Modern plant tissue culture is practiced under aseptic or favourable conditions under filtered
air. Living plant materials which was taken from the environment or free space are being
naturally contaminated or will show contamination on their external surfaces (and sometimes
interiors) with pathogenic microorganisms, so surface sterilization of initial materials
(explants) in chemical solutions (usually Sodium or calcium hypochlorite or mercuric
chloride) is done. Mercuric chloride is mostly used as a plant sterilant now a days, unless other
sterilizing agents which require for removal of contamination are dispose found to be least
effective, as it is very dangerous and harmful to use, and is very difficult to dispose of (Aniel,
et al., 2011) [1].
Explants are then generally placed or maintained on the upper surface of a
solidified culture medium (normally MS) (Pawar & Maheshwari, 2004) [11], but during some
cases it will be inoculate directly into a liquid nutrient medium, normally when cell suspension
cultures are required. Solid nutrient and liquid nutrient media usually consists of hormones.
Solidified media are prepared or produced from the liquid media with the use of a gelling or
solidified agent, generally a purified agar. The constituents of the
solid medium or the
requirement of the plant growth hormones and the nitrogenous source (nitrate/ammonium salts
and amino acids) have produces morphological effects of the grown tissues that from the
initial explants. For example, high concentration of auxin responsible for the high
multiplication of roots whereas an excess cytokinin may yield shoots.
A balance concentration
of auxin and cytokinin will often generate an unorganised and undifferentiated growth of cells,
i.e. callus (Sabir, et al., 2008) [14], but the morphology of the outgrowth depends on the
different plant varieties as well as the suitable medium composition (Joshi & Padhya, 2010) [5].
As cultures grow, explants transferred to new media (sub-cultured) to allow for growth or to
change the morphology of the culture.
Gita Rani and Avinash, (2003) [4] observed or reported the callus induction or cultivation fromthe hypocotyls, root and cotyledonary leaf segments of
Ashwagandha grown on MS medium provided with various
different concentrations and combinations of the 2,4-dichlorophenoxyacetic
acid (2,4-D) and Kinetin plant growth
harmone. Maximum callusing (near about 100%) was
observed with root and cotyledonary leaf explants segments
grown on Murashige and Skoog medium provided with a
combination of 2,4-di-chlorophenoxyacetic acid (2,4-D) and
Kinetin plant growth harmone.
Gita Rani and Avinash (2003) [4] also told that or suggested
that the frequency of the formation of root was
comparatively lower or become slow with the higher
concentrations of Indole 3- butyric acid (IBA) either alone or
in combination with indole 3- acetic acid (IAA) or NAA and
with all concentrations of indole 3- acetic acid (IAA) alone
in Ashwagandha.
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