**4.1 Organogenesis**

There are several methods to regenerate cultured plants, one of it can be through organogenesis, either through direct organogenesis or indirect organogenesis. Organogenesis is the development of individual plant organ such as shoots or roots from the cells in culture (can be callus (indirect) or plant tissue (direct)) by the process of differentiation. Organogenesis in plant tissue culture involves two stages: Dedifferentiation and redifferentiation. Dedifferentiation starts shortly after explant initiated rapid cell division and consequently forms a mass of undifferentiated cells (callus). Redifferentiation, also referred as budding, is the process where the callus starts to differentiate to form plant organ (organ primordia). This primordia organ is then develop into small meristems (which contains of large nuclei).

In direct organogenesis, the formation of plant organs such as shoot and root is straight from organized tissue (explant) without undergoes callus formation. Progenies that are produced through this technique have identical genetic content as parent. However, this technique is depending on several factors such as explant type, age of explant and size of explant. If meristem tissues is used as explant, the genetic content of the progeny (offspring) are identical as parent. Conversely, if embryos are used as explant, the genetic content between progeny and parent are not identical as embryo formed through fertilization of gamete cells (male and female gametes) and some plants have dormancy period.

In indirect organogenesis, callus is produced first from explant. Callus is disorganized group of cells, rapid dividing, and undifferentiated into specialized tissues such as shoots and roots. Callus can be induced from callus culture, explant (tissues) or cell suspension culture of that callus. Subsequently, organ formation is induced from the callus where shoots and roots are stimulated by plant growth hormones. However, the disadvantage is the changes or variation in the genetic content of somatic cells of the progeny (somaclonal variation) due to repetitive subcultures. Direct organogenesis can be opted to avoid somaclonal variation. Direct organogenesis is often used to plants that are difficult to propagate and do not have abundance of meristem tissues. Therefore, these plants are propagated by using leaves, stems, and root tips as explants.

The rule of thumbs in organogenesis technique are the proportion of growth hormones combinations in the culture medium used to stimulate the respective organs. In direct organogenesis, high ratio of auxin to cytokinin will produce roots while high ratio cytokinin to auxin will induce shoots. In indirect organogenesis, balance or same ratio of auxin and cytokinin (1:1) will produce callus.

Organogenesis starts with induction process caused by the plant hormones in the medium, substances carried over from the explants and endogenous hormones produced by the explants. Organogenesis was first induced by Skoog in 1944 on the formation of root and inhibition of shoot after the addition of auxin. It was then proposed that the regulation of organogenesis is depending on the balance between auxin and cytokinin. The research team then later discovered that high ratio of auxin to cytokinin stimulated the root formation in tobacco callus, but a low ratio of auxin to cytokinin led to shoot formation (**Table 1**).


**Table 1.**

*Standard concentrations of auxins and cytokinins to induce* in vitro *organogenesis.*

### **4.2 Somatic embryogenesis**

Other than organogenesis, somatic embryogenesis is another major regeneration technique in plant tissue culture. Embryo production is an important feature of the flowering plants. The process of embryo formation is called embryogenesis which starts from a single embryogenic cell and subsequently develops into either a zygote or undifferentiated callus cells. Embryo that develops from zygotes is called as zygotic embryos. Meanwhile, embryo that develops from somatic cells is called as somatic embryos where it is artificially induced in cultured plant tissues.

Somatic embryogenesis was first induced in cell suspension culture and callus culture of carrot. Other plants like *Coffea Arabica*, *Citrus cincensis*, *Nicotiana tabacum*, *Pinus ponderosa* and *Cocos nucifera* are among successful species in somatic embryogenesis. In plant tissue culture, somatic embryogenesis is the formation of somatic embryoids from somatic tissues of callus or cells of suspension culture, which can then develop into complete plants in a similar way to the zygotic embryos (sexual reproduction). Somatic embryoid (asexual embryo) is small and wellorganized structure that is resemblance to zygotic embryo (sexual embryo), which is produced from embryogenic somatic cells. Somatic and zygotic embryoids share the same pattern of development where both undergo globular, heart, torpedo shaped and cotyledon stage for dicots and conifers. Embryo growth is bipolar which produces a shoot and a radicular pole at the other end. When encapsulated with suitable nutrient, somatic embryos become artificial or synthetic seeds and they as they can produce plantlets and planted directly into the field.

Somatic embryos can be produced through direct or indirect somatic embryogenesis. In direct somatic embryogenesis, the embryo is induced directly from cells or tissues without the formation of intervening callus. However, this technique is rare and uncommon compared to indirect somatic embryogenesis. In indirect somatic embryogenesis, callus if first formed from explant. Somatic embryos can be then induced from the callus or cell suspension culture of that callus. The embryoids are initiated from superficial callus aggregates where the cells contain large vacuole, dense cytoplasm, large starch granules and nucleus.

Two types of medium with different compositions are required to induce somatic embryoids. First medium contains auxin to initiate embryogenic cells. Second medium is lacked or reduced of auxin, is needed to support the development of the embryogenic cells into embryoids and plantlets. Similar to zygotic embryos, the embryogenic cells pass through 3 different stages i.e. globular, heart shaped, and torpedo shaped, to form embryoids. The embryoids can be separated from the non-embryoids callus cells by using glassbeads or filter paper.

The importance of somatic embryogenesis in agriculture, horticulture, and plant conservation is the zygotic and nucellar embryogenic can be obtained separately from the polyembrogenic plants such as citrus. Since somatic embryo has no food reserves, they can be preserved as encapsulated seeds (surrounded with

*An Overview of Oil Palm Cultivation via Tissue Culture Technique DOI: http://dx.doi.org/10.5772/intechopen.99198*


**Table 2.**

*Comparison between zygotic and somatic embryo.*

nutrients). This makes international exchange of germplasm possible. This artificial seeds provide an advantage for embryos of big and heavy fruits like coconut which can be preserved in a test tube for months and then cultured on medium. In addition, some plants that are crossed interspecific or intergeneric are failed to develop at maturity stage, therefore, before the embryos undergo maturity, they can be taken and cultured on artificial medium and grown into whole plants. As somatic embryogenesis produces many somatic embryos in cell culture, this technique is regarded as the ideal mass propagation system. The somatic embryo is a bipolar system which can develop directly into complete plant, hence, there is no need for separate rooting and shooting induction steps. Plants that derived from somatic embryo may be free of viral and pathogens. Therefore, it is another option in disease-free plants generation (**Table 2**).
