Genetic variation among different accessions of Onobrychis viciifolia Scop. (Fabaceae) revealed by genomic ISSR fingerprinting.

Abstract

The genus Onobrychis Miller with about 130 species belongs to the family Fabaceae and tribe Hedysareae. The most widespread species of this genus is Onobrychis viciifolia SCOP. O. viciifolia is of significant agricultural use as a perennial forage and fodder legume. In this study 16 specimens from 8 representative Iranian populations of Onobrychis viciifolia SCOP. were collected from their natural habitats. ISSR genetic fingerprinting was used to analyze the genetic variation because ISSR analysis has become a powerful tool to detect similarities between and within species. Total genomic DNA was isolated from dried leaves using the standard CTAB extraction protocol and purified. Among different primer, (GACA)4 primer providing the most reliable and consistent results was chosen. PCR amplifications were performed. DNA fragments were separated by vertical polyacrylamide gel electrophoresis for 3 h at 65 W. After drying, the denaturing gels were exposed for 24 h to X-ray film. Each ISSR band was considered as a character and the presence or absence of the band was scored in binary code. A data matrix was assembled and analyzed using PAUP. The genetic relationships among the populations were analyzed using Neighbor-joining based on distance measure of total character difference. ISSR PCR generated a total of 56 distinguishable bands, of which 44 (78.5%) were parsimony-informative characters, 8 characters being constant, and 4 variable characters were parsimony-uninformative. The size of the bands ranged from 247 to 1700 bp. A dendrogram was generated using NJ showing three main groups which were consistent morphologically. ISSR profiles were shown to be of high value to reveal genotypic diversity and, in the longer term, to provide potential molecular markers that can be linked to phenotypic properties.