SHORT COMMUNICATION

In vitro direct rhizogenesis from Gerbera jamesonii Bolus leaf

Saikat Gantait • Uma Rani Sinniah

Received: 18 January 2014 / Revised: 18 July 2014 / Accepted: 20 July 2014

� Franciszek Gorski Institute of Plant Physiology, Polish Academy of Sciences, Krakow 2014

Abstract The present report describes an original proto-

col for in vitro direct induction of roots from leaf explants

of gerbera for the first time. Since gerbera has immense

potential as a premium cut-flower, the major attempts were

made on in vitro mass propagation chiefly through in vitro

multiple shoot proliferation or callus regeneration. Never-

theless, rhizogenesis could be impending an unattempted

method with its yet-to-be known advantages. In our study,

the optimum conditions for direct root induction from leaf

explants were assessed employing tissue culture technique.

Leaves were inoculated to MS medium containing no or

variable auxin sources and concentrations namely, 2,4-

dichlorophenoxyacetic acid, indole-3-acetic acid (IAA),

indole-3-butyric acid or a-naphthaleneacetic acid for root

induction. It was evident that the maximum root induction

(with a frequency of 92.6 %) occurred on MS media for-

tified with 1.5 mg l-1 IAA, wherein root induction was

observed as early as 11 days of culture and an average of

*19 roots with *13 mm length was obtained from 4 cm2

leaf segment after 45 days of culture. Stereo microscopic

observation revealed the induction of roots and gradual

developmental stages of rhizogenesis. The efficiency of

direct root induction without any interim growth stages

(such as, callus or shoots) in our study offers a reproducible

system that could provide a model protocol for more

comprehensive developmental studies on root growth.

Keywords Adventitious rooting � Auxins � Coumarin �Developmental stages � Leaf explant

Abbreviation

2,4-D 2,4-Dichlorophenoxyacetic acid

IAA Indole-3-acetic acid

IBA Indole-3-butyric acid

MS Murashige and Skoog medium

NAA a-Naphthaleneacetic acid

PGR Plant growth regulator

PPFD Photosynthetic photon flux density

Introduction

Gerbera (Gerbera jamesonii Bolus), belonging to the

family Asteraceae, is considered as one of the most popular

cut-flowers, as well as potted floriculture plant. Owing to

its array of colourful inflorescences it has attained out-

standing commercial importance as cut-flowers (Gantait

et al. 2011). The gerbera plant is also used in the prepa-

ration of traditional Chinese medicine (Singh 2006). It

contains naturally occurring three new coumarin deriva-

tives 1–3 and a known coumarin 4 (Inoue et al. 1989; Liu

et al. 2010; Qiang et al. 2011). Coumarins from Gerbera

were found to have broad spectrum anti-tumor and anti-

bacterial activities in vitro and in vivo and are considered

to be potential new cancer chemoprevention agents (He

et al. 2014). It is a clonally propagated plant and shows

restricted growth of apical meristem which restrains the

Communicated by B. Borkowska.

S. Gantait � U. R. Sinniah

Department of Crop Science, Faculty of Agriculture, Universiti

Putra Malaysia, 43400 Serdang, Selangor, Malaysia

S. Gantait (&)

Sasya Shyamala Krishi Vigyan Kendra, Ramakrishna Mission

Vivekananda University, Arapanch, Sonarpur,

Kolkata 700150, India

e-mail: saikatgantait@yahoo.com

123

Acta Physiol Plant

DOI 10.1007/s11738-014-1643-4

axillary buds from developing offshoots. Consequently, a

low rate of multiplication is evident during conventional

propagation. Tissue culture systems for mass propagation

of gerbera, as a substitute, have been effectively com-

menced by several researchers. Shoot tip, young capitulum

and axillary bud culture are the most frequently used

methods in gerbera for direct organogenesis such as mul-

tiple shoot, somatic embryo (Gantait et al. 2010).

Direct induction of only roots exclusive of any inter-

mittent organs such as callus or shoots or somatic embryos

could be a simple yet efficient way to mass produce root-

specific secondary metabolites escaping transgenic inter-

vention (Tennant 2010). Direct root induction from leaf

explant as a method for the rapid root regeneration has

been reported earliest by Tran Thanh Van et al. (1974) on

Begonia rex Putz., later on several researchers reported this

system was employed for several plants viz, Lycopersicon

esculentum Mill. (Coleman and Greyson 1976), Nicotiana

tabacum Linn. (Attfield and Evans 1991), Helianthus oc-

cidentalis Riddell (Samaj et al. 1999), Eustoma grandiflo-

rum Shinn. (Tuan et al. 2006), and Ophiorrhiza prostrata

D. Don (Martin et al. 2008).

Compositions of plant growth regulator (PGR) have

been reported to encompass species-specific or even cul-

tivar-specific effect on in vitro root induction, with auxins

(both in types and concentrations), showing the most

imperative effect on direct root induction (Gantait et al.

2009; Teixeira da Silva 2013). It is noteworthy to mention

that auxin-induced rooting through the application of single

doses of auxins, usually singly, was effective in Chrysan-

themum (Dendranthema 9 grandiflora Ramat. Kitamura)

(Teixeira da Silva 2003), Anthurium andreanum Lind., and

Gerbera jamesonii Bolus (Gantait et al. 2008, 2010). The

influence of auxins on direct rhizogenesis from leaf seg-

ments has been reported on Boerhaavia diffusa Linn.

(Shrivastava and Padhya 1995), Rauwolfia serpentina (L.)

Benth (Pandey et al. 2010), Costus igneus Linn. (Nagarajan

et al. 2011), and Carica papaya Linn. (Teixeira da Silva

2013).

In our knowledge, to the current date, no study has been

done on direct induction of root in gerbera. Hence, con-

sidering the competence of leaf explants, the establishment

of a protocol for direct induction of roots from leaf and

their developmental study will be of practical importance.

Counting Gerbera jamesonii Bolus, there are several

ornamental plants species of Asteraceae such as Calendula

officinallis Linn., Chrysanthemum indicum (L.) Desr.,

Spilanthes acmella Auct. Plur., Tagetes erecta Linn.,

Wedelia calendulacea (L.) Less. those are also well known

for their pharmaceutical values due to the presence of

secondary metabolites (Rothe et al. 2011) which can be

accumulated in root system (Qiang et al. 2011). A repro-

ducible rhizogenic system thus would be if practical

importance to several other species of Gerbera carrying

secondary metabolites. Hence, in this article we propose an

accelerated rhizogenesis system of gerbera, a renowned

cut-flower with pharmaceutical values, via direct root

induction from leaf segments by adjusting types and con-

centrations of auxins, followed by the developmental study

at morpho-anatomical level.

Materials and methods

Plant material

One-year old gerbera plants (Gerbera jamesonii Bolus cv.

Sciella) were collected from commercial nursery and

kept growing under polyhouse, with a maximum

200 lmol m-2 s-1 photosynthetic photon flux density

(PPFD) at Universiti Putra Malaysia, Selangor, Malaysia.

Young fully opened leaves of 4–5 cm length were taken

from the source plant 20 days following their emergence

and used for in vitro study.

Surface disinfection and explant preparation

Collected leaves were rinsed for 3 min in tap water; next

submerged in 1 % (v/v) Tween-20 (dissolved in sterile-

distilled water) in a glass beaker for 5 min, 70 % (v/v)

ethanol for 30 s, 3 % (v/v) hydrogen peroxide for 5 min,

and lastly in 20 % Clorox� (Clorox Co., Broadway, Oak-

land, CA, USA) (1.2 % sodium hypochlorite) solution for

15 min, with thorough shaking under a laminar air flow.

Instant sonication in an ultrasonic cleaner was done with

the leaves at full power for 10 min. Subsequently; the

leaves were thoroughly rinsed three times with sterile-

distilled water, every time after decanting the surface dis-

infectant. Then these were shifted to a sterile Petri dish

(15 9 150 mm) for brief drying for 5 min. Each leaf was

cut perpendicularly to its axis into 4 cm2 sections by a

sharp sterile scalpel.

Culture conditions

The basal MS (Murashige and Skoog 1962), containing

3 % (w/v) sucrose and 100 mg l-1 myo-inositol, gelled

with agar was used. Auxins (Sigma-Aldrich, St. Louis,

USA) were added to the medium as specified below. The

pH of the medium was adjusted to 5.8 using 1 N NaOH or

HCl, prior to addition of solidifying agent. After the

addition of 7 % (w/v) agar, media were sterilized in an

autoclave (Hirayama, Japan) at 121 �C and 1 kg cm-2 for

20 min. Auxin solutions were filter-sterilized (membrane

filtration 0.22 lm) and added to autoclaved basal medium

when the temperature fallen to 50 �C. Medium (20 ml) was

Acta Physiol Plant

123

dispensed into each of the culture tubes (50 mm

height 9 25 mm inner diameter with 1 mm thick) and kept

those slant. Polypropylene autoclavable lids (24 mm

height 9 26 mm inner diameter with 1 mm thick) were

used as closures for culture tubes. The cultures were

maintained at 25 ± 1 �C under a 12 h photoperiod, which

was provided by cool, white fluorescent light (Phillips Life

Max, Thailand) with an irradiance of 60 lmol m-2 s-1

PPFD (measured with LI-COR, Lincoln, USA).

Optimization of auxins in culture medium for root

induction

Sterilized explants were placed adaxial side down in culture

tube containing MS medium supplemented with 0.5, 1.0, 1.5,

and 2.0 mg l-1 of either 2,4-dichlorophenoxyacetic acid

(2,4-D), indole-3-acetic acid (IAA), indole-3-butyric acid,

(IBA) or a-naphthaleneacetic acid (NAA), to assess the

effects of the auxin types and concentrations on root induc-

tion and development. MS medium without any PGR was

treated as control. All cultures were grown under environ-

mental conditions as mentioned above. The cultures were

sustained till initiation and development of roots occurred.

Observation was carried out frequently in order to record the

time (in days) taken to root induction. Data collection on

frequency of root induction from explants, and mean number

of root induced per explant was made at 45 days after the

inoculation of explants. In addition, the lengths of the indi-

vidual root were measured. Microscopic observations were

carried out at 109 magnification using a Leica stereozoom

microscope (Leica, Germany) at every week of culture. The

frequency of root induction was determined after 30 days,

using the formula: F = [(E/E0) 9 100] %; wherein: F fre-

quency of root induction, E number of explants inducing

roots, and E0 number of explants inoculated.

Morpho-anatomical observation

Detailed morphological and anatomical examination and

photography of the cultures were carried out using a digital

camera (Panasonic LX5, Japan) and stereozoom micro-

scope (Leica, Germany), correspondingly. Once the

spherules (a protruding structure induced from the undif-

ferentiated tissue prior to root formation) were formed,

observations were recorded weekly to trace stage-wise root

development. For stereo microscopic examination, about

10 roots at each developmental stage were indiscriminately

collected from culture tubes.

Experimental design and statistical analysis

The experiments were arranged in completely randomized

design (CRD). All the aforementioned experiments were

replicated three times, using 20 samples per replicate. Each

inoculated leaf segments were considered as single exper-

imental unit. Visual observations were made every single

day. Data were statistically analysed by means of SAS�-

vers. 6.12 (SAS Institute, Cary, NC, USA) (SAS Institute

1999) by ANOVA. Statistical significance between

mean ± SE were compared based on Duncan’s multiple

range test (DMRT) (Duncan 1955) at a P value = 0.05.

Data expressed as percentage were transformed using arc

sine before ANOVA and converted back to the initial scale

(Compton 1994).

Results and discussion

Gerbera, employed in the present study to establish a direct

root induction protocol using leaf segments, gave a number

of significant results. It was found that leaf segments could

be used to induce roots even in PGR-free media. Never-

theless, the competence of leaf explants inducing roots

significantly varied with the auxin types and levels tested,

in terms of percentage induction, the number of roots and

time taken for root induction. Interestingly, auxin-free

medium induced in vitro roots but with very low frequency

and less number. In addition, the time taken to induce roots

was much longer as compared to the media supplemented

with auxins (Table 1). Fortification of three different aux-

ins (IAA, IBA and NAA) at four levels of 0.5–2 mg l-1 in

MS medium was indispensable for enhanced root induction

and well developed root system within a brief culture

period, whilst 2,4-D was not beneficial like control and was

able to result in analogous response (Table 1). All treat-

ments inclusive of control but excluding 2,4-D with higher

concentrations, resulted in root induction with varying

frequencies (from 17.3 to 92.6 %). There was a significant

difference (P = 0.05) among the effects of four auxin

sources according to Duncan’s multiple range test. Root

induction in the presence of 2,4-D was very low in com-

parison to other auxins used, to the point no root induction

was observed when concentration was more than 1 mg l-1.

Amid IBA and NAA, the latter showed improved effect on

root induction, whereby very low concentration of

0.5 mg l-1 triggered 74.3 % root induction from the

wounded leaf surface. Rising the concentration of these

two PGRs, appeared to have inhibitory influence on rhi-

zogenesis. Among the auxins employed, induction of root

was most proficient in the presence of IAA where

*81–93 % induction was recorded with IAA supplemen-

tation ranging from 0.5 to 2 mg l-1. The superlative

response was recorded for the medium fortified with

1.5 mg l-1 IAA, on which 92.6 % of the explants induced

on an average of 19 roots per 4 cm2 leaf segment, within

45 days of culture. The experiment was evaluated at

Acta Physiol Plant

123

45 days of culture since, at the initial stage roots were

appeared as a mass of structures, making them unfeasible

to count, even with a stereomicroscope.

The morphogenetic phases from direct induction to

development of roots are reviewed in Fig. 1. The initial

symptom of morphogenesis was observed in the form of

swelling from the cut portions (Fig. 1a). As reviewed by

Blakesley et al. (1991) the first visible sign of a new

meristematic locus was the expansion of a single cell, with

simultaneous nuclear swelling. The cytoplasm of such cells

was much denser than that of adjacent cells. However, this

morphogenesis turned either into callus mass (Fig. 1b) with

the influence of higher 2,4-D levels or induced roots

directly. Initial morphological signal of root induction

appeared as swollen, small nodule-like meristemoids on the

wound abaxial surface of leaves, within 11 days after

inoculation of explants (Fig. 1c). Subsequently the nodules

advanced into slender cylindrical bodies with globular root

primordial tip, rough surfaces and were pale pinkish in

color (Fig. 1d). The basal part of the protruding structures

appeared to have tiny root hairs. The whole leaf segment

disorganized with subsequent rhizogenesis. This formula-

tion also resulted in the highest mean length (*13 mm) of

roots. The progressive morphological change during

growth and development of the root system appearance

during the maturity stage is shown in Fig. 1e. Roots started

to disintegrate and became prominent from each other as its

growth progressed to a more advanced developmental

stages. However, with excess auxin concentration, the

development of rhizogenesis was affected with reduced

root length and profuse root hairs formation (Fig. 1f).

Although little research has been carried out on in vitro

direct root induction or organogenesis from leaf explants in

ornamentals, there are a few describing structural details

(Blakesley et al. 1991; Gantait et al. 2012). However, this

is the first study, which traced the various developmental

stages of induction of roots through direct organogenesis

with naked eye with corroboration to the details derived

from micromorphological observations in gerbera. The

formation of initial swelling, induction of primordial

structure and complete rootlets with root cap has been

revealed to be imperative events in the development of

in vitro rhizogenesis. Direct in vitro induction of roots from

leaf explants was first documented by Tran Thanh Van

et al. (1974). Since then a number of studies on direct root

induction using leaf explants have been reported (Coleman

and Greyson 1976; Attfield and Evans 1991; Samaj et al.

1999; Tuan et al. 2006; Martin et al. 2008). The leaf seg-

ments of gerbera used in our study displayed high regen-

erative competence by inducing direct root formation from

the cut surface of leaves which inflicted wound. Wounded

surface have been often reported to induce callus as an

indicator of repair system. In several ornamentals, instead

of producing callus, roots were induced directly. As a

consequence of the wounding-response phenomenon, dor-

mant unwounded cells near the cut surface become active

and initiate cell proliferation. The commencement of cell

proliferation from intact, unwounded cells have been

Table 1 Effect (growth period

45 days) of variable levels of

auxins on direct root induction

from leaf segment of gerbera

(Gerbera jamesonii Bolus)

Data represent

mean ± standard error of 20

replicate explants per treatment

in three repeated experiments

(C control)

Data expressed as percentage

were transformed using arc sine

prior to ANOVA and converted

back to the original scale for

demonstration in the table

(Compton 1994)

Means within a single column

followed by different letters are

significantly different according

to Duncan’s multiple range test

(Duncan 1955) at P = 0.05

MS ? auxins (mg l-1) Frequency of root

induction (%)

Days to root

induction

No. of roots Root length

(mm)2,4-D IBA IAA NAA

C 33.3 ± 0.3 gh 19.6 ± 0.4 ab 5.2 ± 0.6 fg 5.2 ± 0.1 d

0.5 26.2 ± 0.6 i 21.3 ± 0.4 a 6.6 ± 0.6 f 4.4 ± 0.2 de

1 17.3 ± 1.2 j 19.6 ± 0.3 ab 4.5 ± 0.7 g 4.8 ± 0.0 de

1.5 0.0 ± 0.0 k 0.0 ± 0.0 e 0.0 ± 0.0 h 0.0 ± 0.0 f

2 0.0 ± 0.0 k 0.0 ± 0.0 e 0.0 ± 0.0 h 0.0 ± 0.0 f

0.5 34.6 ± 0.4 gh 17.6 ± 0.4 b 4.3 ± 0.4 g 4.6 ± 0.3 de

1 37.8 ± 0.2 fg 17.1 ± 0.3 b 11.6 ± 0.3 cd 4.8 ± 0.2 de

1.5 41.3 ± 0.5 f 19.8 ± 0.2 ab 8.6 ± 0.2 e 6.2 ± 0.2 cd

2 28.6 ± 0.2 hi 22.6 ± 0.3 a 5.3 ± 0.2 fg 5.9 ± 0.3 cd

0.5 84.6 ± 0.7 c 15.3 ± 0.6 c 12.3 ± 0.6 c 7.6 ± 0.1 c

1 87.4 ± 1.3 b 15.6 ± 0.4 c 15.4 ± 0.3 bc 9.6 ± 0.1 b

1.5 92.6 ± 0.7 a 11.3 ± 0.4 d 19.3 ± 1.1 a 12.8 ± 0.1 a

2 81.3 ± 0.6 cd 13.6 ± 0.3 cd 16.4 ± 0.3 b 9.2 ± 0.3 b

0.5 74.3 ± 0.3 d 16.3 ± 0.3 bc 12.8 ± 0.6 c 4.7 ± 0.3 de

1 71.8 ± 0.6 de 17.1 ± 0.6 b 13.6 ± 0.4 c 5.3 ± 0.1 d

1.5 66.7 ± 0.3 e 17.6 ± 0.3 b 11.4 ± 0.5 cd 6.5 ± 0.3 cd

2 65.3 ± 0.7 e 19.8 ± 0.7 ab 7.3 ± 0.2 ef 3.6 ± 0.1 e

Acta Physiol Plant

123

attributed to a range of biochemical transformation such as

degradation of starch, raise in peroxidase and polyphenol

oxidase which have been activated within the quiescent cell

(Imaseki 1985). Conversely, the relationship between these

metabolic changes and root induction is still indistinct and

leftover unexamined.

The other remarkable observation in our study is the

typical regulatory consequence of auxin concentrations on

direct root induction. It was evident that the presence of

auxin enhanced the incidence of root number as well as

reduced the time taken for its initiation. In consideration of

the fact that the survival of explants was a role of the auxin

present in the medium, it is rational to infer that a passable

endogenous PGR level may have been present in those

explants that survived in the control (Gantait and Sinniah

2012). Consequently, one rational explanation for the

incidence of rhizogenesis in the control medium would be a

potential role of endogenous auxin. This opinion is based

on a preceding proposition that the endogenous auxin

content of initial explants might have an imperative func-

tion in organogenic competence (Pandey et al. 2010). In the

present study, the occurrence of root induction increased

with raising concentrations of auxins, but started declining

following an optimum level. Reduced root induction might

be attributed to the excess cumulative level of accumulated

endogenous and raised exogenous auxins. As reviewed by

Blakesley et al. (1991) the presence of exogenous auxin

was most effective for direct root formation but the con-

centration required for optimum response was dependent

upon the type of auxin used. Significant differences in root

induction rate of gerbera were observed between IAA

treatments when compared to that of the 2,4-D, IBA or

NAA. It has been suggested in several experiments that

IAA proved to be the most effective for root induction in

comparison to other auxins (Gantait et al. 2008, 2010). Its

mode of action may be attributed to its ability to influence

the biosynthesis and accumulation of endogenous auxins

(Reviewed by Blakesley et al. 1991). According to Murch

Fig. 1 Direct induction and development of root system in Gerbera

jamesonii Bolus. a Initial symptom of morphogenesis in form of

swelling from the cut portions due to expansion of a cells (Bar 5 mm),

b initiation of callus from the wounded surface at higher 2,4-D

concentrations (Bar 3 mm), c root induction appeared as swollen,

small nodule-like meristemoids on the wound abaxial surface of

leaves, within 11 days after inoculation (Bar 3 mm), d the nodules

advanced into slender cylindrical bodies with globular root primordial

tip, rough surfaces and were pale pinkish in color (Bar 5 mm),

e growth and development of the disintegrated and prominent

rhizohenesis at advanced stage (Bar 5 mm), f rhizogenesis affected

with reduced root length and profuse root hairs at with excess auxin

concentration (Bar 5 mm)

Acta Physiol Plant

123

et al. (1997) PGR induces stress in plants, under in vitro

condition and induction of roots might be an adaptive

organogenic mechanism of leaf segments to overcome the

combined stress induced by wound and IAA. The modes of

actions of IAA are not absolutely explored. IAA may be

involved in the reprogramming and expression of the

competent cells necessary for them to undergo differenti-

ation and development (Reviewed by Blakesley et al.

1991). Although earlier researches have revealed that the

regenerative response might be due to stimulation of the

endogenous PGR system, it can be suggested based on the

present results that IAA presumably induces an array of

biochemical pathways to generate particular metabolites

essential for root induction procedure. In the present study,

IAA at 1.5 mg l-1 alone induced 92.6 % roots compared to

33.3 % in control. On the contrary, a number of reports

have shown the inadequacy of IAA alone on in vitro root

induction from young leaf explants and leaf segments

(Pandey et al. 2010; Nagarajan et al. 2011).

A competent system for the accelerated organogenesis

in terms of direct root induction from gerbera leaf has been

described in the current study. To our knowledge, such an

effort concerning the complete protocol and developmental

study of direct root induction has been made for the first

time on any ornamental plant.

Author contribution S. Gantait, U.R. Sinniah—Con-

ceived the research idea and designed the experiments;

S. Gantait—Executed the experiments; S. Gantait,

U.R. Sinniah—Wrote the manuscript.

Acknowledgments The authors are indebted to Department of Crop

Science, Faculty of Agriculture, Universiti Putra Malaysia for pro-

viding the research facilities.

Conflict of interest We, the authors of this article, declare that there

is not conflict of interest and we do not have any financial gain from it.

References

Attfield EM, Evans PK (1991) Developmental pattern of root and

shoot organogenesis in cultured leaf explants of Nicotiana

tabacum. J Exp Bot 42:51–57. doi:10.1093/jxb/42.1.51

Blakesley D, Weston GD, Hall JF (1991) The role of endogenous

auxin in root initiation. Part I: evidence from studies on auxin

application, and analysis of endogenous levels. Plant Growth

Regul 10:341–353. doi:10.1007/BF00024593

Coleman WK, Greyson RI (1976) Root regeneration from leaf

cuttings of Lycopersicon esculentum Mill: application of the leaf

plastochron index and responses to exogenous Gibberellic acid.

J Exp Bot 27:1339–1351. doi:10.1093/jxb/27.6.1339

Compton ME (1994) Statistical methods suitable for the analysis of

plant tissue culture data. Plant Cell Tissue Organ Cult

37:217–242. doi:10.1007/BF00042336

Duncan DB (1955) Multiple range and multiple F test. Biometrics

11:1–42

Gantait S, Sinniah UR (2012) Rapid micropropagation of monopodial

orchid hybrid (Aranda Wan Chark Kuan ‘Blue’ 9 Vanda

coerulea Grifft. ex. Lindl.) through direct induction of proto-

corm-like bodies from leaf segments. Plant Growth Regul

68:129–140. doi:10.1007/s10725-012-9698-y

Gantait S, Mandal N, Bhattacharyya S, Das PK (2008) In vitro mass

multiplication with pure genetic identity in Anthurium andrea-

num Lind. Plant Tissue Cult Biotechnol 18:113–122. doi:10.

3329/ptcb.v18i2.3361

Gantait S, Mandal N, Das PK (2009) Impact of auxins and activated

charcoal on in vitro rooting of Dendrobium chrysotoxum Lindl.

cv Golden Boy. J Trop Agric 47:84–86

Gantait S, Mandal N, Bhattacharyya S, Das PK (2010) An elite

protocol for accelerated quality-cloning in Gerbera jamesonii

Bolus cv. Sciella. In Vitro Cell Dev Biol Plant 46:537–548.

doi:10.1007/s11627-010-9319-2

Gantait S, Mandal N, Bhattacharyya S, Das PK (2011) Induction and

identification of tetraploids using in vitro colchicine treatment of

Gerbera jamesonii Bolus cv. Sciella. Plant Cell Tissue Organ

Cult 106:485–493. doi:10.1007/s11240-011-9947-1

Gantait S, Sinniah UR, Mandal N, Das PK (2012) Direct induction

of protocorm-like bodies from shoot tips, plantlet formation,

and clonal fidelity analysis in Anthurium andreanum cv.

CanCan. Plant Growth Regul 67:257–270. doi:10.1007/

s10725-012-9684-4

He F, Wang M, Gao M, Zhao M, Bai Y, Zhao C (2014) Chemical

composition and biological activities of Gerbera anandria.

Molecules 19:4046–4057. doi:10.3390/molecules19044046

Imaseki H (1985) Hormonal control of wound-induced responses. In:

Pharis RP, Reid DM (eds) Hormonal regulation of development.

Springer, New York, pp 485–512. doi:10.1007/978-3-642-

67734-2_14

Inoue T, Toyonaga T, Nagumo S, Nagai M (1989) Biosynthesis of

4-hydroxy-5-methylcoumarin in a Gerbera jamesonii hybrid.

Phytochem 28:2329–2330. doi:10.1016/S0031-9422(00)97977-9

Liu SZ, Feng JQ, Wu J, Zhao WM (2010) A new monoterpene-

coumarin and a new monoterpene-chromone from Gerbera

delavayi. Helv Chim Acta 93:2026–2029. doi:10.1002/hlca.

201000017

Martin KP, Zhang CL, Hembrom ME, Slater A, Madassery J (2008)

Adventitious root induction in Ophiorrhiza prostrata: a tool for

the production of camptothecin (an anticancer drug) and rapid

propagation. Plant Biotech Rep 2:163–169. doi:10.1007/s11816-

008-0057-4

Murashige T, Skoog F (1962) A revised medium for rapid growth and

bioassays with tobacco tissue cultures. Physiol Plant

15:473–479. doi:10.1111/j.1399-3054.1962.tb08052.x

Murch SJ, KrishnaRaj S, Saxena PK (1997) Thidiazuron-induced

morphogenesis of regal geranium (Pelargonium domesticum): a

potential stress response. Physiol Plant 101:183–191. doi:10.

1111/j.1399-3054.1997.tb01835.x

Nagarajan A, Arivalagan U, Rajaguru P (2011) In vitro root induction

and studies on antibacterial activity of root extract of Costus

igneus on clinically important human pathogens. J Microbiol

Biotechnol Res 1:67–76

Pandey VP, Cherian E, Patani G (2010) Effect of growth regulators

and culture conditions on direct root induction of Rauwolfia

serpentina L. (Apocynaceae) Benth by leaf explants. Trop J

Pharm Res 9:27–34

Qiang Y, Chen YJ, Li Y, Zhao J, Gao K (2011) Coumarin derivatives

from Gerbera saxatilis. Planta Med 77:175–178. doi:10.1055/s-

0030-1250176

Rothe SP, Bokhad MN, Kakpure MR (2011) Medicinal significance

of ornamental plants in human welfare from Akola district

(M.S.) India. J Exp Sci 2:30–32

Acta Physiol Plant

123

Samaj J, Bobak M, Kubosnikova D, Kristin J, Kolarik E, Ovecka M,

Blehova A (1999) Bundle sheet cells are responsible for direct

root regeneration from leaf explants of Helianthus occidentalis.

J Plant Physiol 154:89–94

SAS Institute, Inc. 1st Printing SAS/STATS User’s Guide Version 8,

vol. 2. SAS Institute, Inc., Cary; 1999

Shrivastava N, Padhya MA (1995) Punarnavine profile in the

regenerated roots of Boerhaavia diffusa from leaf segments.

Curr Sci 68:653–656

Singh AK (2006) Flower crops: cultivation and management. New

India Publishing, India, pp 127–145

Teixeira da Silva JA (2003) Thin cell layer technology for induced

response and control of rhizogenesis in chrysanthemum. Plant

Growth Regul 39:67–76. doi:10.1023/A:1021854320969

Teixeira da Silva JA (2013) In vitro rhizogenesis in papaya (Carica

papaya L.). J Plant Dev 20:51–55

Tennant PF (2010) Transgenic papaya. Transgenic Plant J 4(SI

1):1–96

Tuan NS, Ngoc HM, Mai NT, Quyen NT, Hai NT, Nhut DT (2006)

Shoot and root direct regeneration from cultured leaf segment of

Lisianthus (Eustoma grandiflorum) in vitro. TC Cong Nghe Sinh

Hoc 4:249–256

Van Tran Thanh M, Chlyah H, Chlyah A (1974) Tissue culture and

plant science. Academic Press, London, p 174

Acta Physiol Plant

123