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Clonal Propagation of Dracaena Fragrans Cv. Victoria Through Tissue Culture Technology

Ahshan Jazib
Department of Botany, Jahangirnagar University, Savar, Dhaka-1342, Bangladesh

Mohd Talim Hossain
Department of Botany, Jahangirnagar University, Savar, Dhaka-1342, Bangladesh

Raihan I Raju
Department of Botany, Jahangirnagar University, Savar, Dhaka-1342, Bangladesh

Abstract/ Executive Summary:

Micropropagation of Dracaena fragrans cv.Victoria was conducted using the young, tender and disease-free leaves and nodal segments as explants collected from the local market of Savar, Dhaka. Surface sterilization of the explants pretreated with a liquid detergent and then 0.2% HgCl₂ for 4-5 minutes produces maximum contamination free explants without any toxicity. After surface sterilization, different explants were inoculated on gelrite gelled MS medium supplemented with different concentrations of 2,4-D for callus induction and with different concentrations and combinations of BAP and NAA for direct shoot induction. Nodal explants showed high callus induction potentiality (80%) on MS medium supplemented with 1.5 mg/l 2,4-D. The highest frequency of direct shoot induction from nodal segment was 80% and the number of shoots per nodal segment was(5.28±1.17) when they were cultured on MS medium supplemented with 3.0 mg/l BAP and 0.3 mg/l NAA. The highest shoot multiplication (83.33%) with maximum number of shoot per unit callus (5.62±1.24) and maximum shoot length (3.27±0.82 cm) was observed when the nodal calli were transferred in gelrite gelled MS medium in combination with 4.5 mg/l BAP and 0.5 mg/l NAA. Additionally, the incorporation of 4% sucrose and 10% coconut water with the above mentioned medium showed the satisfactory shoot growth and development with an average 7.84±1.30 shoots per unit of callus which was 4.21±0.78 cm in length. Moreover, addition of 3.0 mg/l GA₃ with the above mention medium showed highest rate of shoot elongation (5.83±2.31cm). For root induction, in vitro raised shoots were transferred onto half-strength of MS liquid medium augmented with different concentrations and combinations of auxins (IBA and NAA). Maximum rooting (75%) were observed in halfstrength MS liquid medium supplemented with 0.5 mg/l IBA. After appropriate rooting the plantlets were successfully acclimatized (85% survival) when they were cultured in polybag containing (1:1:1) garden soil, sand and compost mixture before transferred to soil. Regenerated plants were morphologically identical with mother plants and showed their uniform growth in field condition.

Keywords: Clonal propagation, Dracaena fragrans, Tissue culture technology

Location: Local market of Savar, Dhaka

Start Date:

End Date:

Program Area: Development of Host and Medicinal Plants

Commodity: Medicinal Plants

Objectives:

To establish an efficient protocol for direct and indirect shoot regeneration, rapid multiplication of shoots, rooting of in vitro raised shoots and acclimatization of the in vitro raised plantlets of Dracaena fragrans cv. Victoria under in vivo condition.

Methodology:

Plant materials: Healthy and disease free plants of Dracaena fragrans cv. Victoria were collected from the local market of Savar, Dhaka and used as mother plant. Two type of explants such asnodal segments (2-3 cm) and young leaf disc (1 cm 2 )were used as the experimental material. Surface sterilization: For surface sterilization, explants were washed under running tap water for 30-45 min., followed by washing in sterilized distilled water with several drops of liquid detergent for 20 min. and then washing with autoclaved distilled water for 3 times. Further sterilization of the explants have been done under laminar airflow with (0.1% and 0.2%) HgCl₂ solutions for 3-7 min. to ensure contamination free culture. After sterilization, they were rinsed several times with sterile distilled water and used as explants. Cut ends of the explants which have been in contact with HgCl₂ solution were removed by cutting with a sterilized scalpel. Initial culture medium: For the initiation and proliferation of callus, nodal segment (2-3 cm) and leaf explants (1 cm² ) were inoculated singly in the culture bottles containing MS basal medium (Murashige & Skoog, 1962) supplemented with different concentrations of 2,4-D (1.0-4.0 mg/l). On the other hand, nodal segments (2-3 cm) were inoculated individually on MS basal medium supplemented with different concentrations of BAP alone or in combinations with NAA for direct shoot induction. Shoot induction and multiplication: Different explants derived callus were then excised into small pieces and transferred onto gelrite gelled MS medium containing different concentrations and combinations of cytokinins (BAP, Kn, TDZ and 2-iP) and auxin (NAA) for induction and proliferation of shoot. At this stage, the induced shoots were recovered aseptically from the culture vessels and small shoots were again cultured on freshly prepared medium containing same or different concentrations and combinations of hormonal supplements for shoot development and subsequent multiplication of shoots. Data were scored in terms of number and percentage of responsive callus, number of shoots per culture and shoot length. For subsequent multiplication of regenerated shoots the effect of different concentrations of sucrose (1-5%) and coconut water (5-20%) were also observed. The cultures were exposed to 16 hours light and 8 hours dark per day with constant temperature at 24 ± 2°C. The light intensity of the growth chamber was 3000 lux. Overall, an aseptic environment was maintained throughout the whole process. Rooting in solid and liquid MS medium: After three times of the sub-culturing the in vitro raised shoots obtained from the multiplication media with 5-6 cm in length, were excised aseptically from the culture vessels and implanted separately on freshly prepared solid or liquid half strength MS media for rooting containing different concentrations and combinations of auxins (IBA and NAA). For root induction, the newly transferred cultures were kept in dim light for 3 days and then they were kept in light. Data will be scored in terms of root development percentage (%), root number per shoot and root length (cm). The plantlets with well developed root system will then be acclimatized. Acclimatization of the regenerated plants: The in vitro micropropagated plantlet is not readily fit for field plantation. For acclimatization, the in vitro developed plantlets have been removed from the culture vessels and washed out all medium residues from the roots. Then they were implanted in plastic pots, containing garden soil, sand and compost with 1:1:1 ratio; and thoroughly covered with pored polythene bags and sprayed with water at every 8 hours to maintain high humidity. By this process the plantlets have been established in the soil.

Source of Information: Jahangirnagar University J. Biol. Sci. 8(2): 1-11, 2019 (December)

Article Link (Online Journal): DOI: https://doi.org/10.3329/jujbs.v8i2.49833

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

The present investigation was carried out in order to find out suitable explants for culture establishment, to standardize method for their surface sterilization, to evaluate different concentrations of growth regulators for shoot multiplication, rooting and hardening. This protocol may be helpful for rapid propagation of Dracaena. However, further research is needed to get fuller benefit of the technique in large-scale commercial application.

Knowledge Management: Journal