American Journal of Plant Sciences, 2011, 2, 669-674
doi:10.4236/ajps.2011.25080 Published Online November 2011 (http://www.SciRP.org/journal/ajps)
669
Effect of Vitamins on In Vitro Organogenesis of
Plant
Peter Abrahamian, Arumugam Kantharajah*
Department of Agricultural Sciences, American University of Beirut, Riad El Solh, Beirut, Lebanon.
Received June 10th, 2011; revised July 25th, 2011; accepted September 1st, 2011.
ABSTRACT
Vitamins are necessary compounds synthesized and utilized in plants. In tissue culture media, vitamin addition is not
always common; since the amount needed by plants is relatively unknown and varies. Vitamins, in combination with
other media constituents, have been shown to have direct and indirect effects on callus growth, somatic growth, rooting,
and embryonic development. For example, different studies have shown that thiamine is associated with cytokinin and
has a role in inducing callus growth and rooting. Moreover, thiamine was essential in facilitating the production of
more secondary metabolites such as proteases in pineapple. Both biotin and riboflavin play a role in callus develop-
ment as well. Specifically, riboflavin exerts different effects on plant rooting either positively and negatively. Vitamin D
known to cause uptake of calcium in animal tissue, exerts a similar effect in plants. In addition, vitamin D causes cell
elongation and meristematic cell division. Vitamin C, known for its anti-oxidative properties, has also enhanced shoot
growth and rooting.
Keywords: Vitamin, Organogenesis, In Vitro, Plant Tissue Culture, Plant Propagation
1. Introduction
Plants are a major source of essential vitamins for hu-
mans and animals. Their function and synthesis pathways
have been extensively studied. Vitamin syntheses in plants
are mainly used as essential intermediates in biochemical
reactions or as catalysts in various pathways. Vitamins are
divided into two main groups, the water-soluble (Ascor-
bic acid “C”; thiamine “B1”; riboflavin “B2”; pyridoxine
“B6”; nicotinic acid; cobalamin “B12”; folic acid; pan-
tothenic acid “B5”; biotin) and fat-soluble (A, D, E, K)
vitamins [1]. According to Bonner [2], working on water-
soluble vitamins is of higher interest than fat-soluble vi-
tamins. In tissue culture, some plants can become defi-
cient in vitamin synthesis [3]. Hence, supplementing plant
tissue with sub-optimal levels is essential to obtaining
vigorous growth. Plant cell requirements for vitamin con-
centration vary according to the plant species and type of
culture.
Thiamine pyrophosphate (TPP) is a derivative of Thia-
mine (Vit. B1) [1]. Thiamine’s physiological functions in
plants are diverse and serve as cofactors in enzymatic
reactions including pentose phosphate pathway, glycoly-
sis, tricarboxylic acid cycle (TCA), pyruvate dehyrdro-
genase complex, transketolase, and pyruvate decarboxy-
lase [4]. Pyruvate decarboxylase has shown to be im-
perative in energy production in Arabidopsis [5]. Thia-
mine has also been associated with disease resistance,
and expression of PR-1 gene with local acquired resis-
tance, but not systemic acquired resistance (SAR) [6],
however, Ahn et al. [7] showed induced SAR in Arabi-
dopsis. Under conditions of abiotic stress in Arabidopsis,
endogenous thiamine increases dramatically to cope with
oxidative stresses by supplying NADH and NADPH [8].
Vitamin C or ascorbate is oxidized by oxygen, hydrogen
peroxides, and superoxides into monodehydroascorbate
(MDHA) radicals. Ascorbate oxidase is possibly related
to cell wall expansion and growth. MDHA, a product of
ascorbate oxidase, radicals obtained depolarize the pla-
sma membrane hence causing ion uptake and wall loo-
sening [9]. Other vitamins such as riboflavin, a precursor
of FAD and FMN coenzymes, and nicotinic acid, pre-
cursor of NAD and NADP, participate in cellular redox
reactions. In this review paper, we will provide a basic
summary of how these pathways are exhibited, at the
macro level, upon vitamin addition to plant tissue culture
media.
2. Vitamins in Tissue Culture
In tissue culture media, thiamine, nicotinic acid, pyri-

Effect of Vitamins on In Vitro Organogenesis of Plant 670
doxine and myo-inositol found in Murashige and Skoog
[10] (MS) medium at 0.1 mg·l–1, 0.5 mg·l–1, 0.5 mg·l–1,
and 100 mg·l–1 respectively are the most commonly used,
while the addition of other essential vitamins to media is
uncertain. Myo-inositol remains a controversial compound
being either classified as a water-soluble plant vitamin or
as a sugar alcohol [3]. Earlier studies in pea embryos
done by Ray [11] have shown that it is possible to achi-
eve good in vitro growth by increasing vitamin C content.
This finding cannot be broadly applied as some plants are
less receptive to increasing concentrations of vitamin C,
indicating more autotrophism than other plants (tomato
and oat) [2]. It has also been noticed that adding biotin
increased the shoot dry weights of peas, similar to the
response observed in Ricciocarpus plants treated with
pantothenic acid. Unlike other vitamins, thiamine addi-
tions to pea embryos in vitro affect rooting and shoot
growth simultaneously. In vitro studies have shown that
tomato roots are capable of exhibiting prolonged thia-
mine dependency. [2].
2.1. Micropropagation
In the presence of 25 mg·l–1 of vitamin D3 micro propa-
gated potato plantlets absorbed Ca2+ efficiently [12].
However, vitamin D3 concentrations higher than 25 mg·l–1
i.e. 50 mg·l–1 did not stimulate higher absorption levels.
On the other hand vitamin D2 suppressed Ca2+ uptake. It
was concluded that combining both vitamins D2 and D3
did not improve calcium absorption hence claiming the
superiority of vitamin D3 for calcium ion uptake [12].
2.2. Callus & Somatic Growth
Gamborg et al. [13] cultured soybean root cells unto
several media containing different concentrations of thia-
mine, and to a complete B5 culture media. An initial
amount of 53 mg of soybean cell culture was grown in 0
mg·l–1 and 10 mg·l–1 of thiamine. After 5 days, 138 mg
and 203 mg of soybean cells were produced, respectively.
Pyridoxine, nicotinic acid and myo-inositol present in the
media had no adverse effects on growth individually.
Consequently, Gamborg et al. [13] concluded the neces-
sity of providing thiamine to the media to sustain growth
of soybean root cell.
Eriksson [14] have concluded that nicotinic acid and
pyridoxine are essential vitamins accompanying thiamine,
when studying the optimum growth of Haplopappus gra-
cilis Nutt. on a modified medium of MS [10].
Polikarpochkina et al. [15] reported that maize calli
decreased in weight from 110 mg/ml to nil after 3 suc-
cessive passages when thiamine is eliminated. However,
the removal of inositol and pyridoxine from the MS [10]
medium did not give any significant difference on gr-
owth [15]. The change recorded from the first passage
until the third passage, when omitting inositol and pyri-
doxine was 9% and 2.5%, respectively.
Digby and Skoog [16] found a relation between ki-
netin and thiamine in the normal callus culture of tobacco.
It has been shown that high levels of kinetin are needed
to induce thiamine synthesis in the absence of any ex-
ogenous thiamine added. However, sustaining growth on
a low level kinetin media was not possible except if
thiamine was added. Whereas, Linsmaier and Skoog [17]
maintained tobacco cultures with 1000 μg/l of kinetin
and nil thiamine over 17 passages. Dravnieks et al. [18]
later confirmed that thiamine synthesis was subject to
feedback control mechanism thus sensitive to the amount
of thiamine in tissue, regardless of kinetin concentration.
Both thiamine and biotin significantly affected callus
growth of date palm [19]. Increasing thiamine from 0.1
mg·l–1 to 0.5 mg·l–1 caused maximum callus growth; fur-
thermore, increasing thiamine to 2 mg·l–1 gave reduced
callus weights. Moreover, increasing biotin from 0 to 1
mg·l–1 gave a maximum callus weight similar to thiamine
[19]. On the other hand, an earlier report by Drew and
Smith [20] showed that presence of riboflavin reduced
callus growth of Papaya. A significant decline in mean
callus weight was recorded from 89.32 mg to 0.10 mg
per explant, in the absence and presence of riboflavin,
respectively [20].
Ascorbic acid, functioning primarily as an antioxidant,
is used to prevent browning of tissue [1,3]. However, in
tobacco cells, ascorbic acid has been shown to function
as a stimulant of mitotic cell division [21].
2.3. Rooting
Vitamin D3 stimulates rooting of Phaseolus vulgaris L. in
culture [22]. In a control treatment without any vitamins
43.75% of the roots were longer than 14 mm, while vi-
tamin D3 addition achieved 78.75%. The effect of vita-
min D3 shown by Boland et al. [22] at 10–9 M, on root
growth was associated with an uptake of calcium ions, an
increase in cell elongation in root zone at 0.5 - 1 mm
from the apex, and stimulation of mitotic division of
meristematic cells.
In vitro rooting of peach rootstock GF677 (Prunus
amygdalus × P. persica Batsch.) was studied by adding
different concentrations of riboflavin ranging from 0 to
2.0 mg·l–1 [23]. As more riboflavin was added rooting
decreased in a linear form until it was completely inhi-
bited. The smallest concentration of 0.5 mg·l–1 of ribofla-
vin caused the average number, length, fresh weight and
dry weight to decrease sharply [23]. Whereas at 1.5 and 2
mg·l–1 of riboflavin chlorotic and necrotic symptoms
appeared. Moreover, adventitious root formation in the
control was long and thin, while in the treated media,
roots were short and thick. Also, callus formation was

Effect of Vitamins on In Vitro Organogenesis of Plant 671
inhibited in the rooting MS media, due to the suppressing
action of auxin by photo-degradation [23].
On the contrary, riboflavin has been shown to stimu-
late and help rooting significantly [24-26]. Rooting in
apple tissue culture was studied in the presence of ribo-
flavin. In the dark riboflavin stimulated rooting signifi-
cantly in the presence of auxin (IBA), whereas rooting
decreased when the vitamin was omitted and exposed to
light [24]. Trindade and Pais [25] showed that Eucalyp-
tus globulus Labill. produced 80% rooting ability on a
revised De Fossard [27] media containing riboflavin (Ta-
ble 1). On the other hand, 60% rooting was achieved on
the same media excluding the latter compound [25].
Carica papaya L. rhizogenesis was optimal when 31 μM
of riboflavin and 10 μM of IBA were added to the De
Fossard [27] media in the dark for 2 days, but losses oc-
curred during media preparation [26]. However, Drew et
al. [26] found a way to avoid the loss of IBA due to light
exposure. The procedure involved injecting riboflavin at
300 μM per ml into 10 ml of media, which is equivalent
to the optimum riboflavin level 31 μM, after 1 day of
IBA rich medium in the light [26].
Thiamine is another vitamin shown to have significant
rooting on pacific yew, an evergreen, Taxus brevifolia
Peattie [28]. Upon adding thiamine, Chee et al. [28] ob-
tained 61.5% of adventitious rooting in T. brevifolia Pea-
ttie compared to 30% without thiamine. In a literature
review on Eucalyptus propagation, vitamin E, other than
being an antioxidant, affected rooting and speeded up the
rooting process upon addition to culture media [29].
Table 1. Effect of vitamins on plant growth and development in in vitro.
Vitamin Function1 Culture Medium Concentration Common Name
(Species) Effect2 Reference
B5 + 2 mg·l–1
2,4-D 10 mg·l–1 Stimulate cell growth Gamborg et al. 1968
MS basal medium
+ 1.7 gM BAP +
0.2 gM IBA
1.0 μM
Soybean
(Glycine max L.) Increase
embryogenesis Barwale et al. 1986
MS + 2 mg·l–1
2,4-D 0 mg·l–1 Maize (Zea mays L.)
Decrease callus
weight
Polikarpochkina et al.
1979
MS (Hormone free
MS)
0.5 mg·l–1;
0.5 mg·l–1 or 2 mg·l–1
Palm
(Phoenix
dactylifera L.)
Increase callus
weight, embryo
number; embryo
length
Al-Khayri 2001
MS + 4.2 μM GA 0.3 μM Pineapple (Ananas
comosus L.)
Reduce shoot
fresh mass Pérez et al. 2004
Thiamine (B1)
Cofactor in
carboxylase
reactions and amino
acid
biosynthesis
Linsmaier and
Skoog + 5 mg·l–1
2,4-D + 0.1 mg·l–1
BA
0.4 mg·l–1 Turf grass (Zoysia
japonica Steud.)
Increase embryonic
callus
Asano et al. 1996
MS + 1 mg·l–1
IBA 0.5 - 2 mg·l–1
Peach (Prunus
amygdalus x
persica Batsch.)
Inhibit rooting and
reduces callus Dimassi et al. 2005
De Fossard + 1.11
μM BA + 0.1 μM
IBA
7.97 μM Eucalyptus
globulus Labill.Stimulate rooting Trindade and Pais 1997
De Fossard + 10
μM IBA 31 μM
Papaya
(Carica papaya
L.)
Stimulate rooting Drew et al. 1993
Riboflavin (B2)
Oxidation-reduction
reactions (Transfer
of electrons)
MS + 3.2 μM IBA Not Known Apple (Malus
domestica Borkh.)Stimulate rooting Van der Krieken 1992
Shenk-Hildebrandt
(Hormone free)
10–9 M
Common Bean
(Phaseolus
vulgaris L.)
Stimulate rooting,
mitotic division, and
calcium absorption
Boland et al. 1989
Vitamin D3 -
MS (Hormone
free) 25 mg·l–1 Potato (Solanum tuberosum L.)
Enhance Calcium
absorption
Habib and Donnelly
2003
Biotin Cofactor of en-
zymes
MS (Hormone
free) 2 mg·l–1; 1 mg·l–1 Palm (Phoenix dactylifera L.)
Increase callus
weight, embryo
number; embryo
length
Al-Khayri 2001
Vitamin C
(Ascorbate) Reducing Agent MS + 10 μM IAA + 10 μM Kinetin 4 - 8× 10–4 M
Tobacco
(Nicotiana
tabacumn L.)
Increases shoot
number Joy et al. 1988
Nicotinic Acid Oxidation-reduction
reactions
MS basal medium
+ 1.7 gM BAP +
0.2 gM IBA
32.4 μM Soybean
(Glycine max L.)
Increase
embryogenesis Barwale et al. 1986
1Biochemical pathway in plant cell; 2Effects reported have been due to mixed interaction between vitamin and hormones in media, unless stated otherwise.

Effect of Vitamins on In Vitro Organogenesis of Plant 672
2.4. Embryo & Organ Development
Thiamine and nicotinic acid have been shown to affect
embryogenesis [30]. Barwale et al. [30] studied the effect
of different concentration of both vitamins on 40 imma-
ture soybean embryos cultured to a modified MS [10]
medium. Thiamine at 1.0 μM, or more, has induced 68%
embryogenesis compared to 0.2 μM, the level of salts in
MS medium, at 33% of the immature embryos. Also, a
concentration of 32.4 μM nicotinic acid induced 76%
embryogenesis [30].
Asano et al. [31] showed that enhancing embryonic
callus of Zoysia japonica Steud., a warm season turf
grass native to Japan, is obtained by adding thiamine and
riboflavin to the media. When thiamine was excluded
from the medium 50.3% callus was obtained, on the con-
trary, 0.4 and 4 mg·l–1 gave 53 and 60.3% respectively,
both insignificantly different. Furthermore, riboflavin was
not effective alone, except in the presence of thiamine at
4 mg·l–1 or higher concentration [31].
Thiamine and biotin have shown to be essential com-
ponents of tissue culture media for optimizing embryo-
genesis of date palm (Pheonix dactylifera L.) [19]. The
effect of thiamine has been shown to be dependent on
biotin for maximizing the number of somatic embryos,
which was also mentioned by Bonner [2]. The highest
number of embryos obtained was with a treatment of 0.5
or 2 mg·l–1 thiamine and 2 mg·l–1 biotin. However, the
optimum concentration for embryo number was a media
containing 0.5 mg·l–1 thiamine and 2 mg·l–1 biotin. Em-
bryo elongation also was affected by an interaction be-
tween both biotin and thiamin. The maximum embryo
length was achieved by 0.5 or 2 mg·l–1 thiamine and 1
mg·l–1 biotin [19].
Pérez et al. [32] studied the effect of thiamine and
other compounds on protease excretion in pineapple cul-
ture. Exogenous amounts of thiamine in the range of 0.3 -
1.2 μM had a negative effect on pineapple shoot fresh
mass, forming a plateau [32]. On the other hand, thia-
mine produced a maximum protein content at 0.6 μM,
while proteolytic and specific proteolytic activities both
at 0.3 μM [32].
Shoot weight of Papaya significantly increased in the
presence of both cytokinin and riboflavin, compared to a
medium of cytokinin only [20]. While a decrease of shoot
weight in the presence of riboflavin and auxin, possibly
related to photo-oxidation of auxin, also conveyed in
Gorst et al. [33] on Eucalyptus ficifolia F. Muell, was
reported in comparison to a medium containing only
auxin [20]. Also Drew et al. [34] reported that auxin
(Indole-3-butyric acid) concentrations at 10 μM with
more or less than, but not, 1 μM of riboflavin caused a
small rooting percentage. Moreover, increasing the con-
centration of riboflavin gradually from 0.1, 1, to 10 μM
degraded IBA, in the presence of light [34]. When 10 μM
of IBA is used, a complete destruction of IBA occurs
after 16 days versus 2 days when riboflavin is absent and
present, in light, respectively [34].
Exogenous application of 8 × 10–4 M and 4 - 8× 10–4
M of ascorbate to a shoot-forming media enhanced shoot
formation increased by 45% and 450% when using
young callus tissue (4 - 12 subcultures) and old callus
(>30 subcultures) of tobacco (Nicotiana tabacum L), re-
spectively, after 35 days in culture [35]. In the non-shoot
forming media, containing gibberellic acid, shoot-growth
of the young callus was significant at 4 × 10–4 M and
almost negligible for the old callus [35]. The former phe-
nomenon indicates an inhibitory action by ascorbate on
gibberellic acid. In addition, ascorbic acid reduced the
shoot-forming period [35].
Roest and Bokelmann [36] have shown that a high
number of adventitious shoot formation and transferable
shoots of Chrysanthemum was obtained when vitamins
were kept in the complete MS [10] medium. Whereas, a
medium where vitamins were eliminated suppressed
shoot formation although all other minerals were retained
[36].
On the contrary, omitting vitamins (thiamine, pyri-
doxine, nicotinic acid, folic acid, and biotin) from a
Bourgin and Nitsch [37] media in vitro did not affect 16
cultivars, except one, of Begonia x hiemalis shoot and
root formation [38]. Also, Soczek and Hempel [39] stud-
ied the shoot multiplication of three Gerbera cultivars in
the presence and absence of thiamine, pyridoxine, nico-
tinic acid and other compounds. It was concluded that
reducing the concentration, to half or quarter of the Mu-
rashige et al. [40] medium, or removing the vitamins, did
not have any significance on growth over three passages
(each 4 weeks), except in the case of one cultivar requir-
ing nicotinic acid [39].
3. Conclusions
Vitamins in culture media should be further studied in
order to justify their addition. For instance, little is known
about vitamin E (α-tocopherol), a phenol anti-oxidant,
presence in culture media. In the last few decades, little
interest has been observed in studying certain vitamins,
such as biotin and pantothenic acid. Plant species and
cultivars require different amount of vitamins, while other
do need any at all. For instance, after several passages,
thiamine is essential to soybean, rice, and tobacco cul-
tures but non-essential to peanut cells, which contain
high thiamine concentration [41]. The physiological and
morphological output varies between plants when using
the same vitamins. According to our desired outcome
culture media remain open to modifications, especially

Effect of Vitamins on In Vitro Organogenesis of Plant673
the common Murashige and Skoog [10]. Although sig-
nificant vitamins such as thiamine impose their applica-
tion in culture media; others are poorly applied such as
ascorbic acid.
Scientific knowledge on plant propagation was not the
only significant outcome; however, some experiments
offered economic solutions. In order to reduce costs, Drew
et al. [27,34] suggested adding riboflavin, which degrades
auxin, to the tissue culture media rather than transferring
the tissue to a hormone (i.e. auxin) free media. In the
future, studying the effect on a wider range of vitamins
and plant simultaneously is needed for an enhanced fea-
sibility outcome.
REFERENCES
[1] R. H. Horton, “Coenzymes and Vitamins,” Principles of
Biochemistry, Pearson Education International, Upper
Saddle River, 2006.
[2] J. Bonner, “The Role of Vitamins in Plant Development,”
Botanical Review, Vol. 3, No. 12, 1937, pp. 616-640.
doi:10.1007/BF02872294
[3] E. F. George, M. A. Hall and G.-J. De Klerk, “The Com-
ponents of Plant Tissue Culture Media II,” Plant Propa-
gation by Tissue Culture, Springer, Dordrecht, 2008, pp.
115-119.
[4] A. Goyer, “Thiamine in Plants: Aspects of Its Metabolism
and Functions,” Phytochemistry, Vol. 71, No. 14-15,
2010, pp. 1615-1624.
doi:10.1016/j.phytochem.2010.06.022
[5] O. Kursteiner, I. Dupuis and C. Kuhlemeier, “The Pyru-
Vate Decarboxylase1 Gene of Arabidopsis Is Required
During Anoxia but Not Other Environmental Stresses,”
Plant Physiology, Vol. 132, No. 2, 2003, pp. 968-978.
doi:10.1104/pp.102.016907
[6] J. Malamy, P. S. Casas, J. Hennig, A. L. Guo and D. F.
Klessig, “Dissection of the Salicylic Acid Signaling Path-
way in Tobacco,” Molecular Plant—Microbe Interaction,
Vol. 9, No. 6, 1996, pp. 474-482.
doi:10.1094/MPMI-9-0474
[7] I. P. Ahn, S. Kim and Y. H. Lee, “Vitamin B1 Functions
as an Activator of Plant Disease Resistance,” Plant Physi-
ology, Vol. 138, No. 3, 2005, pp. 1505-1515.
doi:10.1104/pp.104.058693
[8] M. Tunc-Ozdemir, “Thiamin Confers Enhanced Toler-
ance to Oxidative Stress in Arabidopsis,” Plant Physiol-
ogy, Vol. 151, No. 1, 2009, pp. 421-432.
doi:10.1104/pp.109.140046
[9] J. A. Gonzalez-Reyes, F. J. Alcain, J. A. Caler, A.
Serrano, F. Cordoba and P. Navas, “Stimulation of Onion
Root Elongation by Ascorbate and Ascorbate Free Radi-
cal in Allium cepa L,” Protoplasma, Vol. 184, 1995, pp.
31-35. doi:10.1007/BF01276898
[10] T. Murashige and F. Skoog, “A Revised Medium for
Rapid Growth and Bio Assays with Tobacco Tissue Cul-
tures,” Physiologia Plantarum, Vol. 15, No. 3, 1962, pp.
473-497. doi:10.1111/j.1399-3054.1962.tb08052.x
[11] S. Ray, “On the Nature of the Precursor of the Vitamin C
in the Vegetable Kingdom. I. Vitamin C in the Growing
Pea Seedling,” Biochemical Journal, Vol. 28, No. 3, 1934,
pp. 996-1003.
[12] A. Habib and D. J. Donnelly, “Vitamin Ds Improve Cal-
cium Uptake in Micropropagated Potato Plantlets,” Acta
Horticulturae, Vol. 619, 2003, pp. 263-269.
[13] O. Gamborg, R. Miller and K. Ojima, “Nutrient Require-
ments of Suspension Cultures of Soybean Root Cells,”
Experimental Cell Research, Vol. 50, No. 1, 1968, pp.
151-158. doi:10.1016/0014-4827(68)90403-5
[14] T. Eriksson, “Studies on the Growth Requirements and
Growth Measurements of Cell Cultures of Haplopappus
gracilis,” Physiologia Plantarum, Vol. 18, No. 4, 1965,
pp. 976-993. doi:10.1111/j.1399-3054.1965.tb06994.x
[15] R. T. Polikarpochkina, K. Z. Gamburg and E. E. Khavin,
“Cell-Suspension Culture of Maize (Zea mays L.),” Zeits-
chrift fr Pflanzenphysiologie, Vol. 95, No. 1, 1979, pp.
57-67.
[16] J. Digby and F. Skoog, “Cytokinin Activation of Thia-
mine Biosynthesis in Tobacco Callus Cultures,” Plant
Physiology, Vol. 41, No. 4, 1966, pp. 647-652.
doi:10.1104/pp.41.4.647
[17] E. M. Linsmaier-Bednar and F. Skoog, “Thiamine Requi-
rement in Relation to Cytokinin in ‘normal’ and ‘mutant’
Strains of Tobacco Callus,” Planta, Vol. 72, No. 2, 1966,
pp. 146-154. doi:10.1007/BF00387478
[18] D. E. Dravnieks, F. Skoog and R. H. Burris, “Cytokinin
Activation of De Novo Thiamine Biosynthesis in To-
bacco Callus Cultures,” Plant Physiology, Vol. 44, No. 6,
1969, pp. 866-870. doi:10.1104/pp.44.6.866
[19] J. M. Al-Khayri, “Optimization of Biotin and Thiamine
Requirements for Somatic Embryogenesis of Date Palm
(Phoenix dactylifera L.),” In Vitro Cellular & Develop-
mental Biology—Plant, Vol. 37, No. 4, 2001, pp. 453-
456. doi:10.1007/s11627-001-0079-x
[20] R. A. Drew and N. G. Smith, “Growth of Apical and Lat-
eral Buds of Papaya (Carica papaya L.) as Affected by
Nutritional and Hormonal Factors,” Journal of Horticul-
tural Science, Vol. 61, No. 1, 1986, pp. 535-543.
[21] M. C. De Pinto, D. Francis and L. Gara, “The Redox
State of the Ascorbate-dehydroascorbate Pair as a Speci-
fic Sensor of Cell Division in Tobacco BY-2 Cells,” Pro-
toplasma, Vol. 209, No. 1-2, 1999, pp. 90-97.
doi:10.1007/BF01415704
[22] R. Boland, L. Talmon, M. Vega and B. Mujica, “Cyto-
histological Studies on the Action of Vitamin D3 and
Stigmasterol on Phaseolus Vulgaris Roots Growing in
Vitro,” Plant Science, Vol. 59, No. 2, 1989, pp. 183-190.
doi:10.1016/0168-9452(89)90136-2
[23] K. Dimassi, C. Antonopoulou, I. Therios, C. Chatzi-
ssavvidis and V. Tsirakoglou, “Inhibitory Effects of Ri-
boflavin (Vitamin B) on the in Vitro Rooting and Nutrient
Concentration of Explants of Peach Rootstock GF 677
(×),” Scientia Horticulturae, Vol. 106, No. 2, 2005, pp.
268-272.

Effect of Vitamins on In Vitro Organogenesis of Plant 674
[24] W. M. Van der Krieken, H. Breteler, M. H. M. Visser and
W. Jordi, “Effect of light and riboflavin on indolebutyric
acid-induced root formation on apple in vitro,” Physiolo-
gia Plantarum, Vol. 85, No. 4, 1992, pp. 589-594.
doi:10.1034/j.1399-3054.1992.850405.x
[25] H. Trindade and M. S. Pais, “In Vitro Studies on Euca-
lyptus Globulus Rooting Ability,” In Vitro Cellular &
Developmental Biology—Plant, Vol. 33, No. 1, 1997, pp.
1-5. doi:10.1007/s11627-997-0032-8
[26] R. A. Drew, J. A. McComb and J. A. Considine, “Rhizo-
genesis and Root Growth of Carica Papaya L. in Vitro in
Relation to Auxin Sensitive Phases and Use of Ribofla-
vin,” Plant Cell, Tissue and Organ Culture, Vol. 33, No.
1, 1993, pp. 1-7. doi:10.1007/BF01997591
[27] R. A. Fossard, A. Myint and E. C. M. Lee, “A Broad
Spectrum Tissue Culture Experiment with Tobacco
(Nicotiana tabacum L.) Pith Tissue Callus,” Physiologia
Plantarum, Vol. 31, No. 2, 1974, pp. 125-130.
doi:10.1111/j.1399-3054.1974.tb03116.x
[28] P. P. Chee, “Stimulation of Adventitious Rooting of
Taxus Species by Thiamine,” Plant Cell Reports, Vol. 14,
No. 12, 1995, pp. 753-757. doi:10.1007/BF00232916
[29] J. J. Le Roux and J. Van Staden, “Micropropagation and
Tissue Culture of Eucalyptus—A Review,” Tree Physio-
logy, Vol. 9, No. 4, 1991, pp. 435-477.
[30] U. B. Barwale, H. R. Kerns and J. M. Widholm, “Plant
Regeneration from Callus Cultures of Several Soybean
Genotypes via Embryogenesis and Organogenesis,” Plan-
ta, Vol. 167, No. 4, 1986, pp. 473-481.
doi:10.1007/BF00391223
[31] Y. Asano, H. Katsumoto, C. Inokuma, S. Kaneko, Y. Ito
and A. Fujiie, “Cytokinin and Thiamine Requirements
and Stimulative Effects of Riboflavin and Alpha-Ke-
toglutaric Acid on Embryogenic Callus Induction from
the Seeds of Zoysia japonica Steud,” Journal of Plant
Physiology, Vol. 149, No. 3-4, 1996, pp. 413-417.
[32] A. Pérez, L. Nápoles, C. Carvajal, M. Hernandez and J. C.
Lorenzo, “Effect of Sucrose, Inorganic Salts, Inositol, and
Thiamine on Protease Excretion during Pineapple Culture
in Temporary Immersion Bioreactors,” In Vitro Cellular
& Developmental Biology—Plant, Vol. 40, No. 3, 2004,
pp. 311-316. doi:10.1079/IVP2004529
[33] J. R. Gorst, M. Slaytor and R. A. De Fossard, “The Effect
of Indole-3-Butyric Acid and Riboflavin on the Morpho-
genesis of Adventitious Roots of Eucalyptus ficifolia F.
Muell. Grown in Vitro,” Journal of Experimental Botany,
Vol. 34, No. 11, 1983, pp. 1503-1515.
doi:10.1093/jxb/34.11.1503
[34] R. A. Drew, B. W. Simpson and W. J. Osborne, “Degra-
dation of Exogenous Indole-3-Butyric Acid and Ribofla-
vin and Their Influence on Rooting Response of Papaya
in Vitro,” Plant Cell, Tissue and Organ Culture, Vol. 26,
No. 1, 1991, pp. 29-34. doi:10.1007/BF00116606
[35] R. W. Joy, K. R. Patel and T. A. Thorpe, “Ascorbic Acid
Enhancement of Organogenesis in Tobacco Callus,”
Plant Cell, Tissue and Organ Culture, Vol. 13, No. 3,
1988, pp. 219-228. doi:10.1007/BF00043670
[36] S. Roest and G. Bokelmann, “Vegetative Propagation of
Chrysanthemum morifolium Ram. in Vitro,” Scientia
Horticulturae, Vol. 3, No. 4, 1975, pp. 317-330.
doi:10.1016/0304-4238(75)90046-1
[37] J. P. Bourgin and J. P. Nitsch, “Obtention de Nicotiana
Haploïdes à Partir d’Étamines Cultivées in Vitro. (Produc-
tion of Haploid Nicotiana from Excised Stamen),” Annales
de Physiologie Végétale, Vol. 9, 1967, pp. 377-382.
[38] T. Welander, “In Vitro Organogenesis in Explants from
Different Cultivars of Begonia x Hiemalis,” Physiologia
Plantarum, Vol. 41, No. 2, 1977, pp. 142-145.
doi:10.1111/j.1399-3054.1977.tb05546.x
[39] U. Soczek and M. Hempel, “The Influence of Some Or-
ganic Medium Compounds on Multiplication of Gerbera
in Vitro,” Acta Horticulturae, Vol. 226, 1988, pp. 643-
646.
[40] T. Murashige, M. Serpa and J. B. Jones, “Clonal Multi-
plication of Gerbera through Tissue Culture,” Horticul-
ture Science, Vol. 9, No. 3, 1974, pp. 175-180.
[41] K. Ohira, M. Ikeda and K. Ojima, “Thiamine Require-
ments of Various Plant Cells in Suspension Culture,”
Plant Cell Physiology, Vol. 17, No. 3, 1976, pp. 583-588.