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Thidiazuron and other plant bioregulators for axenic culture of Siam

cardamom (Amomum krervanh Pierre ex Gagnep.)

Article  in  Kasetsart Journal - Natural Science · January 2012

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 Kasetsart J. (Nat. Sci.) 46 : 335 - 345 (2012)

Thidiazuron and Other Plant Bioregulators for Axenic Culture

of Siam Cardamom (Amomum krervanh Pierre ex Gagnep.)

Surawit Wannakrairoj1,* and Wondyifraw Tefera2

ABSTRACT

Siam cardamom, one of the most common spice and medicinal plants in Southeast Asia, is
usually propagated by vegetative means. Identification of suitable plant bioregulators for high shoot
proliferation and growth rate is one of the most important steps for an effective micropropagation protocol.
The axillary bud from the rhizome of Siam cardamom could be used as an explant for micropropagation.
High axillary shoot proliferation (mean = 10.25 shoots per explant) and plant growth rate were attained
using a modified Murashige and Skoog (MS) medium with 0.5 mg.L-1 thidiazuron (TDZ) and 2 mg.L-1
imazalil. An MS medium supplemented with 0.75 mg.L-1 TDZ and 3.0 mg.L-1 6-benzyladenine also
enhanced shoot multiplication (8.45 shoots per explant). Proliferated shoots successfully elongated and
rooted when transferred to the basal MS medium. Inclusion of paclobutrazol to the medium exerted a
negative effect on the growth and development of the Siam cardamom.
Keywords: 6-benzyladenine, imazalil, micropropagation, paclobutrazol, Zingiberaceae

INTRODUCTION related problems from conventional propagation

Siam cardamom (Amomum krervanh),
also known as ‘best cardamom’ or ‘krawan’, is a
common spice and medicinal plant species native
to Thailand and Kampuchea, is a member of the
Zingiberaceae family and its seed has been used
as a best substitute for the Indian cardamom
(Elettaria cardamomum) in the world market
(Purseglove et al., 1981). Conventionally, Siam
cardamom is propagated through rhizome
division (Paisooksantivatana, 1996). However,
the technique imposes considerable loss of
potential productive plant stands. In many
instances, micropropagation is believed to be
the best alternative to alleviate this and other
techniques, as it also enables rapid true-to-type
multiplication of an elite clone within a relatively
short time (Prathanturarug et al., 2003).
Development of an efficient
micropropagation protocol requires the optimization
of plant bioregulator supplements. The type and
concentration of plant bioregulators affect the
capacity of in vitro propagation because they play
a major role in cell division, differentiation and
morphogenesis in plant tissue cultures (Toteva
et al., 2000). Axillary bud outgrowth, considered
a process of apical dominance release, could
hence be enhanced in response to exogenous
cytokinins. Cytokinins are a group of plant
bioregulators that play a major role in cell division

1 Department of Horticulture, Faculty of Agriculture, Kasetsart University, Bangkok 10900, Thailand.

2 Plant Biotechnology Division, Jimma Agricultural Research Center (JARC), Ethiopian Institute of Agricultural Research
(EIAR), P.O. Box 1161, Jimma, Ethiopia.
* Corresponding author, e-mail: agrsuw@ku.ac.th

Received date : 06/12/11 Accepted date : 24/04/12

336 Kasetsart J. (Nat. Sci.) 46(3)
and cell differentiation, inducing cell division and
organogenesis in plant cell cultures and affecting
many other physiological and developmental
processes in plants (Kamínek et al., 1997).
Thidiazuron (TDZ), a phenylurea derivative, is
reported to be much more effective in axillary
bud break and adventitious shoot proliferation
in different plant species than the commonly
employed adenine-based cytokinins (Huetteman
and Preece, 1993; Toteva et al., 2000; Faisal et al.,
2005). According to Murthy et al. (1998), various
reports stated the effects of TDZ to be related to
its modulation of plant hormones, either directly
or indirectly because of induced stress. Different
workers (Nielsen et al., 1995; Murthy et al., 1998)
have also reported further improvement in the
efficacy of TDZ through its combined use with some
other plant bioregulators.
The effects of TDZ on shoot proliferation
have been reported on turmeric (Salvi et al., 2000;
Prathanturarug et al., 2003) and korarima (Tefera
and Wannakrairoj, 2004a and 2006). In addition,
Werbrouck and Debergh (1996, 1997) reported the
influence of imazalil (IMA) in enhancing multiple
shoot producing effect of 6-benzyladenine (BA)
and TDZ. Tefera and Wannakrairoj (2006)
also reported a synergistic effect between TDZ
and paclobutrazol (PBZ) on multiplication of
korarima. Nevertheless, no in-depth study has yet
been undertaken to investigate the interactive effects
of TDZ with other plant bioregulators in Siam
cardamom. Thus, the current study was undertaken
to investigate the effects of TDZ in combination with
BA, IMA and /or PBZ on in vitro shoot proliferation
and development of Siam cardamom.
MATERIALS AND METHODS
Plant material and culture conditions
Siam cardamom rhizomes were planted
in a nursery in Kasetsart University, Thailand.
Pieces of rhizomes with sprouting axillary buds
(3–5mm) were collected and some scale leaves
were removed prior to cleaning. The buds with
the rhizome pieces were washed with a laboratory
detergent and then kept under running tap water
for 90 min. The axillary buds were excised and
washed. Outer scales were removed from the
buds and were rinsed with 70% ethanol for 1 min
under aseptic conditions. Further disinfestations
were carried out aseptically using 20 and 10%
Hyter® (active ingredient 6% sodium hypochlorite
v/v) added with 2 mL.L-1 Tween-80 for 10 and 5
min, respectively. Explants were then rinsed four
times using sterilized distilled water and further
trimmed before transfer to the culture tubes. Single
disinfested explants were cultured on modified
Murashige and Skoog (1962) (MS) medium for
culture establishment. Further multiplication of
explants was attained using MS medium added
with 5% coconut water (CW) and 3 mg.L-1 BA
and 1 mg.L-1 kinetin for production of stock
cultures (Tefera and Wannakrairoj, 2004b). The
stock cultures were then subsequently transferred
and maintained on the basal MS medium for
45 d until the commencement of the respective
experiments.
Three experiments were carried out. In
the first experiment, the effect of adding 0, 0.25, 0.5
and 0.75 mg.L-1 thidiazuron (TDZ) was evaluated
with 0, 1.5, 3 and 4.5 mg.L-1 6-benzyladenine
(BA). In the second experiment, the influence of
TDZ at the same four concentrations with 0, 2, 4
and 8 mg.L-1 imazalil (IMA) was determined in
a factorial combination. In the third experiment,
these same levels of TDZ combined with 0,
1.5, 3 and 4.5 mg.L-1 paclobutrazol (PBZ) were
evaluated.
In all instances, plant bioregulators and
30 g.L-1 table sugar were added to the media prior
to autoclaving. The media were gelled with 7 g.L-1
agar-agar after adjusting the pH to 5.7 with 1 N
KOH or 1 N HNO3. The molten media of 30 mL
was dispensed into 100 mL baby food jars and
autoclaved at 121 oC at 1.06 kg.cm-2 pressure
for 20 min. Cultures were maintained at 25 ±
 Kasetsart J. (Nat. Sci.) 46(3) 337

2 oC under a 16-h photoperiod of 28 μmol.m-2
irradiance provided by cool white fluorescent
tubes.
Statistical analyses
Experiments were set up in a completely
randomized design with 10 replications and
repeated at least three times. Only data from
the last two repetitions were used for statistical
analyses. In each experiment, observations on the
number of shoots, the shoot height and dry weight
of plantlets were recorded after 2 mth of culture.
Data were analyzed using the general linear model
(GLM) using a PC-SAS program (Version 8e, SAS
Institute Inc.; Cary, NC, USA). Treatment means
were separated using Duncan’s multiple range test
(DMRT) with significance tested at the P < 0.05
level.
RESULTS AND DISCUSSION
Effects of thidiazuron (TDZ) and 6-benzyladenine
(BA)
The shoot number and height, as well as
the dry weight of Siam cardamom plantlets were
strongly enhanced by the inclusion of TDZ and
BA in the culture medium. For TDZ, the shoot
number increased as the concentration of TDZ
increased, though no significant differences were
observed between 0.5 and 0.75 mg.L-1 TDZ. The
highest plantlet dry weight was obtained from 0.5
mg.L-1 TDZ. For BA, the use of 3.0 mg.L-1 BA
gave both the highest number of shoots and dry
weight of plantlets. The shoot height was observed
to decrease consistently with increasing levels of
each of the two bioregulators (Table 1).
The combined use of TDZ and BA had an
interactive effect upon all the parameters evaluated.
The highest number of shoots per explant (mean
= 8.45) was obtained from the medium with 0.75
mg.L-1 TDZ and 3 mg.L-1 BA. No significant
differences were observed with regard to plantlet
dry weight when 0.5 or 0.75 mg.L-1 TDZ was
used in combination with 3 mg.L-1 BA. The tallest
shoots (mean = 7.02 cm) were obtained from the
basal medium (Table 2). The shoots produced on a
medium supplemented with 0.75 mg.L-1 TDZ and
3.0 mg.L-1 BA were of high quality (Figure 1a) that
easily elongated and produced roots upon transfer
onto a medium devoid of plant bioregulators.
The significantly higher efficacy of
TDZ observed in this study could be associated

Table 1 Siam cardamom mean shoot growth after 8 wk of culture on media containing thidiazuron
(TDZ) and 6-benzyladenine (BA).
Plant bioregulator No. of Shoots Shoot length (cm) Dry weight (mg)
TDZ (mg.L ) -1

0 3.28c 4.96a 76.51c

0.25 5.00b 3.20b 87.76b
0.5 5.81 a 2.62 c 94.55a
0.75 5.98a 2.72c 88.06b
BA (mg.L )-1

0 4.26c 5.00a 82.66c

1.5 5.14 b 3.17 b 90.16b
3.0 6.42a 2.65c 97.19a
4.5 4.22 c 2.22 d 76.74d
% CV 27.87 22.88 15.64
Means within a column followed by the same lowercase letter are not significantly different at P < 0.05 based on Duncan’s
multiple range test.
338 Kasetsart J. (Nat. Sci.) 46(3)

with the high potency of its cytokinin action. The
biological activity of TDZ was stated to be higher
than or comparable to that of most active adenine-
type cytokinins at unusually low concentrations
(Mok et al., 1987; Murthy et al., 1998; Rai, 2002;
Stoeva, 2002; Mithila et al., 2003). This could
again be ascribed to its counteracting effect on
the usual shortage of cytokinin in plant tissues,
which results from their degradation by cytokinin
oxidase (Mok et al., 1987; Hare and van Staden,
1994). In this regard, TDZ is much more resistant
to oxidases, is stable and biologically active at
lower concentrations than adenine-type cytokinins
(Mok et al., 1987).
The use of TDZ in the culture medium
effectively enhanced shoot proliferation of Siam
cardamom. However, the growth and development
of the shoots were much more enhanced when BA
was added to the TDZ-supplemented medium.
This finding concerning the interactive effects
from the combined use of BA and TDZ is in direct
agreement with other reports that described the
benefits from the combined use of these two plant
bioregulators (Khalafalla and Hattori, 1999; Tefera
and Wannakrairoj, 2006).
With regard to shoot height, several
authors (Murthy et al., 1998; Arinaitwe et al.,
2000; Roussos and Pontickis, 2002; Haensch,
2004) reported the suppression of shoot elongation
due to the use of TDZ in the culture medium.
Tiwari et al. (2001) also obtained large shoot buds
from BA-supplemented media, while the use of
TDZ resulted in clusters of stunted shoots. These
findings were in agreement with the results from
the present study. In line with this, Nielsen et al.
(1995) reported stimulation of shoot proliferation
from the use of cytokinins to be traded off with
inhibition of their elongation.

Table 2 In vitro growth and development (mean ± SE) of Siam cardamom after 8 wk of culture on
media containing thidiazuron (TDZ) and 6-benzyladenine (BA).
TDZ BA No. of Shoots Shoot length Dry weight
-1
(mg.L ) -1
(mg.L ) (cm) (mg)
0 0 1.80j ± 0.16 7.02a ± 0.16 61.54g ± 3.03
0 1.5 3.40i ± 0.36 5.28b ± 0.30 80.65efg ± 2.91
0 3 ghi
4.15 ± 0.17 c
4.32 ± 0.29 89.20b-e ± 2.16
0 4.5 3.75hi ± 0.19 3.24d ± 0.21 74.66g ± 4.55
0.25 0 e-h
4.61 ± 0.33 b
5.62 ± 0.15 88.22cde ± 2.92
0.25 1.5 4.90d-g ± 0.27 3.22d ± 0.23 90.82bcd ± 2.05
0.25 3 5.65cd ± 0.34 gf
2.09 ± 0.10 94.10a-d ± 5.72
0.25 4.5 4.80d-g ± 0.26 2.11gf ± 0.09 77.96fg ± 4.57
0.5 0 def
5.25 ± 0.30 c
4.68 ± 0.09 96.26abc ± 2.50
0.5 1.5 6.55c ± 0.29 2.15gf ± 0.11 98.62ab ± 1.29
0.5 3 b
7.45 ± 0.63 gf
1.92 ± 0.05 103.16a ± 2.27
0.5 4.5 4.00ghi ± 0.30 1.71g ± 0.07 80.15efg ± 1.31
0.75 0 de
5.40 ± 0.34 e
2.75 ± 0.21 85.16def ± 2.14
0.75 1.5 5.70cd ± 0.26 2.03gf ± 0.12 90.56bcd ± 2.12
0.75 3 a
8.45 ± 0.31 f
2.27 ± 0.10 102.32a ± 1.77
0.75 4.5 4.35f-i ± 0.22 1.83gf ± 0.08 74.19g ± 3.39
Means within a column followed by the same lowercase letter are not significantly different at P < 0.05 based on Duncan’s
multiple range test.
 Kasetsart J. (Nat. Sci.) 46(3)
This phenomenon could be attributed
to the effect of TDZ in inducing accumulation
of endogenous cytokinin (Murthy et al., 1998;
Arinaitwe et al., 2000). It could also be due to the
comparatively higher cytokinin activity of TDZ
even at a lower concentration (Arinaitwe et al.,
2000). Either way, the resulting higher cytokinin
activity is known to have a negative impact upon
shoot elongation (Murthy et al., 1998; Roussos
and Pontickis, 2002). However, TDZ-induced
shoot stunting was not a permanent phenomenon
and could be reversed by transferring the explants
to media that contained BA or were PGR-free
(Arinaitwe et al., 2000; Rai, 2002; Tefera and
Wannakrairoj, 2006). In the current experiment
using Siam cardamom, shoot elongation and
rooting were also successfully realized by
subculturing the TDZ-induced multiple shoots to
a plant bioregulator-free MS medium.
Effects of thidiazuron and imazalil (IMA)
The inclusion of TDZ and IMA to the
culture medium considerably affected all the
parameters evaluated in this study. Siam cardamom
shoot number showed a significant increase with
the inclusion of TDZ to the culture medium. This
Table 3 Siam cardamom shoot growth after 8 wk of culture on media containing thidiazuron (TDZ)
and imazalil (IMA).
Plant bioregulator No. of Shoots Shoot length (cm)
TDZ (mg.L ) -1
0 2.96c
0.25 5.88b
0.5 7.78 a
0.75 5.62b
IMA (mg.L )-1
0 4.19c
2 7.32 a
4 6.06b
8 4.71 c
% CV 37.76
Means within a column followed by the same lowercase letter are not significantly different at P < 0.05 based on Duncan’s
multiple range test.
339
same trend was observed for the dry weight of
plantlets, except at 0.75 mg.L-1 which gave the
least value. On the contrary, shoot height showed
a consistent decrease at all levels of TDZ studied
in the experiment. The use of IMA in the culture
medium also improved the number of shoots and
dry weight of plantlets, as well, while it reduced
shoot height (Table 3).
The combined use of these two
bioregulators resulted in a significant interactive
effect on the number and height of Siam cardamom
shoots, as well as the dry weight of plantlets.
The highest shoot number (mean = 10.25 shoots
per explant) and dry weight of plantlets (mean =
104.18 mg per explant) were obtained from the
medium with 0.5 mg.L-1 TDZ and 2 mg.L-1 IMA.
No significant variation in shoot number or shoot
height was observed between the different levels
of IMA used in this experiment, when the media
were devoid of TDZ (Table 4). Shoots obtained
from the combined use of TDZ and IMA were
relatively dwarfed in size (Figure 1b). The shoots
obtained from the media could elongate and root
easily after one to two subcultures on a basal MS
medium. On the other hand, the plant bioregulator-
free medium produced well-elongated shoots that
Dry weight (mg)
7.20a 79.48b
2.63b 74.34c
2.05 c 87.62a
1.29d 73.01c
3.84a 90.50a
3.12 b 91.70a
3.14b 75.31b
2.87 b 56.10c
25.54 14.43
340 Kasetsart J. (Nat. Sci.) 46(3)

had a number of functional roots (Figure 1c).
Similar to the current results, enhanced
shoot multiplication from a TDZ-supplemented
medium has been observed in different herbaceous
plant species including banana (Arinaitwe et
al., 2000), Curcuma longa (Salvi et al., 2000;
Prathanturarug et al., 2003) and Aframomum
corrorima (Tefera and Wannakrairoj, 2006). This
could be attributed to the high cytokinin activity of
TDZ which could be associated with the induction
of the synthesis or accumulation of endogenous
cytokinins (Hutchinson and Saxena, 1996).
Werbrouck and Debergh (1996, 1997)
reported the influence of IMA in enhancing the
multiple-shoot-producing effect of cytokinins,
including BA and TDZ. Inducing changes in the
metabolism of exogenously applied cytokinins
was suggested as the possible mechanism for the
enhancement of cytokinin actions observed from
the use of IMA in the culture medium. However,
no cytokinin effects were observed from the sole
use of IMA in the culture medium.
As reviewed by Fletcher et al. (2000),
certain azole compounds interfere with the
biosynthesis of gibberellins and influence the
morphogenesis of plants, indicating their possible
use as plant bioregulators. Werbrouck and
Debergh (1996) reported that imadizol (IMA)
fungicide, shared a similar structural feature with
triazoles, that is, a hetrocyclic ring containing an
sp2-hybridized nitrogen with a lone electron pair,
a common feature of molecules that can inhibit
ergosterol biosynthesis in fungi and can block
gibberellins (GA) synthesis in plants. This impact
of the chemicals on the in planta GA level was
thought to cause the subsequent cytokinin effects
observed from their use (Fletcher et al., 2000).
This could be the reason why the shoot height was
reduced with the use of IMA, instead of using BA
together with TDZ (Figures 1A and 1B).

Table 4 In vitro growth and development (mean ± SE) of Siam cardamom after 8 wk of culture on
media containing thidiazuron (TDZ) and imazalil (IMA).
TDZ IMA No. of Shoots Shoot lg. Dry wt
-1
(mg.L ) -1
(mg.L ) (cm) (mg)
0 0 2.56i ± 0.24 7.66a ± 0.28 88.84b ± 2.32
0 2 3.06hi ± 0.26 7.74a ± 0.38 78.64c ± 2.90
0 4 hi
3.50 ± 0.29 a
7.61 ± 0.16 76.31c ± 3.09
0 8 2.72hi ± 0.16 7.75a ± 0.26 74.02cd ± 3.14
0.25 0 ghi
3.90 ± 0.12 b
3.71 ± 0.22 90.69b ± 1.35
0.25 2 7.90bc ± 0.72 2.68c ± 0.15 91.26b ± 1.62
0.25 4 de
6.25 ± 0.35 cde
2.18 ± 0.18 66.16ef ± 2.45
0.25 8 5.44ef ± 0.74 1.87def ± 0.18 46.48g ± 4.52
0.5 0 efg
5.12 ± 0.35 cde
2.18 ± 0.16 94.12b ± 2.71
0.5 2 10.25a ± 0.63 2.29dc ± 0.16 104.18a ± 3.32
0.5 4 b
8.70 ± 0.75 cde
2.09 ± 0.15 91.19b ± 2.54
0.5 8 6.39de ± 0.52 1.61efg ± 0.13 59.48f ± 2.80
0.75 0 efg
5.33 ± 0.11 efg
1.66 ± 0.09 88.71b ± 2.09
0.75 2 7.22cd ± 0.68 1.30fg ± 0.11 89.96b ± 2.49
0.75 4 ef
5.55 ± 0.37 g
1.12 ± 0.15 67.68de ± 1.59
0.75 8 4.25fgh ± 0.59 1.06g ± 0.16 42.96g ± 2.01
Means within a column followed by the same lowercase letter are not significantly different at P < 0.05 based on Duncan’s
multiple range test.
 Kasetsart J. (Nat. Sci.) 46(3) 341

Figure 1 In vitro shoot multiplication of Siam cardamom: (A) Shoots from a Murashige and Skoog (MS)
medium, supplemented with 0.75 mg.L-1 thidiazuron (TDZ) and 3.0 mg.L-1 6-benzyladenine;
(B) Shoots from 0.5 mg.L-1 TDZ and 2 mg.L-1 imazalil, supplemented MS medium; (C)
A shoot from 0.5 mg.L-1 TDZ and 3 mg.L-1 paclobutrazol, supplemented MS medium; (D)
Shoots from an MS basal medium (control).

Effects of thidiazuron and paclobutrazol
The use of TDZ in the culture medium
enhanced the production of Siam cardamom
shoots, but reduced the shoot height and dry weight
of plantlets. However, the addition of PBZ to the
culture medium exerted a negative effect upon the
shoot proliferation and growth of Siam cardamom
(Table 5).
The combined use of these two plant
bioregulators also showed an interactive effect on
the shoot number. In most cases, PBZ was toxic for
the growth and development of Siam cardamom
shoots (Figure 1C). Thus, the highest numbers of
shoots (mean = 5.89 and 5.50 per explant) were
obtained from media supplemented with 0.5 and
0.75 mg.L-1 TDZ, respectively, but both without
342 Kasetsart J. (Nat. Sci.) 46(3)

PBZ. The plant bioregulator-free medium gave
the tallest shoots (mean = 7.20 cm) and plantlets
with the highest dry weight (mean = 78.17mg)
as shown in Table 6. The negative effects of
PBZ observed from this experiment with Siam
cardamom were in contrast to the findings of
Tefera and Wannakrairoj (2006). They were also
contrary to the comprehensive reviews of Davis
et al. (1988) and Fletcher et al. (2000), that stated
several benefits of this particular plant bioregulator
including reduced hyperhydricity and increased
cytokinin effects. All these inconsistencies could
be ascribed to the differences in the response of
Siam cardamom and the other species studied so
far.
CONCLUSION
In the present study, a high rate of shoot
proliferation was obtained in Siam cardamom
from the inclusion of TDZ in the culture medium.
The use of another plant bioregulator supplement
to the culture medium together with TDZ
further enhanced the shoot proliferation of Siam
cardamom. Therefore, TDZ at 0.5 mg.L-1 gave
the highest rate of shoot proliferation (mean =
10.25 shoots per explant) when combined with 2
mg.L-1 IMA in the culture medium. The highest
plantlet dry weight was obtained from this same
medium. The use of 0.75 mg.L-1 TDZ and 3.0
mg.L-1 BA in the culture medium also gave more
than eight shoots of Siam cardamom every 2 mth.
However, the highest dry weight (mean = 103.16
mg per explant) of plantlets was obtained from the
medium with 0.5 TDZ and 3.0 mg.L-1 BA. Shoot
elongation and rooting were easily attained when
explants were transferred to plant bioregulator-
free MS medium. The high multiplication rate of
plantlets having no detectable phenotypic variation
among the regenerants makes this protocol suitable
for large-scale propagation of this culinary plant
species of high medicinal importance. On the other
hand, the inclusion of PBZ in the culture medium,
with or without TDZ, retarded the growth and
development of Siam cardamom.

Table 5 Siam cardamom shoot growth after 8 wk of culture on media containing thidiazuron (TDZ)
and paclobutrazol (PBZ).
Plant bioregulator No. of Shoots Shoot length (cm) Dry weight (mg)
TDZ (mg.L-1)
0 2.06c 3.64a 51.10a
0.25 2.46 b 2.57 b 43.09b
0.5 2.71a 2.08c 40.81b
0.75 2.32 b 1.60 d 34.63c
PBZ (mg.L-1)
0 4.77a 3.79a 58.47a
1.5 1.75b 2.22b 38.90b
3.0 1.60 bc 1.91 c 36.55b
4.5 1.41c 1.89c 34.95b
TDZ × PBZ *** *** **
% CV 29.65 30.62 38.44
Means within a column followed by the same lowercase letter are not significantly different at P < 0.05 based on Duncan’s
multiple range test.
*** = significantly different at P < 0.01; ** = significantly different at P < 0.05.
 Kasetsart J. (Nat. Sci.) 46(3) 343

Table 6 In vitro growth and development (mean ± SE) of Siam cardamom after 8 wk of culture on
media containing thidiazuron (TDZ) and paclobutrazol (PBZ).
TDZ PBZ No. of Shoots Shoot length Dry weight
-1
(mg.L ) -1
(mg.L ) (cm) (mg)
0 0 2.89c ± 0.20 7.20a ± 0.50 78.17a ± 8.82
0 1.5 2.06d ± 0.15 2.59c ± 0.07 43.65cd ± 3.83
0 3 1.70def ± 0.11 2.58c ± 0.05 44.09cd ± 3.14
0 4.5 efg
1.50 ± 0.16 def
1.96 ± 0.16 36.49de ± 5.61
0.25 0 4.79b ± 0.29 3.29b ± 0.05 60.64b ± 3.56
0.25 1.5 de
1.78 ± 0.15 c
2.57 ± 0.22 40.80cde ± 2.97
0.25 3 1.53d-g ± 0.14 1.99de ± 0.11 35.13de ± 2.65
0.25 4.5 def
1.67 ± 0.14 cd
2.42 ± 0.04 35.23de ± 2.88
0.5 0 5.89a ± 0.24 2.28cd ± 0.10 51.43bc ± 4.34
0.5 1.5 def
1.74 ± 0.13 cd
2.28 ± 0.20 37.49de ± 3.06
0.5 3 2.06d ± 0.15 1.67efg ± 0.11 34.99de ± 2.94
0.5 4.5 fg
1.22 ± 0.10 ecd
2.08 ± 0.14 39.50cde ± 2.64
0.75 0 5.50a ± 0.23 2.40cd ± 0.23 43.53cd ± 3.27
0.75 1.5 efg
1.44 ± 0.12 fg
1.44 ± 0.09 33.73de ± 1.78
0.75 3 1.11g ± 0.08 1.43fg ± 0.09 32.47de ± 2.34
0.75 4.5 efg
1.26 ± 0.10 g
1.14 ± 0.08 29.08e ± 2.39
Means within a column followed by the same lowercase letter are not significantly different at P < 0.05 based on Duncan’s
multiple range test.

ACKNOWLEDGEMENTS Faisal, M., N. Ahmad and M. Anis. 2005. Shoot

The authors would like to acknowledge
the Agricultural Research and Training Project
(ARTP) of the Ethiopian Agricultural Research
Organization (EARO) for the financial support
to undertake this research and thank Associate
Professor Yingyong Paisooksantivatana for
making available literature and the provision of
the Siam cardamom planting material.
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