Development of DNA barcode for rapid identification of Epimedium elatum (Morren & Decne) from Northwestern Himalayas in India

Abstract

Epimedium elatum (Morren & Decne) is a rare monotypic high altitude medicinal species, endemic to shady forests in Northwestern Himalayas, India. Its recent chemoprofiling has revealed high concentration of Epimedin B, C & Icariin, thus matching quality standards of Herba Epimedii. DNA barcode was hitherto unreported in E.elatum. Three plastid genes (matK, rbcL & trnH-psbA) and one nuclear region gene (ITS) were partially sequenced to develop unique barcode and also investigate nucleotide diversity and phylogenetic relationship among five geographically diverse accessions of E.elatum. We used the Kimura 2-parameter (K2P) evolutionary model to calculate basic sequence statistics like conserved, variable, parsimony informative and singleton sites, including pairwise genetic distance in MEGA X software. Besides, we employed a maximum likelihood phylogenetic tree method for estimating the evolutionary divergences. The present study revealed correct blast validation of E.elatum populations with other Epimedium species. A total of 18 new sequences were obtained and submitted to NCBI Genbank via BankIT submission tool (MH615782–MH615799). Highest variable (polymorphic) sites were found in matK (269) followed by ITS (115); rbcL (18) and least in trnH-psbA (16). The evolutionary sequence divergence varied from 0.01 in rbcL/trnH-psbA; 0.09 in ITS to 0.19 in matK. Haplotype and single nucleotide polymorphism profile was rich in matK &ITS sequencing. Both primers developed a reliable and highly efficient way of determining nucleotide sequence diversity in E.elatum. ITS & matK met the ideal DNA barcode characteristics laid by Consortium of Barcode of Life and hence can be endorsed as ideal barcode markers for the identification of E.elatum in NW Himalayas, India.