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Table 2.4: Topographic stain evaluation classes for the TTC test (LEADEM, 1984). Class Description Viability 1 Embryo completely stained Germinable 2 Very pale staining Possibly germinable Literature review 3 Cotyledons unstained Non-germinable or possibly germinable 4 Radicle unstained Non-germinable or probably not germinable 5 No staining Non-germinable The meristems should be well stained in order to insure healthy germination and growth of the embryo (LEADEM, 1984). A typical viable embryo should be at least 75% stained and its tissue should be firm with a smooth surface (LEADEM, 1984). The TTC stain test leaves a number of uncertainties. A sample may, for example, not be stained because the stain never penetrates the tissue and the non-enzymatic reduction of TTC is also possible in dead and living tissue (BAND & HENDRY, 1993). A second viability test should be used to confirm the tetrazolium result (BAND & HENDRY, 1993). 2.8.9 After-ripening In the period of after-ripening individual or collective changes take place so that a seed that was once dormant can germinate (COPELAND, 1976). This period may be determined by physical and chemical stimuli in the environment such as a phytochrome response and or the balance of inhibiting and promoting substances in the environment (COPELAND, 1976). It may also be determined by the effect of the environment on the balance of inhibiting and promoting substances within the seed and embryo itself and morphological growth and development of the embryo (COPELAND, 1976). After-ripening can sometimes be hastened by growth promoting substances, low- temperature stratification, alternating temperature and light exposure treatments 59
Literature review (COPELAND, 1976). Such growth promoting substances are synthesised by the seed during this after-ripening process (KHAN, 1977). These include gibberellins, cytokinins and auxins. 2.8.10 Embryo-excision as a tool for investigating mechanisms behind dormancy and testing viability Embryo excision serves as a good test for determining whether seed dormancy is endogenous or exogenous. This test can also be used to test seeds in cases where embryos require long periods of after-ripening before germination will take place (HARTMANN & KESTER, 1965). It is also useful to determine the viability of slow germinating seed (INTERNATIONAL SEED TESTING ASSOCIATION, 1999b). The embryo is essentially excised from the seed and germinated. Before excision is attempted, seeds must be soaked thoroughly, changing the water once or twice daily to avoid the accumulation of seed exudates, to retard the growth of contaminants and to circumvent the evolution of anoxic conditions (INTERNATIONAL SEED TESTING ASSOCIATION, 1999b). During the excision the embryo should be kept moist and working conditions should be aseptic. Observations such as predated, empty, decayed and discoloured seeds and deformed embryos should be noted and included in a calculation of viability (INTERNATIONAL SEED TESTING ASSOCIATION, 1999b). A viable embryo shows some indication of germination, whereas a non-viable embryo becomes discoloured and deteriorates (INTERNATIONAL SEED TESTING ASSOCIATION, 1999b). Signs of germination include the expansion of cotyledons, the development of chlorophyll and the growth of the radicle and plumules. The time required for this test ranges from 3 days to 3 weeks (HARTMANN & KESTER, 1965). Viability is calculated by dividing the number of viable embryos with the total number of seed tested and is reported as a percentage (INTERNATIONAL SEED TESTING ASSOCIATION, 1999b). 60
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Propagation of Romulea species Pier
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Contents 2.7 GERMINATION PHYSIOLOGY
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Contents 5.3.1 Explants from seedli
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Abstract The suitability of various
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Declarations I Pierre André Swart,
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Declarations DETAILS OF CONTRIBUTIO
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Publications from this thesis ASCOU
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List of Figures Figure 2.13: Suther
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was significantly different from th
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List of Tables Table 2.1: Names of
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List of Abbreviations 2,4-D 2,4-Dic
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Die vlakhaas spits sy oor hy het di
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As die mens dan ook so sy dankbaarh
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Introduction where a great number o
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Introduction The subgenera Romulea
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2 Literature review Leef van daad e
Page 33 and 34: Literature review Figure 2.2: Life
Page 35 and 36: Literature review Figure 2.4: Life
Page 37 and 38: Literature review The plants are sh
Page 39 and 40: Literature review flowers (DE VOS,
Page 41 and 42: Literature review often found on cl
Page 43 and 44: Literature review flowers of R. lei
Page 45 and 46: Literature review flattened upper s
Page 47 and 48: 2.2.16 Romulea tabularis Literature
Page 49 and 50: Literature review extinct, R. papyr
Page 51 and 52: Literature review R. leipoldtii occ
Page 53 and 54: Literature review Figure 2.8: Calvi
Page 55 and 56: Literature review Figure 2.14: Fras
Page 57 and 58: Figure 2.20: Nieuwoudtville weather
Page 59 and 60: Literature review Figure 2.26: Grah
Page 61 and 62: Literature review Figure 2.29: Diag
Page 63 and 64: Literature review Figure 2.30: A te
Page 65 and 66: Literature review Macronutrients ca
Page 67 and 68: Literature review The soils of the
Page 69 and 70: Literature review The embryo is the
Page 71 and 72: Literature review Phase 2 is seen a
Page 73 and 74: Literature review endogenous gibber
Page 75 and 76: Literature review tolerable level (
Page 77 and 78: Literature review these channels pe
Page 79 and 80: Literature review 2.8.7 Seed dorman
Page 81 and 82: Literature review the embryo (COPEL
Page 83: Literature review germinated under
Page 87 and 88: 2.9 BRIEF REVIEW OF IN VITRO CULTUR
Page 89 and 90: Literature review (DEBERGH, 1994).
Page 91 and 92: Literature review PIERIK (1997) sta
Page 93 and 94: Literature review The distinguishin
Page 95 and 96: Literature review The essential fun
Page 97 and 98: 2.9.3.4 Abscisic acid Literature re
Page 99 and 100: Table 2.6: Example of a matrix to e
Page 101 and 102: Literature review Young embryos req
Page 103 and 104: Literature review and the addition
Page 105 and 106: Literature review organ is then pro
Page 107 and 108: Literature review environmental str
Page 109 and 110: 2.11 IN VITRO FLOWERING Literature
Page 111 and 112: Literature review higher regenerati
Page 113 and 114: Subfamily Ixioideae (continued) Tri
Page 115 and 116: Literature review Table 2.8: Corm i
Page 117 and 118: 3 Investigation into the habitat of
Page 119 and 120: Soil sampling and analysis The firs
Page 121 and 122: Soil sampling and analysis Table 3.
Page 123 and 124: 4 Germination physiology 4.1 INTROD
Page 125 and 126: 4.2.2 Water content and imbibition
Page 127 and 128: Germination physiology (-N), phosph
Page 129 and 130: Germination physiology rosea seeds
Page 131 and 132: Germination physiology Figure 4.4:
Page 133 and 134: Germination (%) 50 40 30 20 10 0 77
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Germination (%) 100 80 60 40 20 0 c
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Germination physiology The relative
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Germination physiology seeds of R.
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5.2 MATERIALS AND METHODS In vitro
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In vitro culture initiation and mul
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5.2.3 Explant comparison In vitro c
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5.4 DISCUSSION In vitro culture ini
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6 In vitro corm formation and flowe
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In vitro corm formation and floweri
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6.3 RESULTS 6.3.1 Corm formation In
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Commercialization potential of Romu
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Commercialization potential of Romu
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BASKIN, C. C., and BASKIN, J. M. (1
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Literature cited DOWLING, P. M., CL
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Literature cited HILTON-TAYLOR, C.
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Literature cited KUMAR, A., SOOD, A
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Literature cited PIERIK, R. L. M. (
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Literature cited STEINITZ, B., COHE
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