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Assessment of Genetic Diversity of Chili Cultivars Using Rapd Markers

M. M. Uddin
Department of Biotechnology, Bangladesh Agricultural University, Mymensingh

M. I. Khalil
Plant Pathology Division, Bangladesh Institute of Nuclear Agriculture, Mymensingh

M.S. Haque
Department of Biotechnology, Bangladesh Agricultural University, Mymensingh

M. B. Meah
Department of Plant Pathology, Bangladesh Agricultural University, Mymensingh

Abstract/ Executive Summary:

Random amplified polymorphic DNA (RAPD) assay was performed to estimate genetic polymorphisim in ten chili cultivars. Out of 12 primers four (OPA11, OPB03, OPB04 and OPB17) showed amplification of genomic DNA and generated 21 distinct score able bands of which 17 (80.95%) were polymorphic. The highest percentage (85.71) polymorphic locus was found in OPB03 while the lowest (66.67) in OPA11. The highest genetic distance was computed between Jamalpur Balujuri and Matal marich with the lowest genetic identity as against the lowest genetic distance between Hajari marich and Balujuri marich. The UPGMA dendogram indicated segregation of ten chili varieties and genotypes into two main clusters. Variety Bogra marich and Matal marich formed cluster 1 and Balujuri marich, Deshi marich, Jamalpuri balujuri, Bindu marich, Syloti, Hajari, Biroli city, and the genotype Ausadhebrara grouped in cluster 2. The result indicates the genetic diversity among the chili cultivars and RAPD marker could be used for improvement of chili varieties.

Keywords: Genetic diversity, Chili cultivars, RAPD markers

Location: Mymensingh

Start Date:

End Date:

Program Area: Conservation and Biodiversity

Commodity:

Objectives:

The objective of the study was to assess genetic diversity between chili cultivars using RAPD marker that could facilitate the chili breeders to develop new varieties for higher yield and also for tolerant to stresses.

Methodology:

Ten chili cultivars were used for the molecular characterization through RAPD technique. The experiment was carried out in the Biotechnology Laboratory of the Bangladesh Institute of Nuclear Agriculture, Mymensingh, Bangladesh. Fresh leaf samples collected from 21-day-old seedlings were used for DNA extraction. The harvested leaf samples were kept in –80^oC freezer until extraction. Modified CTAB mini-prep method was followed to extract DNA from leaf samples. The quality of the DNA was verified by electrophoresis on a 0.8% agarose gel in TBE (Tris-boric acid-EDTA) buffer. The concentration of the DNA samples was determined using a UV spectrophotometer at 260 nm. RAPD reactions were maintained following the process of Williams et al. (1990) with some modifications. The screening of primer was done with 12 arbitrary decamer primers (Bengalore Genei, India). Four primers producing a good score apple and reproducible bands were selected for subsequent RAPD analysis of chili cultivars (Table 1). PCRs were performed on each DNA sample in a 10.0 μl reaction mixture containing 1× PCR buffer (10 mM Tris HCl, pH 8.5; 50 mM KCl and 15 mM MgCl2), 10 mM each dNTPs (Bengalore Genei, India), 5 pmols primer, 2 U of Taq DNA polymerase (Bengalore Genei), 100 ng of genomic DNA. The reaction with each primer was replicated thrice to check the reproducibility of DNA. DNA amplification was carried out in a DNA thermocycler (Biometra, Germany) at the following thermal profile: initial denaturation for 3 min at 94°C followed by 41 cycles of 1 min denaturation at 94°C, 1 min annealing at 35°C and extension at 72°C for 2 min. A final extension step at 72°C for 10 min was allowed for complete extension of all amplified fragments. Amplified fragments were separated on a 1.5% agarose (Invitrogen, Canada) gel in 1X TBE buffer along with 20 bp DNA weight marker (Bengalore Genei, India) for 2 hours at 100V. Gel was stained with ethidium bromide solution (0.1 μg/ml) for 20 min. Finally fragments were visualized under UV-transilluminator and photographed by Gel Documentation System (Biometra, Germany). The amplified bands were visually scored as present (1) and absent (0) separately for each individual and each primer. The scores were pooled to create a single data matrix. This was used to estimate polymorphic loci, genetic diversity, genetic distance and a UPGMA dendrogram using computer program, POPGENE (Version 1.31

Source of Information: Plant Tissue Cult. & Biotech. 22(2): 127-136, 2012 (December)

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

The polymorphism detected among the accessions can be used in breeding programme to maximize the use of genetic resources as well as improve chili varieties. From this investigation, it was revealed that the highest genetic identity (0.9524) remains between Balujuri marich and Hajari marich. On the other hand, the lowest (0.4728) genetic identity was observed between the Balujur marich and Matal marich. These findings indicated that Balujuri marich vs Hajari and Matal marich vs Balujuri marich could be used in plant breeding program for development of new chili varieties. RAPD markers provide a fast and efficient tool for genetic variability assessment and are currently used in plant genetic resources management. It is also evident from the dendrogram that the genotypes Jamalpuri balujuri and Matal marich were most distantly related to each other and hence it is recommended that these two genotypes should be used in a hybridization program to create maximum genetic diversity for the improvement of Capsicum crop in Bangladesh.

Knowledge Management: Journal