Coconut Cloning - An Experience
[This has two parts which were conducted simultaneously. In part one Embryos isolated coconuts were culture in test tubes to understand the factors influencing its germination and growth to get a feel of it for the second important part - the production embryoids from leaf and other vegetative tissues and development clonal plantlets of coconut ]
Coconut is an important commercial plant. Unlike in many other trees no vegetative propagation exists for this tree. It is propagated only through the seeds. In nature all plants obtained from seeds shows variation. Importance of cloning comes from the fact that if one can select naturally occurring or one developed by breeding and clone it, one can over come variation and thus predict the end result. For example, if one selects a naturally occurring superior plant and clone it, one should be able to predict the unit productivity of such plantation. That is, if we identify a super clone which yields over 600 nuts /year, with little or no input and if we develop clones from this plant, it means developing a plantation which yields 6 times higher with reduced expenditure. Genetic manipulation can further lead to addition of genes and developing super clones of coconut.
Coconut is a rather difficult plant to clone. Decades of work by many a government and private bodies have produced any significant result.
Our insufficient knowledge of the biological system seems to hinder the end result of all bio-technological project. This work was done between 1981 to 1987. I am releasing it so that it adds to the existing knowledge, with a caution that biological information is dynamic and sensitive and it has to be handled with care in the light full knowledge of the living system. there is a need to review the very foundation of our knowledge of living system.
A brief sketch of the the species
As reported in Manual on Standardized Research Techniques in Coconut Breeding
edited by G.A. Santos, P.A. Batugal, A. Othman, L. Baudouin and J.P. Labouisse
International plant genetic resources institute [http://www.ipgri.cgiar.org]
Coconut (Cocos nucifera L.) is the sole species of the genus Cocos belonging to the subfamily Cocoideae which includes 27 genera and 600 species. It is a diploid with 32 chromosomes (2n=32). As such, hybridization is mainly intraspecific.
The palm has adventitious roots continually produced from the basal 40 cm or so of the trunk, which is the swollen part or what is termed 'bole', in tall types and in some dwarf hybrids. It has no taproot or root hairs but has lots of primary roots which bear large quantities of rootlets.
The main roots grow out somewhat horizontally from the bole and are mostly found within the topsoil. The main branches grow deeper and may extend laterally to as much as 10 m.
The roots, having no cambium, are noticeably uniform - the main roots reaching a maximum diameter of about 1 cm. The root tip is the actively growing region and behind it is the absorbing area whose epidermis is a single layer of thin-walled cells that gradually thicken and become impervious with age. In old roots, the epidermis disintegrates and exposes the hard hypodermis which is generally red.
The root centre has a stele surrounded by a single-celled pericycle sheath from which rootlets and aerenchymatous (respiratory exchange) protuberances or pneumatophores arise. The respiratory exchange occurs more abundantly nearer the soil surface to allow easy diffusion of oxygen into and carbon dioxide out of the root.
The stem develops from the single terminal bud called the 'cabbage' which is the palm's only vegetative growing point. Under favourable conditions, the foundation of the trunk of a young palm reaches full development within 3-4 years.
Stem growth is fastest at early stages, which can be as much as 1.5 m per year. The incremental growth rate levels off as the palms grow old; up to 10-15 cm per year at about the 40th year and over.
The coconut stem has no cambium. Hence, it cannot regenerate damaged tissues. However, a mature palm may have as much as 18 000 vascular bundles which help it to withstand significant physical damage to its trunk, provided pest entry is prevented
The first leaves of a coconut seedling have the pinnae fused together and appear as entire leaves. After eight to ten have been formed, subsequent leaves tend to split into leaflets. After about 3-4 years, the stem starts to form with a single terminal growing point where new leaves develop. Generally, a normal adult palm produces 12-16 new leaves annually, each bearing a corresponding flower cluster (inflorescence). There are about 30-40 leaves in a healthy crown with a similar number of leaf primordia, each differentiated about 30 months before it emerges as a 'sword leaf'. A mature leaf is 3-4 m long and has 200-250 leaflets. A leaf remains on the palm for about 3 years and thereafter, shed leaving a permanent scar on the trunk.
The age of an adult palm is correlated with the number of leaf scars. The number of scars on the stem, divided by 13, gives the approximate age of the palm in years (Mahindapala 1991). This may be important in estimating the age of existing palms used as parent materials in breeding.
The coconut inflorescence is enclosed in a double sheath or spathe, the whole structure known as a 'spadix' which is borne singly in the axil of each leaf. The palm is monoecious, i.e. its inflorescence carries both male and female flowers. The male flowers are more numerous than the female flowers. The former are borne on the top portion of spikelets which are attached to a main axis or peduncle. The female flowers are situated at the base of the spikelets.
The inflorescence primordium can be detected about 4 months after the first leaf primordium is differentiated; the male and female flowers, 22 months thereafter. The opening of the fully grown spathe occurs 1 year later.
The male flowers are the first to open, beginning at the top of each spikelet and proceeding towards the base. After each flower opening, the pollen is shed, and male flowers abscise, the whole process taking just 1 day. The male phase, however, takes about 20 days in most palms but this may vary according to season and variety.
A female flower remains receptive from 1 to 3 days. Depending on the environmental conditions and variety, the female phase may begin a few days or later after the spathe has opened and lasts 3-5 days in tall palms and about 8-15 days in dwarfs. A normal inflorescence may have 10-50 female flowers. With natural pollination, 50-70% usually abort and fall off, especially those which emerge during severe dry weather. The remaining flowers develop into fruits, which take about 12 months to mature.
The length of the male and female phases is affected by climatic environment and usually do not overlap in the tall types, such that self-pollination rarely occurs. In some dwarfs, particularly the Malayan Dwarf, overlapping of the male and female phases and between spadices usually takes place, promoting selfing. Hence, these dwarfs are reasonably homozygous.
Once pollination and fertilization occur, fruits set and develop to maturity in about 12 months, or less than 1 year for some dwarf cultivars. A count of bunch and fruit set can give a reasonable estimate of yield.
The fruit is a fibrous drupe but with a smooth outside skin (exocarp), which may vary from green to red brown or even ivory. The coat (mesocarp) in the young coconut is white and firm. On the other hand, the ripe nut has a fibrous mass, the husk, from which coir is obtained. Within this fibrous mass is the nut with a hard shell (endocarp) enclosing the kernel (endosperm). Between the shell and the kernel is a thin brown seed coat (testa). It adheres firmly to the kernel which is the white flesh, about 12 mm thick lining the central cavity containing the nut water. Towards the end of maturation, the volume of water in the cavity decreases considerably which may be due to absorption by the endosperm tissue or to evaporation. Matured nuts have a sloshing sound of water inside when shaken. Yield is usually estimated in terms of the number of nuts produced per palm or unit area and weight of equivalent copra.