engleski

Analysis of chloroplast DNA variability among some Hypericum species in Croatia

Background and Purpose Hypericum perforatum L. is considered as very variable species divided into three subspecies based on morphological, ecological and citogenetical characteristics. With the aim to test suitability of the cpDNA analyses for the intra and interspecific phylogenetic study of genus Hypericum, preliminary investigations of the cpDNA polymorphism were performed in H. perforatum subsp. typicum Beck, H. perforatum subsp. veronense (Schrank) Beck, H. perforatum subsp. angustifolium DC., H. maculatum Crantz, H. montanum L. and H. androsaemum L. comparing restriction patterns of the cpDNA fragments amplified by PCR with three pairs of universal chloroplast primers. Materials and Methods Plant material was collected from different location in Croatia. Total cellular DNA was extracted from fresh or frozen leaves. PCR amplification have been performed using three pairs of universal chloroplast primers (AS, CD and DT). Amplification products were digested using 4-base recognition endonucleases HinfI and TaqI and analyzed by agarose gel electrophoresis. The cluster analysis using WPGMA method has been performed on obtained data. Results Amplified products of AS and CD pairs of primers, 2.7 kb and 2.5 kb respectively, are the same in all samples. Obtained patterns of PCR products of AS primers pair after digestion with Hinf I are identical for three subspecies of H. perforatum L. as well as for H. maculatum Crantz, while profiles of species H. montanum L. and H. androsaemum L. show some differences. The patterns of PCR products of CD primers pair after digestion with Taq I are the same for three subspecies of H. perforatum L. but different for other investigated species. The cluster analysis demonstrates high species specifity of the cpDNA structure. Conclusion Considerable degree of interspecific polymorphism was detected in both restriction profiles, while intraspecific pattern was identical even between H. perforatum subspecies. This described method of cpDNA analysis appears applicable in interspecific phylogenetical investigation for relationship linkage study, i.e. between Hypericum perforatum L. and Hypericum maculatum Crantz, as well as intraspecific biogeographic investigation in genus Hypericum.

Hypericum spp; chloroplast DNA; universal primers; phylogenetic study

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