism.

Stage 2:Multiplication(contd:)
2. Adventitious shoot formation a. callus organogenesis With most structures other than shoots usually must go through callus first. Get dedifferentiation and then redifferentiation. The process is similar to that seen with root primordia formation. But in tc in most cases want shoot growth first. b. Organ formation directly without callus. Occurs but is not as common. eg. African violet, peperomia. Structures arising not at their usual sites.

Stage 2:Multiplication(contd:)
3. Callus embryogenesis - callus in first stage. Callus then is placed in liquid culture. For callus and liquid culture - use 2,4-D, NAA. Then plate it out without auxin. Then get embryos to form.

Stage 3 : Rooting
Need root formation and adjustment to greenhouse conditions. For root initiation growth regulators are important. Generally aux/cyt>1. NAA used for herbaceous plants. IBA - for woody plants. Other treatments - reduce sugar to encourage autotrophic growth. Light - reduce or increase depending on specific plant. Lower or in darkness for 1 week for root formation. Higher - 10,000 lux to adapt it to higher light intensities in GH or field.

STAGE 4 - Transfer to soil

Must maintain high RH or plants will wilt and desiccate very rapidly. Seems to be less wax on most plants. Stomates on some do not seem to function normally. Either remain open or close too slowly. Disease - succumb to fungus - have not been exposed to any in vitro.

ADVANTAGES OF MICROPROPAGATION
1. Plants often more uniform - only way to propagate them vegetatively. . 2. Plants often grow faster, show improved vigor. 3. Plants often mature earlier than when propagated by seed. Disease free plants are produced in bulk Seasonal independence is one criteria of this technique.

List of technologies which have been perfected for large scale propagation
Plant category Fruits Cash crops Spices Medicinal plants Ornamentals Trees Plants Banana, grapes, pineapple, strawberry Sugarcane, potato Turmeric, ginger, vanilla, large cardamom, small cardamom Aloevera, geranium, stevia, patchouli, neem Gerbera, carnation, anthurium, lily, syngonium,orchids Teak, white teak, bamboo, eucalyptus, populus

and many more genetically enginered plants.

Market projections for TCPs (Volume in thousand nos., Value in Rs. Lakhs)

Crop 2003-04 Volume Banana Pineapple Grapes Sugarcane Potato Turmeric Vanilla Large Cardamom Small Ginger Medicinal Ornamentals Trees Total 200 401 2100 20290 3000 71579 18 28 105 3044 750 7660 300 441 11510 18120 3380 87567 27 31 576 2718 845 8607 400 484 11741 18944 3824 100436 36 34 587 2842 956 9793 500 533 11995 19973 4342 118499 45 37 600 2996 1086 11404 600 586 12275 21172 4951 143731 21613 4618 926 14791 6 634 1000 2000 44 70 140 Value 1945 693 232 592 Volume 27537 5080 975 16271 10 698 1245 2000 49 87 140 2004-05 Value 2478 762 244 651 Volume 35808 5588 1026 18709 22 767 1123 2000 54 79 140 2005-06 Value 3223 838 257 748 Volume 47470 6147 1080 22449 43 844 1123 2000 59 79 140 2006-07 Value 4272 922 270 898 Volume 64060 6761 1137 28055 83 929 1123 2000

Year 2007-08 Value 5765 1014 284 1122

65 79 140

54 41 614 3176 1238 13592