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Molecular Characterization of Jamir (citrus Jambhiri) Accessions of Bangladesh Through Pcr Based Rapd Markers

S. Akhter
Department of Horticulture, Bangladesh Agricultural University, Mymensingh

M.J. Ferdous
Department of Horticulture, Bangladesh Agricultural University, Mymensingh

M.R. Hossain
Department of Genetics and Plant Breeding, Bangladesh Agricultural University, Mymensingh

G. Rabbani
Department of Horticulture, Bangladesh Agricultural University, Mymensingh

Abstract/ Executive Summary:

Fourteen accessions of Jamir (Citrus jambhiri) were used to observe the variability among them by RAPD markers. Out of 16 screened decamer primers 6 primers were used which gave 80 clear and bright fragments. Of the 80 bands, 74 (92.5%) were polymorphic. Pair-wise comparison of DNA profile of the 14 Jamir showed inter-species similarity index (Sij) value 86.45% for CJ-14 vs CJ-6 pair which was higher than all other accession pairs and the lowest was in between CJ-9 vs CJ-5 (30.58 %). The estimate of Nei's (1972) genetic diversity (I) for entire accessions of Jamir was 0.1488 and Shannon's information index (h) was 0.1951. The highest genetic distance (D) value 0.7444 was observed between the accession CJ04 and CJ10 and between CJ10 and CJ12. On the other hand, the lowest distance value 0.1193 was found among the accession CJ15 which revealed that there is a great genetic difference among the accessions. The UPGMA dendrogram produced 2 main clusters and more than 7 sub cluster of the 14 Jamir accessions. Thus, RAPD offer a potentially simple, rapid and reliable method to evaluate the variability among all species of plant.

Keywords: Citrus jambhiri, PCR, RAPD

Location: Horticulture Field Laboratory, Bangladesh Agricultural University, Mymensingh

Start Date: 00-01-2008

End Date: 00-09-2008

Program Area: Variety and Species

Commodity: Orange

Objectives:

1. To characterize the local Jamir accession through RAPD markers and to assess the genetic diversity and relatedness of Jamir accession for future breeding program.

Methodology:

The research work was conducted at the Field Laboratory, A total of 14 jamir accessions were investigated in this study. These accessions were maintained in the farm of “Collection, Molecular Characterization and Conservation of Landraces and Wild Relatives of Different Vegetables and Fruits of Bangladesh” project, at Horticulture Field Laboratory, Bangladesh Agricultural University, Mymensingh. DNA was extracted from 14 jamir accessions separately following standard protocol using SDS and Phenol: Chloroform: IAA followed by alcohol precipitation. DNA was confirmed using 1% agorose gel and quantified at 260nm spectrophotometrically. Out of 16 primers screened, 6 primers viz. OPA-05, OPA-07, OPA-10, OPC-05, OPK-07 and OPK-12 exhibiting good quality banding patterns and sufficient variability were selected by primer test for RAPD analysis. Polymerase Chain Reactions (PCR) were done in a volume of 10 µl containing 10 x PCR Buffer, 0.25 mM each of the dNTPs, 1µM of each of primer, 1 unit Taq DNA polymerase, 75ng template DNA and a suitable amount of sterile deionized water. Amplification was carried out in an oil free thermal cycler with the following thermal profile: initial denaturation step at 94°C for 3 min followed by 40 cycles at 94 °C for 1 min., 36°C for 41 min, and 72 °C for 2 min with 2 replications. PCR was confirmed by electrophoresis. All distinct bands (RAPD markers) were given identification numbers according to their position on gel and scored visually on the basis of their presence (1) or absence (0). The software DNA FRAG version 3.03 was used to estimate allelic length. This was used to estimate polymorphic loci, Nei's (1972) genetic distance (D), gene frequencies, gene diversity, Shanon information index (I) and to construct a Unweighted Pair Group Method of Arithmetic Means (UPGMA) dendrogram among populations using a computer program, POPGENE version 1.31. The similarity index values (SI) between the RAPD profiles of any two individuals were calculated from RAPD markers according to the following formula: SI = 2 Nxy / Nx+ Ny, where Nxy is the total number of RAPD bands shared by individuals x and y, Nx and Ny are the number of bands in individual x and y, respectively (Lynch, 1990). Within population similarity (Sx) was calculated as the population of SI across all possible comparisons between individuals within a population.

Source of Information: J. Agrofor. Environ. 3 (1): 21-24, 2009; ISSN 1995-6983

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

A wide variability was present among the collected accessions of Jamir. And the selection of superior genotypes from the collected accessions will be helpful to increase production through breeding program which in turn could improve varietal characteristics. The RAPD profiles of collected Jamir accessions of this study would be useful in monitoring genetic stability.

Knowledge Management: Journal