Abstract:
Tissue culture media provide ideal conditions for growth of plant cells, but also bacteria and fungi. It is
therefore necessary to sterilize media to remove these microbes prior to incubation of explants. Growth
media are commonly sterilised by autoclaving at 121°C and pressure of 105 kPa for 15 minutes, or longer
for larger volumes (Beyl, 2000). Some components of the growth media such as gibberellins (GA3) and capanthothenate
are heat-labile and would become inactive when autoclaved (Nissen & Sutter, 1990). Such
heat sensitive components are sterilised by filtering through bacteria-proof membrane (0.22μm pores) and
added to the sterilised medium after it has cooled down to at least 60°C. Autoclaving the growth media at
121°C and pressure of 105 kPa for 15 - 20 minutes also breaks down sucrose into D-glucose and Dfructose,
resulting in alteration in the osmotic potential of the growth media. Thus, it is important to consider
these changes when performing osmotic-sensitive procedures such as protoplast culture. Moreover, the
simple sugars resulting from sucrose degradation apparently have inhibitory effects on in vitro regeneration
of some plant tissues (Dodds & Roberts, 1990).
