Elementary plant Biochemistry and Biotechnology (2+1)

Stage 2. Multiplication / Proliferation

1. This is the most crucial stage since it is the point at which most of failures in micropropagation occur. Broadly three approaches have been followed to achieve in vitro multiplication.

1.Through callusing (in direct morphogenesis)

• The potentiality of plant cells to multiply indefinitely in cultures and their totipotent nature permit a very rapid multiplication of several plant types. Differentiation of plants from cultured cells may occur via shoot-root formation (organogenesis) or somatic embryogenesis. Somatic embryogenesis is most appealing from a commercial angle. A somatic embryogenesis system once established lends itself to better control than organogenesis.


• Since somatic embryos are bipolar structures, with defined root and shoot meristems, the rooting stage required for microshoots gets eliminated. Above all, somatic embryos being small, uniform and bipolar are more amenable to automation at the multiplication stage and for field planting as synthetic seeds, offering cost advantages from labour savings, can also be stored through cold storage, cryopreservation or dessication for prolonged periods. These characteristics make somatic embryogenesis potentially a less expensive and flexible system for micropropagation. The most serious objection against the use of callus cultures for shoot multiplication is the genetic instability of their cells since it may lead to somaclonal variation.

2.Adventitious bud formation (direct morphogenesis)

• Buds arising from any place other than leaf axil or the shoot apex are termed adventitious buds. The shoots differentiated from calli should also be treated as adventitious buds. In many crops, vegetative propagation through adventitious bud formation from root (blackberry, raspberry) and leaf (Begonia, Crassula) cuttings is standard horticultural practice. In such cases the rate of adventitious bud development can be considerably enhanced under culture conditions.


• For most bulbous plants (e.g. Lilley) adventitious bud formation is the most important mode of multiplication and the best explants are obtained from bulb scales. A serious problem may arise when this method of propagation is applied to varieties which are genetic chimeras. Adventitious bud formation involves the risk of splitting the chimeras leading to pure type plants. For example, in variegated geranium cv. Mme Salleron, the chimera is perpetuated in meristem culture but broken down in petiole culture.

3.Enhanced axillary branching

• In cultures the rate of shoot multiplication by enhanced axillary branching can be substantially enhanced by growing shoots in a medium containing a suitable cytokinin at an appropriate concentration with or without auxin. Due to continuous availability of cytokinin, the shoots formed by the bud, present on
Stages of somatic embryogeneis in dicotyledons and monocotyledons



4.The explant

• Develops axillary buds which may grow directly into shoots. This process may be repeated several times and the initial explant transformed into a mass of branches. This method is the most preferred one for clonal propagation of the mother plant.