Chapter 3: Micropropagation and in vitro phenolic production in ‘Williams’ bananas
3.3 Results and discussion
3.3.3 Effect of the aromatic cytokinins on in vitro rooting and acclimatization
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3.3.3 Effect of the aromatic cytokinins on in vitro rooting and acclimatization
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Figure 3.4: Effect of rooting compounds on the number of regenerated roots. A= meta-Topolin;
B = meta-Topolin riboside; C = meta-Methoxy topolin; D = meta-Methoxy topolin riboside; E = Benzyladenine and F = meta-Methoxy topolins 9-tetrahydropyran-2-yl.
IAA = Indole-3-acetic acid; IBA = Indole-3-butyric acid; NAA = Naphthalene acetic acid; SW = Smoke-water; KAR1 = Karrikinolide. Concentrations of root inducing compounds: IAA, IBA and NAA = 1 µM; KAR1 = 4.8 X 10-22 M and SW = 1:1000 dilution. In each graph, bars with different letter(s) are significantly different (P = 0.05) according to Duncan‟s multiple range test.
0 2 4 6 8 10 12 (A)
d cd bc cd
ab a
0 2 4 6 8 10
(B) 12
b ab
ab ab a a
0 2 4 6 8 10 12 (C)
ab ab b
a a a
0 2 4 6 8 10
(D) 12
b b a a
a a
Control IAA IBA
NAA SW KAR1
0 2 4 6 8 10 12 (E)
d
bc cd
d
a ab
Mean number of roots per shoot Mean number of roots per shoot
Rooting compounds
Control IAA IBA
NAA SW KAR1
0 2 4 6 8 10
(F) 12
c bc
ab a a ab
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Figure 3.5: Effect of rooting compounds on the length of regenerated roots. A= meta-Topolin; B
= meta-Topolin riboside; C = meta-Methoxy topolin; D = meta-Methoxy topolin riboside; E = Benzyladenine and F = meta-Methoxy topolin 9-tetrahydropyran-2-yl.
IAA = Indole-3-acetic acid; IBA = Indole-3-butyric acid; NAA = Naphthalene acetic acid; SW = Smoke-water; KAR1 = Karrikinolide. Concentrations of root inducing compounds: IAA, IBA and NAA = 1 µM; KAR1 = 4.8 X 10-22 M and SW = 1:1000 dilution. In each graph, bars with different letter(s) are significantly different (P = 0.05) according to Duncan‟s multiple range test.
0 10 20 30 40 50
(A)
a a a
a
a a
0 5 10 15
(B) 20
a a a a a
a
0 5 10 15 20 (C)
b a a a a
a
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(D) 20
a
b b b b b
Control IAA IBA NAA SW KAR1
0 5 10 15
20 (E) a
b b b b
b
Mean root length (cm) Mean root length (cm)
Rooting compounds
Control IAA IBA NAA SW KAR1
0 5 10 15
(F) 20
c bc
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ab a
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In vitro rooting is mainly controlled by the levels of both exogenous and endogenous auxins in plant tissue (De Klerk et al., 1999). Despite the absence of exogenous auxins in the shoot multiplication medium, the CK-treated plantlets still produced roots. In fact, 10 µM MemTTHP-treated plantlets had approximately 3.5-fold more roots than the control. It is possible that the addition of CKs to the medium stimulated the production of endogenous auxins in the regenerated plantlets resulting in the observed roots.
However, higher concentrations of CK in the plantlets were detrimental to rooting as observed in the drastic reduction in number of produced roots across all the treatments.
Rooting inhibition at higher concentrations for both BA and topolins in several micropropagated species is well documented (Escalona et al., 2003; Bairu et al., 2008; Valero-Aracama et al., 2010). Even though the existence of synergistic, antagonistic and additive interactions between auxin and CK (endogenous and exogenous) is well-known, the underlying mechanisms are complex and remain to be fully understood (Nordström et al., 2004).
During the rooting stage, the presence of different rooting compounds produced diverse responses in the evaluated rooting parameters of the „Williams‟ banana plantlets.
Although rooting frequency was high irrespective of the type of CK from which the shoots were obtained or the type of rooting compounds in the medium, the topolins (particularly, MemT and MemTR treatments) had higher off-shoot production frequency compared to BA-derived shoots. Clearly, the topolins maintained a stronger positive carry-over effect that resulted in production of more shoots in the rooting medium which was devoid of any exogenous CK. The production of the extra off-shoots during the rooting stage inevitably contributes to the overall shoot multiplication potency of the topolins for the micropropagation of „Williams‟ bananas.
In addition to the exogenously applied auxins, the rooting ability of in vitro plantlets is influenced by the endogenous auxin levels as well as the presence of an efficient transport mechanism (De Klerk et al., 1999; Fogaça and Fett-Neto, 2005).
Furthermore, the efficiency of different rooting compounds depends on their specific affinities for auxin receptor molecules and the concentration of free auxin reaching
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target cells (De Klerk et al., 1999). In this study, the five rooting compounds generally had no significant effect on the number of roots produced in mTR-, MemT-, MemTR- and MemTTHP-derived shoots compared to the control. In fact, some of the auxins such as IAA and NAA slightly inhibited the number and length of roots produced when compared to the control treatment. Similarly, Ascough et al. (2011) observed that the presence of different auxins (IAA, IBA and NAA) at varying concentrations inhibited root elongation in micropropagated Sisyrinchium laxum. In particular, the negative effect of NAA on rooting has been attributed to its longer persistence in plant tissue where it remains in free form and blocks root emergence (Fogaça and Fett-Neto, 2005). On the other hand when compared to the control, current findings indicate a significant increase in number of roots due to IBA, SW and KAR1 treatments in mT-derived shoots as well as IAA, SW and KAR1 treatments in BA-derived shoots. In addition to IBA and IAA which are frequently used in PTC (De Klerk et al., 1999), the auxin-like activity of SW and KAR1 is a further indication of the great potential of both compounds as a substitute for commonly used rooting compounds. Evidence demonstrating the auxin- and CK-like activity of both SW and KAR1 in PTC have been reported (Jain et al., 2008). A significant increase in root length was achieved with the use of KAR1 on BA- and MemTTHP-derived shoots.