Micropropagtion and DNA-barcoding of the endangered endemic Phlomis aurea plant of Saint Katherine

Abstract

Saint Katherine is considered a “biodiversity hotspot” because of the high level of endemism of reported plant species. In this study, conservation of the endangered endemic plant; Phlomis aurea of Saint Katherine, Southern Sinai, Egypt, was carried out through micropropagation and DNA barcoding. The first efficient micropropagation protocol for Phlomis aurea was established as a mean of ex situ conservation of the plant. Shoot tips and nodal segments of in vitro germinated seedlings were established on Murashige and Skoog medium supplemented with 0.54 μM β-naphthalene acetic acid (NAA) and 2.46 μM N6-(2-isopentenyl) adenine (2iP) in combination with 6-benzylaminopurine (BA) or kinetin (Kin). The medium supplemented with 3.48 μM Kin considered optimum for both explants. For multiplication, BA was the most efficient cytokinin. The percentage of rooted explants reached 100% at the concentration of 14.7 μM indolebutyric acid (IBA), whereas the highest number of roots was recorded for 4.90 μM, which considered the optimum concentration with a percentage of 80% of rooting. Rooted plantlets were transplanted in the greenhouse with 75% survival rate. The present study also aimed to carry out DNA barcoding of Phlomis aurea for accurate identification to provide a database for establishing an efficient conservation program for the plant. Three chloroplast DNA markers were used [ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (rbcL), maturase K (matK) and RNA polymerase C1 (rpoC1)] and all were successful in amplifying target regions, however the performance of both rbcL and matK markers seemed to be species‐specific. The similarity percentage was maximum for rbcL (99.81%) and matK (100%) compared to the database of the same species.