Plant Regeneration Techniques
Plant regeneration refers to the physiological renewal, repair, or replacement of tissue in plant.
An entire plant can be regenerated from an adult tissue or organ, a mass of unorganized calli, or
even a single cell in a process referred to as plant regeneration.
Plant regeneration occurs when plants repair or replace damaged structures based on the
totipotency and pluripotency of their cells.
Totipotency refers to the ability of a cell to differentiate into a complete individual, whereas
pluripotency involves the differentiation of a specific group of tissues or organs from a cell.
Pathways of Plant Regeneration in Tissue Culture
Regeneration pathways in seed plants can be divided into:
- tissue repair,
- somatic embryogenesis, and
- de novo organogenesis.
The tissue repair concerns how young plant tissues, such as root or leaf tips, repair injured parts
and is often used in plant cutting propagation techniques. However, in tissue culture, plants are
regenerated mainly by somatic embryogenesis and de novo organogenesis.
Somatic Embryogenesis
In somatic embryogenesis, plant somatic cells undergo dedifferentiation into embryonic stem
cells and then by way of embryonic development form complete plants, signifying that plant
cells are totipotent by virtue of the embryogenic callus. Somatic embryogenesis leads to an
exchange in cell fate from a somatic cell back into an embryonic stem cell. Dedifferentiation
through this pathway is usually accomplished under stress conditions, hormonal induction (e.g.,
auxin), or gene expression modification. Somatic embryos can be directly induced from
individual somatic cells or indirectly generated from embryonic callus.
Indirect somatic embryogenesis is the most common pathway, especially in crop plants, and
starts with the embryonic callus (an unorganized cell mass). Embryonic callus induction is
followed by the formation of proembryonic masses on the surface or within the callus mass, from
which single cells or cell clusters develop into somatic embryos. Under appropriate conditions,
somatic embryos can develop into shoots and roots.
The direct somatic embryogenesis lacks the callus phase and is less well defined. In this system,
the explant exhibits a more regular compact cell division and is less prolific. The individual
somatic cell in one or more cell layers divides and bulges under appropriate conditions to
develop into a morphologically recognizable new embryo capable of developing into a whole
plant. These somatic embryos could then be directly germinated into plants without the callus
phase.
Direct and indirect somatic embryogenesis pathways can occur in the same explant, but the
periods of obtaining regenerated plants differ. Compared with the direct somatic embryogenesis
pathway, the indirect pathway has a longer period to regenerate plants due to the callus induction
process. Therefore, the indirect somatic embryogenesis pathway is frequently associated with
somaclonal variation. The indirect somatic embryogenesis pathway results in more regenerated
plantlets than the direct pathway due to the plentiful proliferation of callus. Therefore, if the goal
is rapid regeneration of plants, the direct pathway is more efficient than the indirect pathway.
However, for species in which explants are difficult to obtain or situations where many
regenerated plants are desired, the indirect pathway is the better choice.
De novo Organogenesis
De novo organogenesis refers to the regenerative process that does not use a somatic embryo but
rather the differentiation of the meristematic centre, reflecting the pluripotency of plant cells.
Plant-regenerative mechanisms, such as de novo organogenesis, result in regenerating
adventitious roots and/or adventitious shoots in vitro or from injured plant organs. The
regeneration process of adventitious shoots and roots is referred to as de novo shoot
organogenesis and de novo root organogenesis.
Like somatic embryogenesis, de novo organogenesis can also be categorized as either a direct or
indirect regeneration pathway. As with somatic embryogenesis, shoots or roots are directly
induced from pre-existing meristems or injured organs under specific conditions.
Cutting-propagation technology is based on the direct de novo organogenesis used to regenerate
organs. During indirect de novo organogenesis, cells undergo dedifferentiation and plant growth
regulators stimulate cell division after which additional dedifferentiated cells are induced with
further culture time and ultimately generate pluripotent callus. When all conditions are met, the
pluripotent callus undergoes physiological and biochemical changes, resulting in different cell-
division positions and directions.
Factors Affecting Plant Regeneration
The ability of plant regeneration is affected by some environmental factors which include;
- Explant Sources – e.g. age of explants used can affects plant regeneration in tissue
culture. Although every living cell can regenerate entire plants, most studies use cells or
tissues with active growth. Explants in the juvenile-development phase are more
regenerative and possess higher totipotency than those of adult explants
- Plant Growth Regulators (PGRs) e.g. auxin, cytokinin, 2,4-Dichlorophenoxyacetic acid
(2,4-D).
- Basal culture medium (i.e types of culture media) - Murashige and Skoog (MS), N6,
woody plant medium (WPM), and B5, are some of the culture media that are utilized for
callus induction and shoot differentiation and have a big impact on plant regeneration in
tissue culture.
- Light/Dark treatment - Under light conditions, phenolic compounds in explants will be
oxidized by polyphenol oxidases, and the tissue will turn brown. The oxidation products
can darken tissues and inhibit the activity of various proteins, with a potentially adverse
effect on the formation of somatic embryos. Therefore, callus initiation, maintenance, and
maturation require dark conditions in plant for many species.
For further studies:
Long Y, Yang Y, Pan G and Shen Y (2022) New Insights IntoTissue Culture Plant-
Regeneration Mechanisms. Front. Plant Sci. 13:926752.
