Genetic Diversity in Punica granatum Revealed by Nuclear rRNA, Internal Transcribed Spacer and RAPD Polymorphism

Abstract

The morpho-physiological and molecular markers were used to reveal genetic diversity among 13 putative varieties of Punica granatum grown in India. A hybrid protocol of DNA isolation was developed to obtain high molecular weight quality genomic DNA. Ten RAPD primers generated 119 marker bands amounting to 92.44 % polymorphism with 88 % polymorphic information content. The 5.8S rRNA gene region was found to be highly conserved (99.34 %) followed by ITS-1 (96.58 %) and ITS-2 (89.21 %) intron regions encompassing the gene. ITS-2 region was recorded with the highest percentage of parsimony informative sites with high divergence mainly due to SNPs. The PCA accounted for 84.21 % of total variance. The AMOVA of RAPD data revealed the maximum genetic variation within population rather than among populations. Nei’s gene diversity (h) was 0.2684 and Shannon information index (i) was 0.4135. The Nei’s genetic distance ranged from 0.0620 to 0.0878. The coefficient of gene differentiation between populations (Gst) of 0.2584 and gene flow (Nm) of 1.4347 validates that the main proportion of genetic variation was within populations. The present study validates the utility of ITS rDNA region as a reliable indicator of phylogenetic interrelationships, especially ITS-2 as probable DNA barcode at higher levels and can serve as an additional approach for genetic cataloguing of pomegranate germplasm for crop improvement.