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In Vitro Plant Regeneration of Lentil (lens Culinaris Medik.) Using Leaf and Nodal

S. SULTANA
Department of Genetics and Plant Breeding, Hajee Mohammand Danesh Science and Technology University, Dinajpur

U. HABIBA
Department of Biotechnology, Bangladesh Agriculture University, Mymansingh

T. AFROZ
Department of Plant Pathology, Hajee Mohammand Danesh Science and Technology University, Dinajpur, Bangladesh

Abstract/ Executive Summary:

The investigation was carried out in the Tissue Culture Laboratory of the Department of Genetics and Plant Breeding, Hajee Mohammad Danesh Science and Technology University, Dinajpur, Bangladesh during November, 2009 to April, 2010 with a view to study in vitro regeneration of Lentil (Lens culinaris Medik.). For this purpose, three varieties of Lentil viz. BARI Masur-3, BARI Masur-4 and BARI Masur-5 & different concentrations and combinations of hormones (2, 4-D, NAA, BAP, Kn and IAA) were used to assess regeneration ability by using two explants (leaf disc and nodal segments). Leaf disc and nodal segments of the three genotypes of Lentil were cultured on MS medium with different concentrations and combinations of growth regulators. Among the three varieties, BARI Masur-4 showed early callusing (17.67 days) with maximum rate of callus induction (83.33%) incase of leaf disc culture and also showed highest percentage of callus induction (80.55%) with minimum days (17.33 days) with nodal segment as explant. Early and maximum rate of callusing appeared in (MS + 1.5 mg/L 2, 4-D + 0.5 mg/L BAP) and (MS + 2 mg/L 2, 4-D + 0.5 mg/L NAA) from leaf disc and nodal segment respectively. BARI Masur-4 had the highest percentage of shoot regeneration from leaf disc (69.44%) and nodal segment (66.66%) explants. Early and maximum rate of regeneration was found in (MS + 2.0 mg/L Kn + 0.2 mg/L NAA) and (MS medium + 0.5 mg/L BAP + 0.25 mg/L Kn). The highest number of roots per shoot was counted in BARI Masur-4 (47.23%), in (MS medium containing 10 mg/L IAA) and (MS medium containing 15 mg/L IBA) considering leaf disc and nodal segment as explant. Considering the overall performance, BARI Masur-4 appeared to be the best genotype for callus formation, shoot regeneration and root formation.

Keywords: In vitro, Regeneration, Tissue culture, Lentil, Lens culinaris

Location: Tissue Culture Laboratory of the Department of Genetics and Plant Breeding, HSTU, Dinajpur

Start Date: 00-11-2009

End Date: 00-04-2010

Program Area: Crop-Soil-Water Management

Commodity: Lentil

Objectives:

To study in vitro regeneration of Lentil (Lens culinaris Medik.).

Methodology:

Location, time and duration of the experiment: The present study was conducted in the Tissue Culture Laboratory of the Department of Genetics and Plant Breeding, HSTU, Dinajpur, Bangladesh during November 2009 to April 2010. Experimental materials: The experimental material used in the present investigation is the seed of three Lentil (Lens culinaris Medik.) varieties viz. BARI Masur-3, BARI Masur-4 and BARI Masur-5 were used to study different parameters associated with plant regeneration. Sources of the experimental materials: The seed materials of three Lentil (Lens culinaris Medik.) varieties were collected from the Bangladesh Agricultural Development Corporation (BADC), Dinajpur, Bangladesh. Methods: Various culture media were used in the present investigation depending on specific purposes. A list of them is given below: A. For Seed Germination: Half strength MS medium medium was used for seed germination. B. For callus induction: T₁ = MS medium containing 1 mg/L 2, 4-D, T₂ = MS medium containing 1.5 mg/L 2, 4-D + 0.5 mg/L BAP, T₃ = MS medium containing 2 mg/L 2, 4-D + 0.5 mg/L NAA. C. For Shoot regeneration: T₁ = MS medium containing 1.5 mg/L Kn, T₂ = MS medium containing 2.0 mg/L Kn + 0.2 mg/L NAA, T₃ = MS medium containing 0.5 mg/L BAP+ 0.25 mg/L Kn. D. For root formation: T₁ = Hormone free 0.5 strength MS medium, T₂ = MS medium containing 15 mg/L IBA, T₃ = MS medium containing 10 mg/L IAA. Sterilization of experimental materials (Seed): Matured seeds of three varieties of lentil were washed in running tap water for 3-5 minutes for two or three times to remove the surface organism. The floating seeds were discarded. Later the seeds were dipped in 70% ethyl alcohol for 3-5 minutes with gentle shaking followed by washing with sterile distilled water. Surface disinfections was done by the use of sodium hypochlorite solutions (1% active chlorine) containing 1-2 drops of tween-20 for ten minutes with gentle shaking and then rinsed five times in sterile distilled water. These sterilized seeds were then ready for keeping in the MS media. Recording of Data: Number of explants with callus (Percent callus induction) The number of explants producing in each vial was recorded. The percentage of callus induction was calculated on the basis of the number of explants placed and total number of callus induced. Number of callus with shoot was recorded and the percentage of shoot regeneration was calculated by the formula. Average number of shoots with roots calculated by using the formula. Statistical analysis of data: The data for the parameters under present study were statistically analyzed wherever applicable. The Duncan's Multiple Range Test (DMRT) compared the analysis of variance for different parameters.

Source of Information: Int. J. Sustain. Crop Prod. 7(3): 1-7, ( 2012)

Article Link (Online Journal):

Funding Source:

1. Government

Budget:

Total Budget (Tk.) :

Achievement/Findings:

BARI Masur-4 showed best performance followed by BARI Masur-5 and BARI Masur-3. When the combined effect of explant was considered, leaf showed better performance than nodal segment. Incase of callus induction MS + 1.5 mg/L 2, 4-D + 0.5 mg/L BAP and MS + 2 mg/L 2, 4-D + 0.5 mg/L NAA, for shoot induction MS + 2.0 mg/L Kn + 0.2 mg/L NAA and MS medium + 0.5 mg/L BAP + 0.25 mg/L Kn, for root initiation MS medium + 10 mg/L IAA and MS medium containing 15 mg/L IBA were best with leaf disc and nodal segment explant respectively.

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