Somatic Embryogenesis in Dicotyledonous and Monocotyledonous Culture
Somatic Embryogenesis in Dicotyledonous Culture:
Totipotent embryogenic cells can be obtained from explants of embryogenic or young seedling tissue. Excised small tissues from young inflorescence are equally effective in including somatic embryogenesis in cultures. Other explants used are scuttelum, young roots, petiole, immature leaves and immature hypocotyl. Citrus nucellar cells have natural potential for somatic embryogenesis. Somatic embryos germinate in situ or when they are excised and cultured individually on fresh semisolid medium. a special and noteworthy feature may be the development of a fresh crop of adventive embryos which originate from single epidermal cells on the stem surface of the plantlets obtained from germinating embryos.
The essential requirement for induction and promotion of somatic embryo is suspension culture. Presence of auxin- in the medium is generally essential for embryo initiation. First, the callus is initiated and multiplied on a medium rich in auxin (2,4-D, 0.5mg/l ) which induce differentiation of localized groups of meristematic cells called “Embryogenic clump” ( ECs). Second, ECs develop into mature embryos when transferred to a medium with a very low level of auxin (0.01 to 0.1mg/l) or no auxin at all. Consequently the medium with auxin is called a ‘Proliferation medium (PM) and without auxin an ‘ embryo development medium’ (EDM). In some cases, NAA, IBA , BAP , ethephon also induce embryogenesis.
Somatic Embryogenesis in Monocotyledonous Culture:
Many monocotyledonous plants are of agricultural and medicinal importance. Unlike dicot, the vegetative parts of monocot plant donot readily proliferate in culture. Therefore, explants are best taken from embryogenic on meristematic tissues.
Selection of Explant:
i) Zygotic Embryo:
In case of zygotic as explant, young caryopses ( 10-15 day after pollination) or seeds are surface sterilized by normal procedure and zygotic embryos are excised aseptically and transferred to a culture vials containing MS medium supplemented with 2, 4-D in case of cereals and sucrose (2-6%). Cultures are incubated in diffuse light or complete darkness. Culture will develop within 4-6 weeks.
ii) Young Inflorescence:
Premitotic inflorescence, with primordia of individual florets just beginning to protrude is best suitable material in some cases. The inflorescence of 1-2cm in length is excised. Sterilized and each inflorescence is exposed by vertical incision through the surrounding leaves and then cut into 1-2 mm thick segments. Individual segments are then cultured on medium containing 2,4-D for proliferation and initiation of embryogenic callus.
iii) Young Leaf:
Leaves of young seedlings obtained from seeds, germinate under aseptic condition, are removed and cut into 1-2 mm thick transverse segments starting from the level of the shoot meristem upto the leaf apex. Six to eight examples are placed on nutrient medium to obtain a callus.