In Vitro Regeneration and Multiplication of Jackfruit
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M. Ashrafuzzaman et al / Research Journal of Biology (2012), Vol. 02, Issue 02, pp. 59-65 ISSN 2049-1727
Research Paper
In Vitro Regeneration and Multiplication of Jackfruit
(Artocarpus Heterophyllus L.)
M. Ashrafuzzaman*1, Sukarna Kar1, Dilafroza Khanam2 and Shamsul Haque Prodhan1
1
Department of Genetic Engineering and Biotechmology, Shahjalal University of Science and Technology, Sylhet,
Bangladesh
2
Biotechnology Division, Bangladesh Agricultural Research Institute (BARI), Joydebpur, Gazipur 1701, Bangladesh
*E-Mail: azamanbt@gmail.com ; azamangeb-gen@sust.edu
Abstract
Jackfruit (Artocarpus heterophyllus L.), known as national fruit of Bangladesh, found between June to August. Propagation
of jackfruit plant from seeds is not widely accepted because of high heterozygosis. To maintain the true type quality fruit,
tissue culture technique could be used for the propagation of jackfruit throughout the year. Healthy and juvenile shoot tips
were used for explanting purpose and cultured in Murashige-Skoog (MS) medium complemented with different
concentrations (0 mg/L, 1 mg/L, 2 mg/L, 3 mg/L and 4 mg/L) of plant growth hormone i.e. BAP (6-benzyleaminopurine),
for the purpose of multiple shoot development. Regeneration of shoots increased comparatively better when MS medium
was enriched with 2 mg/L of BAP. With the increase of subculture (up to 10th maximum), frequency of shoot proliferation
was enhanced. These shoots were then cultured on half strength of MS medium supplemented with 0 mg/L, 1 mg/L, 2
mg/L, 3 mg/L and 4 mg/L IBA (Indole-3-butyric acid) and observed that 2 mg/L IBA containing medium was highest in
number of roots/explants, root length and early root induction.
Keywords: Jackfruit (Artocarpus heterophyllus L.), In vitro, Micro-Propagation, Shoot Tip
1. Introduction
Jackfruit is a dicotyledonous compound fruit of the jack proteins, vitamins and minerals. The juicy pulp of the ripe
tree (Artocarpus heterophyllus L.) which belongs to the fruit could be eaten either fresh or preserved in syrup. In
family Moraceae grow in many of the tropical countries of the tropical and sub-tropical areas, the seeds, budding,
Southeast Asia but is particularly abundant in India and inarching, air layers, cutting of young wood and tissue
Bangladesh. Fruit usually reach 10-25 kg in weight, grows culture (Morton, 1987), could propagate the jackfruit. The
in summer when staple food-grains are often in short jackfruit tree could be bearing fruits twice yearly (Roy et
supply. The edible portion is considered as a good source al, 1996) which could play a significant role to improve the
of carbohydrate, proteins, vitamins and minerals. The juicy economy of a country. The jackfruit most acceptable in the
pulp of the ripe fruit could be eaten either fresh or full-grown but unripe stage, when it has no objectionable
preserved in syrup. Jackfruit is a dicotyledonous odor and excels cooked green breadfruit. The fruit at this
compound fruit of the jack tree (Artocarpus heterophyllus time is simply cut into large chunks for cooking. The
L.) which belongs to the family Moraceae, grown in many chunks are boiled in lightly salted water until tender, when
of the tropical countries of Southeast Asia but is the really delicious flesh is cut from the rind and served as
particularly abundant in India and Bangladesh. Fruit a vegetable, including the seeds which, if thoroughly
usually reach 10-25 kg in weight, grows in summer when cooked, are mealy and agreeable (Morton, 1987).
staple food-grains are often in short supply. The edible
portion is considered as a good source of carbohydrate, The formation of adventitious shoots or roots was first
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M. Ashrafuzzaman et al / Research Journal of Biology (2012), Vol. 02, Issue 02, pp. 59-65 ISSN 2049-1727
determined by Skoog and Miller (1957) through discovery 2.1. Explants Collection
of the regulation of organ formation (shoots and roots) by
changing the ratio of cytokinin/auxin. When ratio of The explants material (5 cm long shoot tips) for the
cytokinin/auxins is high, it favours the formation of shoot experiment was collected from healthy jack tree.
but root formation is inhibited. The reverse favours the
root formation. The compounds that are most frequently 2.2. Surface Sterilization of Explants
used are called auxins, chemically called as 2,4-
dichlorophenoxy acetic acid (2,4-D) and cytokinins are Jackfruit explants were surface sterilized at first using 3
kinetin, benzyl adenine (BA), 6-benzylaminopurine (BAP) g/L mercuric chloride for 1 minute, and thoroughly rinsed
and 2-isopentenyl aminopurine (2ip) (Chawla, 2002). with sterilized distilled water for three times then
continued surface sterilization using 5.25% sodium
Many investigators, such as Adiga et al (1998), studied in hypochlorite for 20 minutes.
vitro propagation (tissue culture) of the jackfruit. They
reported that sucrose was a good source of Carbon, when 2.3. Sterilization of Equipments
media cultures supplemented with sucrose or sugar
produced more shoots and Gibberelic acid (6.0 mg/L) Glassware was scrubbed with brush in a hot detergent
increased shoot length. Singh and Tiwari (1996) bath. They were washed thoroughly with tap water and
mentioned that jackfruit was successfully micro then rinsed two to three times with distilled water and if
propagated by culturing nodal segments on modified MS necessary autoclaved shortly in order to liquefy the agar
medium containing plant growth regulators i.e. and to kill any contaminates that may be present. All the
combination of 18 mg/L of BA (benzyladenine) + 0.2 glasswares that needed cleaning were soaked in detergent
mg/L of IBA (Indole-3-butyric acid). The highest number water for 2 hours followed by repeated washing in tap
(4-5) of usable shoots was developed on nodal segments water to remove components of detergent. Petri dishes,
taken from in vitro proliferated shoots by enhancement of beaker etc. were wrapped with aluminum foil and left in
auxiliary branches after 4 subcultures on MS with 2 mg/L the autoclaved at 121ºC, 15 1b, for 20 minutes for
BA + 0.2 mg/L IBA, in vitro grown successful rooted sterilization.
(58.7%) in half-strength MS with 1 mg/L. Rajmohan and
Mohankumaran (1988) mentioned that light regime of 2.4. Establishment Stage of Jackfruit Explants
darkness in the establishment stage for 4 weeks only was
necessary for the successful micropropagation of jackfruit. The sterilized explants had been cultured individually into
. Hazarika (2003) reported that paclobutrazol (0.5-4 mg/L) test-tube (150 ml) containing 20 ml. of the following
in the rooting medium reduced stomatal apertures and media:
wilting after transfer to compost, increased epicuticular
MS medium (control)
wax, shortened stems, chlorophyll content and thickened
roots. MS medium supplemented with 1 mg/L BAP
MS medium supplemented with 2 mg/L BAP
To improve the yield of the local varieties/cultivars; an
efficient protocol for plant regeneration needs to be MS medium supplemented with 3 mg/L BAP
developed to achieve transgenic plant. The main objectives
MS medium supplemented with 4 mg/L BAP
of the present study were the followings:
Cultures were incubated at 27 ± 2° C under light provided
To establish a suitable protocol for the
by white fluorescent tubes giving the intensity of about
micropropagation of jackfruit from shoot tip.
2000 lux for 16 hours/day. The explants were subcultured
with the same media every three weeks.
Find out the effect of growth hormone (BAP) on
Frequency of shoot induction was calculated according to
shoot proliferation.
the following formula:
Find out the effect of growth hormone (IBA) on
root induction.
Shoot Induction Frequency (%) = × 100
2. Materials and Methods
2.5. Multiplication Stage
This study was carried out at the Tissue Culture Lab of
Biotechnology division in Bangladesh Agricultural Shoot, induced explants were multiplied in regeneration
Research Institute (BARI), Gazipur, Bangladesh from media and were repeatedly subcultured onto the same
March to June in 2011. media at least twice for every 3 three weeks.
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M. Ashrafuzzaman et al / Research Journal of Biology (2012), Vol. 02, Issue 02, pp. 59-65 ISSN 2049-1727
2.6. Rooting Stage data were recorded (Table 2), after eight weeks as:
Jackfruit shoots at 3 cm length, produced from the No. of survival shoots / explants at different
multiplication stage were cultured on the half strength MS hormone concentration
basal medium supplemented with Indole-3-butyric acid
(IBA) at the concentration of (0 mg/L, 1 mg/L, 2 mg/L, 3 Shoot length in cm
mg/L and 4 mg/L). Each treatment of above experiment
was supplemented with 30 g / 1000 ml sucrose and Number of leaves/culture
solidified with 16 g/ 1000 ml agar.
Table 1. Response of Different Concentration of BAP on Ms
Medium of Shoot Induction
3. Results
BAP
There were three stages of this experiment – establishment Number of Shoots
Concentration Number of
stage, multiplication stage and rooting stage. Data were Inducing
(mg/L) Explants
recorded of each stage carefully basis on some specific Explants
criteria.
0.0 10 0
1.0 10 5
3.1. Establishment Stage
2.0 10 8
Four weeks after the establishment of jackfruit explants 3.0 10 6
data were recorded as No. of shoot inducing explants at 4.0 10 4
different concentration of hormone (Figures 1).
3.4. Shoots Number / Explants
3.2. Shoot Induction
Data obtained from Table 2 showed that, BAP at 2 mg/L
It is clear from the Table 1 that, BAP at 2 mg/L produced produced the highest significant values (100%) of number
the highest significant values of shoot inducing explants of shoots/explants comparing with other concentration 3
(8) which followed by 3 mg/L (6), 4 mg/L (4),1 mg/L (5) mg/L (90%), 4 mg/L (80%), 1 mg/L (80%)
and 0 mg/L (0). After three weeks time, the first subculture
was carried out (Figure 2 and 3). 3.5. Average Shoots Length (cm)
3.3. Multiplication Stage It was obvious from Table 2 that BAP at 2 mg/L gave the
highest significant shoot length (3.84 cm) & the lowest
Multiplication stages have been shown in Figures 4-6 and length (2.7 cm) at 1 mg/L.
1 mg/L 2 mg/L 3 mg/L 4 mg/L
Figure 1. Shoot at Different Concentration of BAP after 15 Days of Establishment
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M. Ashrafuzzaman et al / Research Journal of Biology (2012), Vol. 02, Issue 02, pp. 59-65 ISSN 2049-1727
1 mg/L 2 mg/L 3 mg/L 4 mg/L
Figure 2. After first Subculture at Different Concentration of BAP
80% 80%
Frequency of Shoot Induction (%)
60% 60%
50%
40%
40%
20%
0%
0%
0
1
2
3
4
Concentration of BAP (mg/L)
Figure 3. Frequency of Shoot Induction at Different Concentration of BAP
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M. Ashrafuzzaman et al / Research Journal of Biology (2012), Vol. 02, Issue 02, pp. 59-65 ISSN 2049-1727
Table 2. Response of Different Concentration of BAP in MS Medium on Shoot Multiplication
Number of
BAP Concentration Number of Average Length of Average Number
Shoots/Culture
of (mg/L) Explants Shoot /Culture (cm) Leaves/Culture
Forming Multiple Shoot
1.0 10 08 2.70 2
2.0 10 10 3.84 4
3.0 10 09 3.30 3
4.0 10 08 2.80 3
Figure 4. Multiplication of Explants Figure 5. Multiplied Explants
100% 100%
Parentage Of Explants Forming Multiple Shoot
90%
90%
80% 80%
70%
80%
60%
50%
40%
30%
20%
10% 0%
0%
0
1
2
3
4
Concentration of BAP (mg/ L)
Figure 6. Effect of BAP on Shoot Multiplication
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M. Ashrafuzzaman et al / Research Journal of Biology (2012), Vol. 02, Issue 02, pp. 59-65 ISSN 2049-1727
3.6. Average Leaves Number / Explant 3.8. Average Number of Roots/Explant
Results in Table 2 cleared that, BAP at 2 mg/L gave the Results in Table 3 showed the jackfruit explants cultured
highest number of leaves/explants i.e. 4; after two intervals on ½ MS medium supplemented with 2 mg/L IBA resulted
subculture & the lowest i.e. 2 at 1 mg/L while it was the in the highest number i.e. 4 of roots/explants.
same number i.e. 3, at 3 mg/L & 4 mg/L.
3.9. Average Root Length (cm)
3.7. Rooting Stage
Table 3 results also revealed that, the greatest average
Jackfruit explants were incubated for four weeks in the length (1.20 cm) of jackfruit roots was obtained on 1/2 MS
growth room (Figure 7) to determine the rooting data medium supplemented with IBA at 2 mg/L compared with
(Table 3): other treatment.
1. Days to root induction 4. Discussion
2. No. of roots/explants
3. Root length in cm Any in vitro propagation method developed, should be
Table 3. Effect of Different Concentration of IBA in Half Strength MS Medium on the Initiation & Development of Jackfruit Roots
IBA Concentration Number of Average Number of Average Length of
Days to Roots Induction
of (mg/L) Explants Roots Root / Culture (cm)
0.0 10 No Induction 0.0 0.0
1.0 10 12-17 2.7 0.93
2.0 10 10-15 4.0 1.20
3.0 10 13-16 3.2 1.10
4.0 10 13-17 2.6 0.98
Data in Table 3 revealed that the early induction (10-15 easy to adopt and achieve high multiplication rate. Success
days) of jackfruit roots was obtained on ½ MS medium in micro-propagation of woody perennials has been very
supplemented with IBA at 2 mg/L as compared with other limited due to the problems such as contamination,
treatments & supplemented with 4 mg/L on 1/2 MS phenolic exudation, vitrification, induction of rooting,
medium take the comparatively long time (13-17 days). acclimatization etc. Jackfruit being woody perennial shows
1 mg/L 2 mg/L 3 mg/L 4 mg/L
Figure 7. Root Formation on ½ MS Medium Supplemented with Different Concentration of IBA Days to Root Induction
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M. Ashrafuzzaman et al / Research Journal of Biology (2012), Vol. 02, Issue 02, pp. 59-65 ISSN 2049-1727
many of the above undesirable characteristics. In this Plant Tissue Culture, 2, pp. 27-30.
experiment, BAP at 2 mg/L had given better response with
respect to shoot induction, shoot multiplication, length of Chawla, H.S. (2002) Introduction to plant
shoot, and number of leaves per explants. Adiga (1996) biotechnology. 2nd ed. New Delhi, Oxford & IBH
had obtained similar results in jackfruit. A hundred percent publishing Co. Pvt. Ltd., pp. 16-17.
rooting efficiency has been recorded in jackfruit with IBA
and NAA (1-Napthaleneacetic acid) (1 mg/L each) (Amin, Hazarika, B.N. (2003) Acclimatization of tissue-cultured
1992). plants. Current Science, 85 (12), pp. 1704-1712.
In jackfruit, 90% rooting was achieved on a media Islam M.S., Sen, J., Alam, N., and Roy, S.K. (1993)
consisting of ½ MS with IBA and IAA (Indole-3-Acetic Propagation of Jackfruit (Artocarpus heterophyllus)
acid) (1 mg/L each) (Islam et al, 1993). In this experiment, Through Zygotic Embryo Culture In Vitro. Short
best concentration for shooting was found to be MS Communications, Plant Tissue Cult., 3, pp. 51-55.
medium supplemented with BAP 2 mg/L and for rooting
purpose half-strength MS medium supplemented with IBA Morton, J.F. (1987) Jackfruit. In: Jackfruit of warm
2 mg/L. All other treatments initiated less shooting and climates. Miami, Julia, F. Morton, pp. 58-64.
rooting. The reason may be attributed to-Physiological
status of explants taken for shooting and rooting which Rajmohan, K., and Mohankumaran, N. (1988) Influence of
interacts with growth regulators and environmental factors. explant source on the in vitro propagation of jack.
The other reason may be lack of cell sensitivity to respond Agricultural Research, Journal of Kerala, 26, pp. 69-
to morphogenesis even though growth regulator may or 174.
may not present in abundance or in excess.
Roy, S.K., Royand, P.K., and Brumfield, R.G. (1996) In
5. Conclusion vitro propagation and establishment of a new cultivar of
jackfruit (Artocarpus heterophyllus lam.) bearing fruits
With this experiment, a protocol for the micro-propagation twice yearly. Acta Hort., 429, pp. 497-502.
of A. heterophyllus has been established from shoot tip that
could be used for cultivating jackfruit in fruit gardens and Singh, R., and Tiwari, J.P. (1996) In vitro clonal
reforestation programs and on the other hand, this finding propagation of jackfruit (Artocarpus heterophylluslam.). J.
would be a milestone for genetic transformation studies App. Hort. Nav., 2(1-2), pp. 86-90.
and conservation of endangered varieties in the developing
countries in future. Skoog, F., and Miller, C.O. (1957) Chemical regulation of
growth and organ formation in plant tissues cultivated in
vitro. Symposium Society for Experimental Biology, 11,
Acknowledgements pp. 118-131.
The authors gratefully acknowledge to the Biotechnology
Division, Bangladesh Agricultural Research Institute
(BARI) for providing the facilities to carry out this
research work.
References
Adiga, D.J. (1996) Clonal Propagation of Jackfruit
(Artocarpus heterophyllus Lam) cv. Singapore Jack
through tissue culture. Current Science, 78, pp. 1231-
1234.
Adiga, T.D., Khan, M.M., and Sathyanarayana, B.N.
(1998) Effect of carbon source, culture vessels, gelling
agents and GA 3 on in vitro shoot proliferation of
Singapore jack (Artocarpus heterophyllus lam.).
Karnataka J. Agric. Sci., 11(3), pp. 632-636
Amin, M.N. (1992) In vitro enhanced proliferation of
shoots and regeneration of plants from explants of jack.
Available online at www.scientific-journals.co.uk 65
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