View/Open - ResearchSpace - University of KwaZulu-Natal

In vitro culture initiation and multiplication
Seedling organs of R. flava and R. leipoldtii were placed in culture tubes with MS
media with nine different plant growth regulator treatments; a control with no plant
growth regulators, 4.4 µM BA, 22.2 µM BA, 4.5 µM 2,4-D, 22.6 µM 2,4-D, 4.4 µM BA
and 4.5 µM 2,4-D, 4.4 µM BA and 22.6 µM 2,4-D, 22.2 µM BA and 4.5 µM 2,4-D, and
22.2 µM BA and 22.6 µM 2,4-D. Seedling organs of R. diversiformis and R.
minutiflora were placed in culture tubes with MS media with 12 different plant growth
regulator treatments; a control with no plant growth regulators, 2.3 µM kinetin, 23.2
µM kinetin, 5.4 µM NAA, 26.9 µM NAA, 53.7 µM NAA, 2.3 µM kinetin and 5.4 µM
NAA, 23.2 µM kinetin and 5.4 µM NAA, 2.3 µM kinetin and 26.9 µM NAA, 23.2 µM
kinetin and 26.9 µM NAA, 2.3 µM kinetin and 53.7 µM NAA, and 23.2 µM kinetin and
53.7 µM NAA.
5.2.2 Explants from embryos
The seeds were surface sterilised as with in vitro germination experiments described
in Chapter 4 and imbibed for 5 days. Decontaminated seeds were placed in a Petri
dish with sterile distilled water. The seeds were left to imbibe in the laminar flow
cabinet and were dissected every 24 h to test for ease of dissection. Seeds could
only be dissected after 4 days, as the blade could not penetrate the periderm (fruit
coat) with a clean cut with less time for imbibition. The embryos that were dissected
after 7 days of imbibition did not show any growth response when placed on media.
To excise the embryo some of the surrounding endosperm had to be cut away (See
Figure 5.1).
117