Shoot tip culture of BARI banana 1 (Musa sp.) were cultured on MS medium supplemented with 5.0 mg L-1 BAP for shoot proliferation. Well-developed shoots were used for rooting. Among the six different concentrations of IBA (0.1, 0.2, 0.3, 0.4, 0.5, 0.6 mg L-1) with half strength MS medium, a good number of healthy roots were produced on half MS+0.5 mg L-1 IBA (7.86) followed by half MS+0.6 mg L-1 IBA (6.89) and half MS+0.4 mg L-1 IBA (6.31) with the weight of 0.85, 0.83 and 0.77 g plantlet-1, respectively. However, 95-100% plantlets were survived when they were transferred to small plastic pots after 15-20 days in vitro culture on half MS medium supplemented with 0.40.6 mg L-1 IBA and 7 days hardening at room temperature.
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Banana (Musa sp.) is a most important fruit crop in Bangladesh. The plants are propagated vegetatively by means of sucker, which is a slow process. Moreover, suckers are sometimes infected with banana bunchy top virus as a symptomless carrier, Sigatoka and Panama diseases. Application of tissue culture technique is, therefore, a tool to produce large number of true to type disease free plants in limited period of time and space. Apical meristem culture offers an efficient method for rapid clonal propagation, production of pathogen free material and germplasm preservation in plants[2-10]. Shoot tip culture is a well established adequate and relatively simple in vitro method for the rapid propagation of selected musa materials and the clean planting material. A wide range of plant tissue culture techniques is increasingly being used as an enabling and enhancing technology for the handling of musa germplasms. But in vitro propagation of different cultivar required different culture media for shoot proliferation and root differentiation. However, there is still lack of information on in vitro rooting of banana. Now a day, the plant growth regulators are widely used in modern agriculture to promote rooting. Widiastoety and Soebijanto reported that good rooting and the best survival (96.6%) were obtained with IBA treatment in Hibiscus rosa sinensis. Kundu et al. reported that indole 3-butyric acid (IBA) had a highly significant effect on the percentage success of rooting, number of root and length of root production in Ixora coccinia. Percentage of rooting, number of roots and length of roots were markedly increased with IBA treatment in seedless lemon. Hence, the present study was undertaken to standardize IBA concentration and days for in vitro rooting with 7 days hardening at room temperature for better plant survival of BARI banana-1 in ex vitro condition.
MATERIALS AND METHODS
BARI Banana 1 (Musa sp.) is a high yielding and newly released banana variety in Bangladesh. Shoot tips were collected from the virus free sword sucker of the variety. Roots and outer layer of tissues of the suckers were removed and the remaining portions were washed with tap water using detergent. The explant then surface sterilized in 0.1% mercuric chloride (HgCl2) with a few drops of Tween 20 for 30 min. After washing 4 times with sterilized distilled water, the shoot tips were further cut to a size of approximately 5x8 mm portion containing an intact apex under clean bench. The explants were placed on MS solid medium with 5.0 mg L-1 BAP. All cultures were incubated at 25±1°C with a 16 h photoperiod (2000 lux) provided by cool white fluorescent tubes. The pH of the medium was adjusted to 5.8 prior to autoclaving. The materials were sub-cultured at 30 days interval in the same medium to produce multiple shoots. Well-developed shoots were transferred to rooting medium containing half MS medium supplemented with different concentrations of IBA (0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6 mg L-1). Each treatment contained ten replicate with six shoots/replicate. Every five days interval plantlets from different concentrations, transferred at room temperature for seven days hardening. After that, the plantlets were washed in water carefully to remove the media and transferred to small plastic pots containing a mixture of sterilized soil and sand for five days where high humidity was maintained by covering the plantlets with polyethylene sheet. The plantlets were then planted in poly bags (12.5x17.5 cm) containing soil, sand and cowdung (1:1:1) and were kept in nethouse. After incubation in rooting medium every 5 days interval, number of roots were recorded. After 15 days in vitro culture with 7 days hardening at room temperature average number of roots, length of roots, fresh weight of roots, number of leaves and length of shoots were recorded.
RESULTS AND DISCUSSION
Shoot proliferation and growth patterns of cultured explants are shown in Fig. 1(a-d). Rooting initiation was started within 3-4 days after incubation in half MS medium supplemented with 0.5 mg L-1 IBA where 4-5 days were needed in half MS medium supplemented with 0.1, 0.2, 0.3 and 0.4 mg L-1 IBA. Five to six days were needed in half MS medium supplemented with 0.6 mg L-1 IBA and in control treatment (Table 1). Khanam et al. observed root initiation within 3-4 days with 2 μM L-1 IBA in banana cv. Amritsagar. Number of roots on different concentration of IBA (0.0, 0.1, 0.2, 0.3, 0.4, 0.5 and 0.6 mg L-1) are given in Table 1. In every concentration, number of roots was increased with increasing the duration of incubation in half MS supplemented with IBA medium.
Half MS medium without IBA also produced roots (2.44) but their growth was very poor. Half MS + 0.5 mg L-1 IBA produced highest number (8.28) of roots followed by half MS + 0.6 mg L-1 IBA medium (7.33) and half MS+0.4 mg L-1 IBA (6.33). Half MS+0.1 mg L-1 IBA, half MS+0.2 mg L-1 IBA and half MS+0.3 mg L-1 IBA were produced more or less similar number of roots such as 3.89 and 3.97, respectively (Table 1). After 25 days of in vitro culture, the initial roots were changed in colour and new roots initiation was stopped except secondary and tertiary root initiation.
Length of root was not increased with the increasing of IBA concentration in half MS medium and the range was 2.605.67 cm (Table 2). But fresh weight of roots/plantlet was increased with the increasing of IBA concentration in half MS medium. Highest fresh weight of roots were recorded on half MS+0.5 mg L-1 IBA concentration (0.85 g) followed by half MS +0.6 mg L-1 IBA concentration (0.83) and half MS+0.4 mg L-1 IBA concentration (0.77 g). Lowest fresh weight of roots/plantlet was recorded in half MS medium without IBA (0.23 g). On the other hand, half MS medium with 0.1, 0.2 and 0.3 mg L-1 IBA concentration produced fresh wt. of roots/plant 0.30, 0.31 and 0.35 g, respectively where as poor developed or unsatisfactory fresh weighted roots were produced on 0.0-0.3 mg L-1 IBA supplemented half MS media. Good rooting was observed on 0.4-0.6 mg L-1 IBA concentration on half MS medium.
|Table 1:||Effect of different concentrations of IBA and duration of in vitro culture on rooting of BARI Banana 1|
|Table 2:||Growth and development of plantlets as influenced by IBA at 15 days in vitro culture with 7 days hardening|
|Fig. 1(A-D):||Different stages of in vitro regeneration and rooting of banana|
|(A) Explant initiation (B) Multiple shoot (C) Rooted plantlet (D) Established plantlets in plastic bags|
|Table 3:||Days to in vitro culture with different conc. of IBA and 7 days hardening at room temp. on survival percentage of BARI banana 1 plantlets|
Number of leaves and length of shoots were varied from 5.20 6.40 and 8.12-8.35 cm, respectively.
Plantlets transferred to plastic pots after 10, 15, 20, 25 and 30 days in vitro culture with different concentrations of IBA and 7 days hardening are shown in Table 3. Results indicated that 95-100% plantlets survived when they were transferred after 15 -20 days in vitro culture on 0.4-0.6 mg L-1 IBA concentration with 7 days hardening at room temperature.
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