(a) What is somatic hybridization ? Explain its steps in production of hybrid plants. (10 marks) (b) What are the biochemical…

(a) Somatic hybridisation (10 marks)

Somatic hybridisation is the fusion of protoplasts (cells stripped of their walls) from somatic cells of two different plants, and the regeneration of a hybrid plant from the fused cell. It bypasses sexual incompatibility, and can combine distant species, as in pomato (potato and tomato). Steps in producing hybrid plants:

1. Isolation of protoplasts: leaf or callus tissue is treated with cell-wall-degrading enzymes (cellulase, pectinase, macerozyme) in an osmoticum such as mannitol, to prevent bursting; protoplasts are purified.
2. Fusion of protoplasts: induced by polyethylene glycol, high calcium at high pH, or electrofusion.
3. Identification and selection of hybrid cells: by complementation markers, fluorescent labels, or mechanical isolation of heterokaryons.
4. Culture of fused cells: cell wall regenerates, and the cell divides to form callus.
5. Regeneration of plants: shoots and roots from callus on suitable hormone media; then hardening.
6. Verification of hybrid nature by chromosome count, isozymes and molecular markers.

The result may be a symmetric hybrid or a cybrid, with cytoplasm from one parent, useful for traits such as male sterility.

(b) Biochemical changes in seeds under prolonged storage (10 marks)

During long storage, seeds age and lose vigour and viability, with the following biochemical changes:

• Lipid peroxidation by free radicals, giving toxic products (malondialdehyde) and loss of membrane integrity, which raises leakage of electrolytes and sugars.
• Hydrolysis of stored lipids to free fatty acids by lipases, and rancidity in oilseeds.
• Declining activity of enzymes such as dehydrogenases, catalase, peroxidase and amylases, and loss of respiratory capacity and ATP production, from mitochondrial damage.
• Denaturation and degradation of proteins, and formation of insoluble complexes.
• Hydrolysis of starch and increase in reducing sugars, and Maillard-type reactions between sugars and proteins.
• Damage to DNA and RNA, with breaks and chromosomal aberrations, and reduced protein synthesis and repair.
• Decreased soluble sugars that protect membranes, and accumulation of aldehydes and other toxic metabolites.

These changes are faster at high moisture and temperature, and are slowed by dry and cool storage in airtight containers.

(c) Factors influencing nutrient absorption from the soil (10 marks)

Nutrients reach the root by mass flow, diffusion and root interception, and are absorbed by active and passive processes. The factors are:

• Plant factors: species and variety; size, extent and surface of the root system and root hairs; root cation exchange capacity; mycorrhizal association; stage of growth; and metabolic energy from respiration.
• Nutrient supply: the concentration and form of nutrient in soil solution, and its replenishment from the exchangeable and solid phases.
• Soil moisture, since nutrients move in the water film, and transpiration draws them by mass flow.
• Aeration: oxygen is needed for respiration and active uptake, and waterlogging reduces absorption.
• Temperature: uptake rises with temperature up to an optimum.
• Soil reaction and organic matter: pH controls availability, for example of phosphorus and micronutrients.
• Ionic interactions: antagonism (for example potassium with calcium and magnesium) and synergism (nitrate and cation uptake).
• Salinity, light and photosynthesis (which supplies energy), and rhizosphere microorganisms.

(d) Characteristics of a clone (10 marks)

A clone is a group of genetically identical individuals produced from a single plant by vegetative (asexual) propagation, using parts such as cuttings, tubers, suckers, runners, grafts or tissue culture. Examples are potato, sugarcane, banana, sweet potato, cassava and most fruit varieties. Its characteristics:

• All members are genetically uniform, since they come from mitotic division of one parent, and are identical with it.
• They are often highly heterozygous, and the heterozygosity is preserved without segregation, since no meiosis occurs, so the type is fixed and true to type.
• Propagation is by vegetative means, so a desirable genotype is multiplied quickly and maintained indefinitely, even where seed is scarce or the crop is sterile (banana).
• Variation arises only by somatic mutation (bud sports), and clonal selection acts on it.
• They may carry systemic diseases and viruses from the parent, and a narrow genetic base makes them vulnerable to pests.
• New variability comes by hybridisation and selection among seedlings, with the best genotype then cloned.

(e) Mutagens (10 marks)

A mutagen is a physical or chemical agent that raises the frequency of heritable changes in the genetic material (mutations) above the spontaneous level. Groups of mutagens:

• Physical mutagens, ionising radiation: X-rays, gamma rays (cobalt-60), alpha and beta particles, and fast neutrons, which cause breaks in chromosomes and gene mutations; and non-ionising ultraviolet light, which forms pyrimidine dimers and is used for pollen and microorganisms.
• Alkylating agents (chemical): ethyl methane sulphonate (EMS), methyl methane sulphonate, ethyl imine and nitrogen mustard add alkyl groups to bases and cause mispairing.
• Base analogues (chemical): 5-bromouracil and 2-aminopurine are incorporated in DNA in place of normal bases.
• Acridine dyes (chemical): proflavin and acridine orange intercalate in DNA and cause frameshifts.
• Other chemicals: nitrous acid and hydroxylamine, which deaminate or modify bases, and sodium azide.

Mutagens are used in mutation breeding to create variation in crops, as in the wheat variety Sharbati Sonora developed by gamma irradiation.