(indoleacetic acid), IBA (indolebutyric acid), NAA (naphthaleneacetic acid), and 2,4-D
(2,4-dichlorophenoxyacetic acid). Cytokinins promote cell division and shoot growth. An
auxinlike compound TDZ (thidiazuron) has increased success rate of plant regeneration
in many species. The most commonly used cytokinins are BAP (benzylaminopurine),
zeatin, and kinetin. In addition to auxins and cytokinins, other hormones such as abscisic
acid (Augustine and D’Souza 1997; Cardoza and D’Souza 2002) and jasmonic acid
(Bla ́zquez et al. 2004) have also been used in plant cell culture. Other adjuvants (additional
components that enhance growth) that have known to have a positive effect on morphogen-
esis are polyamines such as spermidine, spermine, and putrescene (Cardoza and D’Souza
2002; El Hadrami and D’Auzac 1992; Potdar et al. 1999). By manipulating the amount
and combination of growth hormones, regeneration of whole plants from small tissues is
possible (Fig. 5.2).
Another critical aspect in plant tissue cultures is the management of the gaseous
hormone ethylene. When plants are grown in vitro in closed culture vessels, there is a
buildup of ethylene, which is typically detrimental to the cultures. The addition of
ethylene biosynthetic inhibitors such as silver nitrate (Giridhar et al. 2001), AVG
(aminoethoxyvinylglycine), and silver thiosulphate (Reis et al. 2003) have been shown
to increase the formation of shoots.
Tissues are transferred to fresh media periodically—every week to monthly, depending
on the species and experiment. Without subculturing, tissues will deplete the media and
often crowd each other, competing for decreasing resources.
Figure 5.2.Brassica junceaplants produced from hypocotyl explants. Shoots are produced when
a combination of auxin and cytokinin is used. (a) Callus from hypocotyl explants (note the green
fluorescent protein fluorescent sectors on some of the calli); (b) shoots from callus; (c) shoots elonga-
ting; (d) whole plantlets transferred to soil. See color insert.
5.3. MEDIA AND CULTURE CONDITIONS 117