What is plant tissue culture : What is plant tissue culture Plant tissue culture is the propagation of plants through“cloning” 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 : 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 : 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:Stage1-Establishment of explant in culture. : Stages of Micropropagation:Stage1-Establishment 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:) : 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:) : 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:) : 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 : 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:) : 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:) : 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 : 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 : 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 : 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 : List of technologies which have been perfected for large scale propagation Plant category Plants
Fruits Banana, grapes, pineapple, strawberry
Cash crops Sugarcane, potato
Spices Turmeric, ginger, vanilla, large cardamom, small
cardamom
Medicinal plants Aloevera, geranium, stevia, patchouli, neem
Ornamentals Gerbera, carnation, anthurium, lily, syngonium,orchids
Trees Teak, white teak, bamboo, eucalyptus, populus
and many more genetically enginered plants.
Market projections for TCPs(Volume in thousand nos., Value in Rs. Lakhs) : Market projections for TCPs(Volume in thousand nos., Value in Rs. Lakhs)