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In Vitro Regeneration of Potato (solanum Tuberosum L.)

M. M. H. Molla
Biotechnology Division Bangladesh Agriculture Research Institute Gazipur 1701, Bangladesh.

K. M. Nasiruddin
Biotechnology Division Bangladesh Agriculture Research Institute, Gazipur 1701, Bangladesh.

M. Al-Amin
Biotechnology Division Bangladesh Agriculture Research Institute, Gazipur 1701, Bangladesh.

M. S. Haque
Biotechnology Division Bangladesh Agriculture Research Institute, Gazipur 1701, Bangladesh.

D. Khanam
Biotechnology Division Bangladesh Agriculture Research Institute, Gazipur 1701, Bangladesh.

Abstract/ Executive Summary:

The experiment was conducted during the period from January to December, 2008 at the Laboratory of Biotechnology, Bangladesh Agricultural Research Institute, Gazipur, Bangladesh. A two step procedure was followed for direct plant regeneration of potato. Murashige and Skoog (MS) basal medium supplemented with 0.5 mg l^-1IBA and 30 g sugar was used for in vitro potato plants development via nodal cuttings. Leaves and internodes of in vitro grown potato variety Asterix was cultured in instant MS basal medium supplemented with 0.01 mg l^-1 IAA, 0.20 mg l^-1 GA3, 44.0 g l^-1 CaCl₂, 20 g sucrose. Eight different concentrations of Zeatin riboside (ZR) viz., 0, 0.1, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0 mg l^-1along with above supplements (step II) were tested for in vitro direct regeneration of potato. MS medium supplemented with 4-5 mg l^-1of ZR performed better in respect of shoot induction from internodal explants. All the internodes produced shoots directly from the basal and apex region within 18-21 days and from the leaves within 21-28 days in 2 - 5 mg l-1 ZR. Maximum 43.20 and 7.25 shoots were recorded from each internode and leaf explant, respectively. In vitro shoots treated with 0.5 mg l-1 of IBA produced roots profusely within 21 days.

Keywords: Direct regeneration, Zeatin riboside (ZR), Xplant, Internode, Leaf, IBA, Rooting and potato

Location: BARI

Start Date: 00-01-2008

End Date: 00-12-2008

Program Area: Variety and Species

Commodity: Potato

Objectives:

To find out an optimum concentration of ZR for direct in vitro regeneration of potato from internode and leaf explants

Methodology:

In vitro ready plant lets of potato cv. Asterix was used as source material. The in vitro plant lets were propagated on MS medium supplemented with 0.5 mg l^-1 IBA and 3% sugars. The cultures were incubated 16 h ours under light in a room with temperature 21±1ºC with 2500-3000 lux. After 18 - 21 days, in vitro plants were ready for explants collection. For culturing the explants, 4.4_g of instant MS basal medium (Duchefa, Netherlands) and 20 _g sucrose (Biobasic, Canada) were used for preparing one liter medium. GA3 (0.20 mg l^-1) and IAA (0.01 mg l^-1) were used according to Cearley and Bolyard (1997). The pH was adjusted to 5.8 before autoclaved and 2 g gelrite (Biobasic, Canada) was added to solidify the medium. Then the medium was autoclaved at 1.06 kg/cm2 with 121ºC for 20 minutes. Zeatin riboside in the medium through filter sterilization and mixed in the medium under the laminar air flow hood when the temperature of medium was come down near about 50ºC. Then 20 ml medium was dispensed into each petri dish and covered by upper plate when the medium was cool down and rapped by Para film. Petri dishes were marked as per treatment and kept in 4ºC. Eight different concentrations of ZR viz., 0, 0.1, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0 mg l^-1mg l^-1 along with GA₃ and IAA were tested in this study. Explants (internode) were collected from in vitro plant lets of 18 - 21 days old. Only the first 5-6 internodes (0.5 – 1.0 cm long) from the top of the plantlet excluding shoot apex was excised. For leaf explants, thick and healthy leaf blades from the upper nodes of the plants excluding tips and basal portions were cut into 5×5 mm pieces. The leaf explants were placed upside down on the medium. Both the leaf and internode explants, fifteen and five were placed at every Petri dish, respectively in the above mentioned medium. Petri dishes were para filmed and placed at 23±1ºC with 16/8 hrs light/dark fluorescent light. Shoots regenerated from the 5 mgl^-1 ZR treated explants were transferred to half strength MS medium havinh different concentration of IBA (0.5, 1.0, 1.5 and 2.0 mgl^-1). Sometimes rooting is prohibited due to the effect of ZR. To reduce the ZR effect, explants were cultured 2 weeks on hormone free MS medium. The cultures were checked daily to note the response and growth of explants. The experiment was designed in CRD with 7 replications. Each petri dish was considered as single replication which contained 15 and 10 internodal and leaf explants, respectively. Stem diameter was measured by a digital slide calipers. Results were analyzed using MSTAT-C statistical package. Differences among the means were compared by the Duncan’s Multiple Range Test at 1% level of significant. The percentage data were subjected to appropriate transformation like arcsine and squire root system according to Gomez and Gomez (1984). The regenerated rooted plant lets with 4-5 leaves stage were transferred from culture room and kept in room temperature (approx. 300C) for 4-5 days. The plant lets were then removed from the culture tubes and all the adhering media were carefully washed out so that, the root damage was the least. Washed plant lets were planted into small plastic pots containing sterile soil, sand and decomposed cow dung at the ratio of 1: 1: 1.

Source of Information: Bangladesh J. Pl. Breed. Genet, 24(1): 07-14

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

Based on the above results, it may be concluded that MS medium supplemented with 5 mg l^-1ZR performed best for in vitro direct regeneration of potato from internodal and leaf explants. Moreover, half strength MS medium supplemented with 0.5 mg l^-1IBA produced well developed roots within 21 days which gave the 100% survival at ex vitro condition (Plate 3).

Knowledge Management: Journal