Molecular Differentiation between Two Varieties of Rosmarinus officinalis Grown in North East Region of Iraq

Abstract

In an attempt to differentiate and study one of the most popular medicinal plants (Rosmarinus officinalis L.) grown in the North East region of Iraq, a number of experiments were carried out to differentiate between subspecies available in Kurdistan Province at the morphological and molecular levels. A survey and morphological study was conducted on these subspecies of rosemary in Erbil and Sulaimania governorates. It was found that there are morphological differences in leaves, flowers, shoot growth directions, number of branches, leaf size and shape, accordingly two subspecies were identified and marked as class A and class B of R. officinalis in North East of Iraq. Subspecies A was characterized by a straight shoot growth, dark green leaves and whitish blue flowers while subspecies B has random growth, green leaves and white flowers. In order to investigate the genetic variation, the DNA was isolated and purified from the two subspecies, and then electrophorized using agarose gel. Chromosomal DNA appeared as a clear smear on gel for both A and B. Six types of restriction enzymes were used to digest the genomic DNA for both subspecies A and B, namely EcoRI, EcoRV, BamHI, HindIII, SalI and SmaI. Resulted genomic DNA was amplified using PCR technique for ribosomal DNA (rDNA) as the target sequence, of 668 bp which includes 18S, 5.8S and 26S RNA genes separated by two ITS regions that represent the non-coding sequence of interest for subspecies identification. It is concluded that there are obvious genomic differences in the rDNA in both subspecies, according to the recognition sites availability of restriction enzymes in template DNA, as there was cleavage sites for each one of EcoRV, BamHI, HindIII and SalI in rDNA sequence of subspecies A, while no one of the six restriction enzymes cleavage sites used was present in subspecies B.