Somaclonal Variation

Somaclonal Variation

• Definition: It is the term used to describe the variation seen in plants that have been produced by plant tissue culture.

• Plant tissue cultures isolated from single explant, or even a single cell can show variation after repeated subculture.

• Dedifferentiation : Callus formation phase.

• Redifferentiation : Morphogenesis phase.

• Cellular totipotency : Phenomenons of dedifferentiation and redifferentiation are inherent in the capacity described as cellular totipotency, a property found only in plant cells and not in animal cells, although totipotent stem cells have now been found in animal tissues also. In other words, while a differentiated plant cell retains its capacity to give rise to a whole plant, an animal cell loses this capacity of regeneration after differentiation.

Somaclones

• Meristem tip culture and micropropagation produced plants with few exceptions, are true to type.

• Regeneration of plants through callus or cell suspension culture can induce changes among cells, tissues and organs.

• Calliclones, protoclones. • Larkin and Scowcroft 1982:

Somaclones.

• Morphological Characters: Plant height, leaf numbers, leaf size and color.

• Physiological Characters: Biotic stress like Disease resistance, or Abiotic stress like temperature resistant, drought resistant etc.

Somaclonal Potential

• The amount of variation that can be expected in vitro will vary with the clone, age of the clone, use of mutagenic agents, and use of selection pressure applied to single cells for stress conditions such as salt level, herbicides, microorganisms or their byproducts, and specific metabolites.

Kinds of Variation

• Epigenetic change

• Cytogentic

• Genotypic change

• Phenotypic change– Genotype X environment interation

Epigenetic change(Transient)

– Change in expression of the gene– Often not permanent– Not expressed in sexual progeny.– Useless for crop improvement.– Dwarfing, leaf shape, growth habits,

drug/stress resistance.

Cytogenetic ChangesStructural DNA changes

• Chromosomal rearrangements– Deletions– Inversions– Duplications– Translocations

Structural Changes in Chromosomes

Deletion Inversion

Duplication Translocation

Genotypic Changes

• The change of a single DNA base sequence.(Point mutation).

• Inherit in mendelian proportion.

• Expressed in sexual progenies

Causes of variation

• Variations pre-exist in the somatic cells: Variable chromosome numbers (diploid, triploid and polyploid) of mother plant.

• Tissues can be acquired from and part of the plant.

• All part of the plant body do not have same tissue system and different tissue systems differ in composition and physiological function

• Age of explant play vital role in variation.

• Different in original plant

• Ploidy numbers

apex

5 cm2n - 50%4n - 50%

20 cm4n - 70%2n - 9%8n - 16%

tobacco

Variability generated due to culture process

• Duration of culture. Age of cultureFrequency of sub-culture

• Role of growth regulators (GR)Both auxins and cytokininsGR can trigger a high rate of cell divisionDirectly induce mutation

• Culture medium, Levels of KNO3, some micro salts and metals destabilize the normal genetic make up.

• Undefined substances like coconut milk and yeast extract may induce somaclonal variation.

To Increase Somaclonal Variation

• Callus and suspension cultures for several cycles

• Regeneration of large number of plants from long-term cultures

• Testing of selected somaclones for genetic stability

• Multiplication of genetically stable somaclones for developing new cultivars.

Isolation for Somaclones

• In vivo screening:

Morphological varaition: like intensity of chlorophyll, leaf shape, size, height, branching patteren and flower numbers, shape and color.

• Physiological changes such as disease resistance and herbicide resistance.

In Vitro Screening

• Major application is the selection of mutant

• Provide the variation for crop improvement

• Assumption is made at cellular level that resistant to particular set of cultural and environmental condition is expressed by intact plant.

Strategy

• Normal media having selective agent like

(salt, herbicides, heavy metals or phytotoxin) at broad range of concentrations.

• Optimize D90 which kills 90% of cultures

• Further batches TC are then exposed to same concentration in the medium.

• Surviving cultures are identified and multiplied on normal medium and finally regenerate.

Studies concerning different aspects of somaclonal variation are important for several reasons.

1. First, it is hailed as a novel source of genetic variation. However, successful utilization of somaclonal variation heavily depends upon its systematic evaluation and judicious utilization in breeding programmes. This necessitates appropriate experimentation.

2. Second, somaclonal variation is of interest as a basic genetic process, since it contradicts the concept of clonal uniformity. The cells and tissues which are expected to produce true to type plants through the processes of de-differentiation, division and re-differentiation, possibly perceive the whole process as stress, as a result of which the genome, known for its plasticity, restructures itself to modulate the expression of genes as demanded by the in vitro conditions.

3. Third, somaclonal variation is unwanted when the objectives is micropropagation of elite genotypes or genetic transformation which partly involve tissue culture. Under such circumstances, prevention or at least minimization of variation is of utmost importance. To achieve this, the frequency, nature and magnitude of somaclonal variation in relation to manipulation of media components, explant source, culture conditions etc. should essentially be understood.

Somaclonal variation induced in

morphological traits in some crop plants Crop CharactersSugar cane diameter, stalk length and weight Cane yield, sugar yield, stalk

number, length diameter, volume, density the weight

Potato Growth habit, maturity period, tuber uniformity and skin color. Photoperiod requirement, fruit production, Late blight.

Tobacco Yield, Days to flowering, plant height, stem diameter, leaf number, leaf length, leaf width and yield Leaf Shape, type of inflorescence.

Rice Number of tillers per plant, number of fertile tillers per plant, average panicle length, frequency of fertile seed, plant stature and flag leaf length. Flowering period, plant height, seed fertility, heading date and acquired resistance to tungro rice virus and leaf hopper.

Oat Plant height, heading date, twin culms, yellow leaf stripe, awn morphology and fertility.

Maize Twin stalks from a single node, reduced pollen fertility and male sterility.

Crop Characters

Brassica Altered leaf way, multiple branching of the stem, precocious flowering from the apex, stem or leaf, abnormal leaves, reduced lamina in leaves, spontaneously absorbing vegetative buds, slow growth, failure to flower, large pollen grains. Delayed flowering altered growth habit and gross morphology

Pelargonium Leaf shape, size and form, flower morphology, plant height, fasciation, pubescence and anthocyanin pigmentation.

Pineapple Leaf colour, foliage density, leaf width and leaf spine formation.

Tomato Male sterility, jointless pedicel, tangerine vifescent leaf, flower and fruit colour.

Wheat showing resistance to rust and resistance to high temperature

Cons and Pros of somaclonal variation

• Disadvantages • No control, may change in undesirable direction.

i.e –ve, may lose important characters.

• Variations are unpredictable in nature– Any kind of variation may occurs– Phenotypic or morphogenic

• May not occur for complex agronomic traits.• May not be genetically stable

– Might bring new variations in up coming progenies– Gentically different from somaclonal variant parent

• Require extensive field testing– To become a new line need several field trails– Vegetative propagation of selected clone– Sexually reproducible characters are preferred– These variants could then be field tested to

ascertain their genetic stability. – Reciprocal crosses between desirable R1'

progeny and seed derived controls would further stabilize the variants and help in seedset among the promising lines.

Advantages• Changes can occur in agronomic traits

– Morphogenic and phenotypic.– Appropriate technology for genetic manipulation of crops with polygenic

traits.

• Changes occur at a high frequency– Rate of high cell division.– Increase in chance of variation.

• Some novel changes – May not occur by conventional breeding.– Significant contribution in plant breeding.– Although variation does effect all Characters, and not always those that are

agriculturally useful. – It is an alternative tool to plant breeding for generation of new varieties that

could exhibit disease resistance and improvement in quality and yield.

• In vitro selection helps in isolation of lines tolerant to abiotic and biotic stresses.

• In vitro selection shortens time in somaclonal isolation with desirable trait.

• somaclonal variation offers an opportunity to uncover the natural variability in plants and to use this genetic variability for new product development.