DNA barcode based species-specific marker for Ocimum tenuiflorum and its applicability in quantification of adulteration in herbal formulations using qPCR

Ocimum tenuiflorum L. is a highly traded and extensively used species in Ayurvedic medicine owing to its immense therapeutic potential, which has led to adulteration of both formulations as well as traded raw drugs. To meet the standard international market requirements, quick and efficient techniques for authentication are required which can keep pace with the increasing demand for the herb. In view of this, a robust and accurate DNA marker for the qualitative and quantitative analyses of this species and derived products is reported. The species-specific markers (designated as CIM-PATH-fwd / CIM-PATH-rev) developed in this study was designed based on the differences in the nucleotide sequence of the psbA-trnH intergenic region. A consistent amplicon of 218 bp was generated across all the O. tenuiflorum accessions and was absent in other Ocimum species studied. Further, developed species-specific primers were assessed for simultaneous authentication as well as quantification of O. tenuiflorum in commercially purchased herbal formulations by SYBR green-based quantitative real-time PCR assay. The established qPCR method was determined to be effective for the authentication and relative quantitative assessments of the products. The species-specific marker developed can be successfully employed as a taxonomic marker for O. tenuiflorum.