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Plant tissue culture is a practice used to propagate plants under sterile conditions, often to produce clones of a plant. Different techniques in plant tissue culture may offer certain advantages over traditional methods of propagation, including:

The production of exact copies of plants that produce particularly good flowers, fruits, or have other desirable traits.

To quickly produce mature plants. The production of multiples of plants in the absence of seeds or necessary pollinators to

produce seeds. The regeneration of whole plants from plant cells that have been genetically modified. The production of plants in sterile containers that allows them to be moved with greatly

reduced chances of transmitting diseases, pests, and pathogens. The production of plants from seeds that otherwise have very low chances of germinating

and growing, i.e.: orchids and nepenthes. To clean particular plant of viral and other infections and to quickly multiply these plants

as 'cleaned stock' for horticulture and agriculture.

Plant tissue culture relies on the fact that many plant cells have the ability to regenerate a whole plant (totipotency). Single cells, plant cells without cell walls (protoplasts), pieces of leaves, or (less commonly) roots can often be used to generate a new plant on culture media given the required nutrients and plant hormones.

Contents[hide]

1 Techniques 2 Choice of explant 3 Applications 4 Laboratories

[edit] TechniquesModern plant tissue culture is performed under aseptic conditions under filtered air. Living plant materials from the environment are naturally contaminated on their surfaces (and sometimes interiors) with microorganisms, so surface sterilization of starting materials (explants) in chemical solutions (usually alcohol or bleach) is required. Mercuric chloride is seldom used as a

plant sterilant today, as it is dangerous to use, and is difficult to dispose of. Explants are then usually placed on the surface of a solid culture medium, but are sometimes placed directly into a liquid medium, particularly when cell suspension cultures are desired. Solid and liquid media are generally composed of inorganic salts plus a few organic nutrients, vitamins and plant hormones. Solid media are prepared from liquid media with the addition of a gelling agent, usually purified agar.

In-vitro tissue culture potato explants

The composition of the medium, particularly the plant hormones and the nitrogen source (nitrate versus ammonium salts or amino acids) have profound effects on the morphology of the tissues that grow from the initial explant. For example, an excess of auxin will often result in a proliferation of roots, while an excess of cytokinin may yield shoots. A balance of both auxin and cytokinin will often produce an unorganised growth of cells, or callus, but the morphology of the outgrowth will depend on the plant species as well as the medium composition. As cultures grow, pieces are typically sliced off and transferred to new media (subcultured) to allow for growth or to alter the morphology of the culture. The skill and experience of the tissue culturist are important in judging which pieces to culture and which to discard.

As shoots emerge from a culture, they may be sliced off and rooted with auxin to produce plantlets which, when mature, can be transferred to potting soil for further growth in the greenhouse as normal plants. Editing Plant tissue culture (section)

[edit] Choice of explantThe tissue which is obtained from the plant to culture is called an explant. Based on work with certain model systems, particularly tobacco, it has often been claimed that a totipotent explant can be grown from any part of the plant. However, this concept has been vitiated in practice. In many species explants of various organs vary in their rates of growth and regeneration, while some do not grow at all. The choice of explant material also determines if the plantlets developed via tissue culture are haploid or diploid. Also the risk of microbial contamination is increased with inappropriate explants. Thus it is very important that an appropriate choice of explant be made prior to tissue culture.

The specific differences in the regeneration potential of different organs and explants have various explanations. The significant factors include differences in the stage of the cells in the cell cycle, the availability of or ability to transport endogenous growth regulators, and the

metabolic capabilities of the cells. The most commonly used tissue explants are the meristematic ends of the plants like the stem tip, auxiliary bud tip and root tip. These tissues have high rates of cell division and either concentrate or produce required growth regulating substances including auxins and cytokinins.

Some explants, like the root tip, are hard to isolate and are contaminated with soil microflora that become problematic during the tissue culture process. Certain soil microflora can form tight associations with the root systems, or even grow within the root. Soil particles bound to roots are difficult to remove without injury to the roots that then allows microbial attack. These associated microflora will generally overgrow the tissue culture medium before there is significant growth of plant tissue.

Aerial (above soil) explants are also rich in undesirable microflora. However, they are more easily removed from the explant by gentle rinsing, and the remainder usually can be killed by surface sterilization. Most of the surface microflora do not form tight associations with the plant tissue. Such associations can usually be found by visual inspection as a mosaic, de-colorization or localized necrosis on the surface of the explant.

An alternative for obtaining uncontaminated explants is to take explants from seedlings which are aseptically grown from surface-sterilized seeds. The hard surface of the seed is less permeable to penetration of harsh surface sterilizing agents, such as hypochlorite, so the acceptable conditions of sterilization used for seeds can be much more stringent than for vegetative tissues.

[edit] ApplicationsPlant tissue culture is used widely in plant science; it also has a number of commercial applications. Applications include:

Micropropagation is widely used in forestry and in floriculture. Micropropagation can also be used to conserve rare or endangered plant species.

A plant breeder may use tissue culture to screen cells rather than plants for advantageous characters, e.g. herbicide resistance/tolerance.

Large-scale growth of plant cells in liquid culture inside bioreactors as a source of secondary products, like recombinant proteins used as biopharmaceuticals.

To cross distantly related species by protoplast fusion and regeneration of the novel hybrid.

To cross-pollinate distantly related species and then tissue culture the resulting embryo which would otherwise normally die (Embryo Rescue).

For production of doubled monoploid plants from haploid cultures to achieve homozygous lines more rapidly in breeding programmes, usually by treatment with colchicine which causes doubling of the chromosome number.

As a tissue for transformation, followed by either short-term testing of genetic constructs or regeneration of transgenic plants.

Certain techniques such as meristem tip culture may be employed that can be used to produce clean plant material from virused stock, such as potatoes and many species of soft fruit.

[edit] LaboratoriesAlthough some growers and nurseries have their own labs for propagating plants via tissue culture, a number of independent laboratories provide custom propagation services. The Plant Tissue Culture Information Exchange lists many commercial tissue culture labs. Since plant tissue culture is a very labour intensive process, this would be an important factor in determining which plants would be commercially viable to propagate in a laboratory.

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1. What is biofertilizer?Biofertilizers are ready to use live formulates of such beneficial microorganisms which on application to seed, root or soil mobilize the availability of nutrients by their biological activity in particular, and help build up the micro-flora and in turn the soil health in general.

2. Why should we use biofertilizers?With the introduction of green revolution technologies the modern agriculture is getting more and more dependent upon the steady supply of synthetic inputs (mainly fertilizers), which are products of fossil fuel (coal+ petroleum). Adverse effects are being noticed due to the excessive and imbalanced use of these synthetic inputs. This situation has lead to identifying harmless inputs like biofertilizers. Use of such natural products like biofertilizers in crop cultivation will help in safeguarding the soil health and also the quality of crop products.

3. What are the benefits from using biofertilizers? Increase crop yield by 20-30%. Replace chemical nitrogen and phosphorus by 25%. Stimulate plant growth. Activate the soil biologically. Restore natural soil fertility. Provide protection against drought and some soil borne

diseases.

4. What are the advantages of bio-fertilizers?1. Cost effective.

2. Suppliment to fertilizers. 3. Eco-friendly (Friendly with nature). 4. Reduces the costs towards fertilizers use, especially

regarding nitrogen and phosphorus.

5. What types of biofertilizers are available?1. For Nitrogen

o Rhizobium for legume crops.

o Azotobacter/Azospirillum for non legume crops.

o Acetobacter for sugarcane only.

o Blue –Green Algae (BGA) and Azolla for low land paddy.

2. For Phosphorous o Phosphatika for all

crops to be applied with Rhizobium, Azotobacter, Azospirillum and Acetobacter

3. For enriched compost o Cellulolytic fungal

culture o Phosphotika and

Azotobacter culture

Biofertilizer

product

s

6. What biofertilizers are recommended for crops? Rhizobium + Phosphotika at 200 gm each per 10 kg of seed

as seed treatment are recommended for pulses such as pigeonpea, green gram, black gram, cowpea etc, groundnut and soybean.

Azotobacter + Phosphotika at 200 gm each per 10 kg of seed as seed treatment are useful for wheat, sorghum, maize, cotton, mustard etc.

For transplanted rice, the recommendation is to dip the roots of seedlings for 8 to 10 hours in a solution of Azospirillum + Phosphotika at 5 kg each per ha.

7. How biofertilizers are applied to crops?1. Seed treatment:

200 g of nitrogenous biofertilizer and 200 g of Phosphotika are suspended in 300-400 ml of water and mixed thoroughly. Ten kg seeds are treated with this paste and dried in shade. The treated seeds have to be sown as soon as possible.

2. Seedling root dip: For rice crop, a bed is made in the field and filled with water. Recommended biofertilizers are mixed in this water and the roots of seedlings are dipped for 8-10 hrs.

3. Soil treatment: 4 kg each of the recommended biofertilizers are mixed in 200 kg of compost and kept overnight. This mixture is incorporated in the soil at the time of sowing or planting.

8. How could one get good response to biofertilizer application?

Biofertilizer product must contain good effective strain in appropriate population and should be free from contaminating microorganisms.

Select right combination of biofertilizers and use before expiry date.

Use suggested method of application and apply at appropriate time as per the information provided on the label.

For seed treatment adequate adhesive should be used for better results.

For problematic soils use corrective methods like lime or gypsum pelleting of seeds or correction of soil pH by use of lime.

Ensure the supply of phosphorus and other nutrients.

9. What would be probable reasons for not getting response from the application of biofertilizers?

1. On account of quality of product o Use of ineffective strain. o Insufficient population

of microorganisms. o High level of

contaminants. 2. On account of inadequate storage facilities

o May have been exposed to high temperature.

o May have been stored in hostile conditions.

3. On account of usage o Not used by

recommended method in appropriate doses.

o Poor quality adhesive. o Used with strong doses

of plant protection chemicals.

4. On account of soil and environment o High soil temperature or

low soil moisture. o Acidity or alkalinity in

soil. o Poor availability of

phosphorous and molybdenum.

o Presence of high native population or presence of bacteriophages.

10. What precautions one should take for using biofertilizers?

Biofertilizer packets need to be stored in cool and dry place away from direct sunlight and heat.

Right combinations of biofertilizers have to be used.

As Rhizobium is crop specific, one should use for the specified crop only.

Other chemicals should not be mixed with the biofertilizers. While purchasing one should ensure that each packet is

provided with necessary information like name of the product, name of the crop for which intended, name and address of the manufacturer, date of manufacture, date of expiry, batch number and instructions for use.

The packet has to be used before its expiry, only for the specified crop and by the recommended method of application.

Biofertilizers are live product and require care in the storage Both nitrogenous and phosphatic biofertilizers are to be

used to get the best results. It is important to use biofertilizers along with chemical

fertilizers and organic manures. Biofertilizers are not replacement of fertilizers but can

supplement plant nutrient requirements.

11. Where can I get further information on biofertilizers?You may visit the following Internet sites:http://www.ikisan.com/links/up_riceBiofertilizers.shtml#tophttp://www.entireindia.com/YellowPg/YpCatList.asp?s=1159&cnm=Biofertilizershttp://www.glsbiotech.com/products.htm#biofertilizershttp://www.us.erc.org/greenchannel/gc7/innovativebiotechnologicalproductsforagriculture.php www.suvash.com http://www.kumarbuilders.com/bio.htm,

Biofertilizers - Biofertilizers are defined as biologically active products or microbial inoculants of bacteria, algae and fungi (separately or in combination), which may help biological nitrogen fixation for the benefit of plants. Biofertilizers also include organic fertilizers (manure, etc.), which are rendered in an available form due to the interaction of micro-organisms or due to their association with plants. Biofertilizers thus include the following: (i) symbiotic nitrogen fixers Rhizobium spp.; (ii) asymbiotic free nitrogen fixers (Azotobacter, Azospirillum, etc.);

(iii) algae biofertilizers (blue green algae or BGA in association with Azolla); (iv) phosphate solubilising bacteria;(v) mycorrhizae;

(vi) organic fertilizers.The need for the use of biofertilizers has arisen, primarily for two reasons. First, because increase in the use of fertilizers leads to increased crop productivity, second, because increased usage of chemical fertilizer leads to damage in soil texture and raises other environmental problems. Therefore, the use of biofertilizers is both economical and environment friendly. The pragmatic approach will be to develop the integrated nutrient supply system involving a combination of the use of chemical fertilizers and biofertilizers. India is not self sufficient in fertilizer production.

An estimated capital investment of Rs. 7,000 crores was needed by the end of Seventh Five Year Plan period to achieve self sufficiency. Realizing the  importance of biofertilizers in supplementing the use of chemical fertilizers, the Government of India had launched the 'National Project on Development and use of Biofertilizers’ during the Sixth Five Year Plan.

Under this 7 project, one national centre and six regional centres and 40 BGA production centres have been established. These centres will produce 800 tonnes of Rhizobia and 600 tonnes of BGA annually.

 

Introduction: Biofertilizers are culture of microorganisms used for inoculating seed or soil or both under ideal conditions to increase the availability of plant nutrients.

Classification of Bio-fertilizers -

How to use the Bio-fertilizers

Biofertilizers are mixed with little water for preparing slurry. In that slurry small quantity of sugar or jaggary or gum is added so that the inoculant may get energy for their prolonged survival. The slurry is poured over the seeds which should be kept in a container. The seed is mixed well with the slurry by pouring the mixture into another container. Thus by pouring fourth and back into both containers the seed is nicely mixed with inoculant. Now the treated seed should be dried in cool and dry shady place and sown immediately in the field.

Application of Azolla Culture: There are two methods.

Azolla can be used as green manure in rice field. In this case Azolla culture is inoculated in the main field about 15-20 days before transplanting of rice seedling so that it can be incorporated in the field at the time of puddling.

The nicely prepared field is divided into smaller plots of 100 sq.m. by raising bunds and 5-10 cm deep water level is maintained upto about 20-25 days after inoculation so that a thick azolla mat may be

formed. After about 25 days of inoculation when thick mat is formed, the water level is reduced and azolla mat is mixed into the soil while weeding.

Water Resources | Soil Conservation & Forestry | Package of Practices

 

Plant tissue cultureIntroductionMost methods of plant transformation applied to GM crops require that awhole plant is regenerated from isolated plant cells or tissue which have beengenetically transformed. This regeneration is conducted in vitro so that the en-vironment and growth medium can be manipulated to ensure a high fre-quency of regeneration. In addition to a high frequency of regeneration, theregenerable cells must be accessible to gene transfer by whatever technique ischosen (gene transfer methods are described in Chapter 3). The primary aimis therefore to produce, as easily and as quickly as possible, a large number ofregenerable cells that are accessible to gene transfer. The subsequent regenera-tion step is often the most difficult step in plant transformation studies. How-ever, it is important to remember that a high frequency of regeneration doesnot necessarily correlate with high transformation efficiency.This chapter will consider some basic issues concerned with plant tissue cul-ture in vitro, particularly as applied to plant transformation. It will also lookat the basic culture types used for plant transformation and cover some of thetechniques that can be used to regenerate whole transformed plants fromtransformed cells or tissue.Plant tissue culturePractically any plant transformation experiment relies at some point on tissueculture. There are some exceptions to this generalisation (Chapter 3 will lookat some), but the ability to regenerate plants from isolated cells or tissues invitro underpins most plant transformation systems.Plasticity and totipotencyTwo concepts, plasticity and totipotency, are central to understanding plantcell culture and regeneration.SPB2 2/27/2003 4:06 PM Page 35Plants, due to their sessile nature and long life span, have developed agreater ability to endure extreme conditions and predation than have animals.Many of the processes involved in plant growth and development adapt to en-vironmental conditions. This plasticity allows plants to alter their metabo-lism, growth and development to best suit their environment. Particularlyimportant aspects of this adaptation, as far as plant tissue culture and regen-

eration are concerned, are the abilities to initiate cell division from almost anytissue of the plant and to regenerate lost organs or undergo different develop-mental pathways in response to particular stimuli. When plant cells and tis-sues are cultured in vitro they generally exhibit a very high degree of plasticity,which allows one type of tissue or organ to be initiated from another type. Inthis way, whole plants can be subsequently regenerated.This regeneration of whole organisms depends upon the concept that allplant cells can, given the correct stimuli, express the total genetic potential ofthe parent plant. This maintenance of genetic potential is called ‘totipotency’.Plant cell culture and regeneration do, in fact, provide the most compelling evidence for totipotency.In practical terms though, identifying the culture conditions and stimuli re-quired to manifest this totipotency can be extremely difficult and it is still alargely empirical process.The culture environmentWhen cultured in vitro, all the needs, both chemical (see Table 2.1) and physi-cal, of the plant cells have to met by the culture vessel, the growth medium andthe external environment (light, temperature, etc.). The growth medium has tosupply all the essential mineral ions required for growth and development. Inmany cases (as the biosynthetic capability of cells cultured in vitro may notreplicate that of the parent plant), it must also supply additional organic sup-plements such as amino acids and vitamins. Many plant cell cultures, as theyare not photosynthetic, also require the addition of a fixed carbon source in theform of a sugar (most often sucrose). One other vital component that must alsobe supplied is water, the principal biological solvent. Physical factors, such astemperature, pH, the gaseous environment, light (quality and duration) andosmotic pressure, also have to be maintained within acceptable limits.Plant cell culture mediaCulture media used for the in vitro cultivation of plant cells are composed ofthree basic components:(1) essential elements, or mineral ions, supplied as a complex mixture of salts;(2) an organic supplement supplying vitamins and/or amino acids; and(3) a source of fixed carbon; usually supplied as the sugar sucrose.362 : Plant tissue cultureSPB2 2/27/2003 4:06 PM Page 36For practical purposes, the essential elements are further divided into thefollowing categories:(1) macroelements (or macronutrients);(2) microelements (or micronutrients); and (3) an iron source.Complete, plant cell culture medium is usually made by combining severaldifferent components, as outlined in Table 2.2.Media componentsIt is useful to briefly consider some of the individual components of the stocksolutions.

MacroelementsAs is implied by the name, the stock solution supplies those elements requiredin large amounts for plant growth and development. Nitrogen, phosphorus,potassium, magnesium, calcium and sulphur (and carbon, which is added sep-arately) are usually regarded as macroelements. These elements usually com-prise at least 0.1% of the dry weight of plants.Plant tissue culture37Table 2.1Some of the elements important for plant nutrition and their physiological function.These elements have to supplied by the culture medium in order to support the growth ofhealthy cultures in vitroElementFunctionNitrogenComponent of proteins, nucleic acids and some coenzymesElement required in greatest amountPotassiumRegulates osmotic potential, principal inorganic cationCalciumCell wall synthesis, membrane function, cell signallingMagnesiumEnzyme cofactor, component of chlorophyllPhosphorusComponent of nucleic acids, energy transfer, component of intermediates in respiration and photosynthesisSulphurComponent of some amino acids (methionine, cysteine) and somecofactorsChlorineRequired for photosynthesisIronElectron transfer as a component of cytochromesManganeseEnzyme cofactorCobaltComponent of some vitaminsCopperEnzyme cofactor, electron-transfer reactionsZincEnzyme cofactor, chlorophyll biosynthesisMolybdenumEnzyme cofactor, component of nitrate reductaseSPB2 2/27/2003 4:06 PM Page 37Nitrogen is most commonly supplied as a mixture of nitrate ions (from theKNO3

) and ammonium ions (from the NH4

NO3

). Theoretically, there is anadvantage in supplying nitrogen in the form of ammonium ions, as nitrogenmust be in the reduced form to be incorporated into macromolecules. Nitrateions therefore need to be reduced before incorporation. However, at high con-centrations, ammonium ions can be toxic to plant cell cultures and uptake ofammonium ions from the medium causes acidification of the medium. Inorder to use ammonium ions as the sole nitrogen source, the medium needs tobe buffered. High concentrations of ammonium ions can also cause culture382 : Plant tissue cultureTable 2.2Composition of a typical plant culture medium. The medium described here is that ofMurashige and Skoog (MS)a

Essential elementConcentration in Concentration stock solution (mg/l)in medium (mg/l)Macroelementsb

NH4

NO3

33 0001 650KNO3

38 0001 900CaCl2

.2H2

O8 800440MgSO4

.7H2

O7 400370KH2

PO4

3 400170Microelements

c

KI1660.83H3

BO3

1 2406.2MnSO4

.4H2

O4 46022.3ZnSO4

.7H2

O1 7208.6Na2

MoO4

.2H2

O500.25CuSO4

.5H2

O50.025CoCl2

.6H2

O50.025Iron sourcec

FeSO4

.7H2

O5 56027.8Na

2

EDTA.2H2

O7 46037.3Organic supplementc

Myoinositol20 000100Nicotinic acid1000.5Pyridoxine-HCl1000.5Thiamine-HCl1000.5Glycine4002Carbon sourced

SucroseAdded as solid30 000a

Many other commonly used plant culture media (such as Gamborg’s B5 and Schenk andHildebrandt (SH) medium) are similar in composition to MS medium and can be thought of as‘high-salt’ media. MS is an extremely widely used medium and forms the basis for many othermedia formulations.b

50 ml of stock solution used per litre of medium.c

5 ml of stock solution used per litre of medium.d

Added as solid.SPB2 2/27/2003 4:06 PM Page 38problems by increasing the frequency of vitrification (the culture appears paleand ‘glassy’ and is usually unsuitable for further culture). Using a mixture ofnitrate and ammonium ions has the advantage of weakly buffering themedium as the uptake of nitrate ions causes OH–

ions to be excreted.Phosphorus is usually supplied as the phosphate ion of ammonium, sodiumor potassium salts. High concentrations of phosphate can lead to the precipi-tation of medium elements as insoluble phosphates.MicroelementsThese elements are required in trace amounts for plant growth and develop-ment, and have many and diverse roles. Manganese, iodine, copper, cobalt,boron, molybdenum, iron and zinc usually comprise the microelements, al-

though other elements such as nickel and aluminium are frequently found insome formulations.Iron is usually added as iron sulphate, although iron citrate can also beused. Ethylenediaminetetraacetic acid (EDTA) is usually used in conjunctionwith the iron sulphate. The EDTA complexes with the iron so as to allow theslow and continuous release of iron into the medium. Uncomplexed iron canprecipitate out of the medium as ferric oxide.Organic supplementsOnly two vitamins, thiamine (vitamin B1

) and myoinositol (considered a B vitamin) are considered essential for the culture of plant cells in vitro. How-ever, other vitamins are often added to plant cell culture media for historicalreasons.Amino acids are also commonly included in the organic supplement. Themost frequently used is glycine (arginine, asparagine, aspartic acid, alanine,glutamic acid, glutamine and proline are also used), but in many cases its in-clusion is not essential. Amino acids provide a source of reduced nitrogen and,like ammonium ions, uptake causes acidification of the medium. Casein hydrolysate can be used as a relatively cheap source of a mix of amino acids.Carbon sourceSucrose is cheap, easily available, readily assimilated and relatively stable andis therefore the most commonly used carbon source. Other carbohydrates(such as glucose, maltose, galactose and sorbitol) can also be used (see Chap-ter 3), and in specialised circumstances may prove superior to sucrose.Gelling agentsMedia for plant cell culture in vitro can be used in either liquid or ‘solid’forms, depending on the type of culture being grown. For any culture typesthat require the plant cells or tissues to be grown on the surface of themedium, it must be solidified (more correctly termed ‘gelled’). Agar, producedfrom seaweed, is the most common type of gelling agent, and is ideal for rou-tine applications. However, because it is a natural product, the agar qualitycan vary from supplier to supplier and from batch to batch. For more Plant tissue culture39SPB2 2/27/2003 4:06 PM Page 39demanding applications (see, for instance, the section on microspore culturebelow and Chapter 3), a range of purer (and in some cases, considerably moreexpensive) gelling agents are available. Purified agar or agarose can be used,as can a variety of gellan gums.SummaryThese components, then, are the basic ‘chemical’ necessities for plant cell cul-ture media. However, other additions are made in order to manipulate the pat-tern of growth and development of the plant cell culture.Plant growth regulatorsWe have already briefly considered the concepts of plasticity and totipotency.The essential point as far as plant cell culture is concerned is that, due to this

plasticity and totipotency, specific media manipulations can be used to directthe development of plant cells in culture.Plant growth regulators are the critical media components in determiningthe developmental pathway of the plant cells. The plant growth regulatorsused most commonly are plant hormones or their synthetic analogues.Classes of plant growth regulatorsThere are five main classes of plant growth regulator used in plant cell culture,namely:(1) auxins;(2) cytokinins;(3) gibberellins;(4) abscisic acid;(5) ethylene.Each class of plant growth regulator will be briefly looked at.AuxinsAuxins promote both cell division and cell growth The most important naturally occurring auxin is IAA (indole-3-acetic acid), but its use in plant cellculture media is limited because it is unstable to both heat and light. Occa-sionally, amino acid conjugates of IAA (such as indole-acetyl-L-alanine and indole-acetyl-L-glycine), which are more stable, are used to partially alleviatethe problems associated with the use of IAA. It is more common, though, touse stable chemical analogues of IAA as a source of auxin in plant cell culturemedia. 2,4-Dichlorophenoxyacetic acid (2,4-D) is the most commonly usedauxin and is extremely effective in most circumstances. Other auxins areavailable (see Table 2.3), and some may be more effective or ‘potent’ than 2,4-D in some instances.402 : Plant tissue cultureSPB2 2/27/2003 4:06 PM Page 40