Internal transcribed spacer guided multiplex PCR for species identification of Convolvulus prostratus and Evolvulus alsinoides

Abstract

Shankhpushpi is a reputed drug from an Indian system of medicine for treating mental disorders and enhancing memory. Two herbs, namely Convolvulus prostratus Forssk. and Evolvulus alsinoides (L.) L., are commonly known as Shankhpushpi. Ambiguous vernacular identity can affect the scientific validity of the Shankpushpi-based herbal drug therapy. In the present investigation, a novel and sensitive multiplex PCR method based on polymorphism in the internal transcribed spacer (ITS) region was developed to establish the molecular identity of C. prostratus and E. alsinoides. DNA was isolated and the ITS region was amplified, sequenced and assembled. Sequences were aligned to identify variable nucleotides in order to develop plant-specific primers. Primers were validated in singleplex reactions and eventually a multiplex assay was developed. This assay was tested for sensitivity and validated by amplifying DNA isolated from the simulated blended powdered plant material. Primers developed for C. prostratus resulted into a 200 bp amplicon and 596 bp for E. alsinoides. The assay was found to be sensitive enough for amplification of low quantities of DNA. The method can detect 10% of the mixing of plants with each other in blended material. This PCR assay can be used for rapid botanical identification of Shankhpushpi plant materials and will improve evidence-based herbal drug therapy.

Plastome-wide comparison reveals new SNV resources for the authentication of Dendrobium huoshanense and its corresponding medicinal slice (Huoshan Fengdou)

“…Furthermore, after intensive processing, the medicinal slices of Dendrobium species become more difficult to distinguish 44 . Recently, well-developed molecular marker techniques, i.e ., SSR, ISSR, AFLP and DNA barcodes became available for the identification of medicinal plants 9 , 10 , 11 , 45 , 46 , 47 , 48 , 49 . However, these methods can only be used to distinguish fresh Dendrobium materials, but are useless for their medicinal slices due to two reasons.…”

Comprehensive analysis using DNA metabarcoding, PCR, and HPLC unveils the adulteration in Brahmi herbal products

“…The emergence and development of DNA-based technology has enabled the cost-effective, rapid, and yet sensitive and species-specific PCR assays to authenticate medicinal plants. Species-specific sequence-characterized amplified region (SCAR) markers, nuclear ribosomal internal transcribed spacer (nrITS) sequences, and different plastid markers (e.g., rbcL, trnH-psbA, and matK) have been extensively used to authenticate plant species(de Boer et al, 2015; Mishra et al, 2016b; Sharma and Shrivastava, 2016; Xin et al, 2018). Earlier studies have revealed a PCR-based assay for the authentication of herbal products.…”

“…There is a lack of centralized authentication systems that provide taxon identification with high-resolution power. The predominance of species-specific PCR assays (Noh et al, 2021; Sharma and Shrivastava, 2016) DNA barcoding(Bansal et al, 2018; Seethapathy et al, 2015; Thakur et al, 2019; Vassou et al, 2016), and metabarcoding (Raclariu et al, 2017; Raclariu et al, 2017; Raclariu et al, 2018b; Seethapathy et al, 2019) based molecular authentication of medicinal plants has recently undergone a revolution, mostly because of DNA’s accessibility, consistency, and independence from tissue characteristics, age, harvesting techniques, and storage conditions. Chinese Pharmacopoeia, United States Pharmacopeia, British Pharmacopoeia, Japanese Pharmacopoeia, and Hong Kong Chinese Materia Medica are advocating DNA-based authentication around the globe (Wu and Shaw, 2022).…”

Recent advances onDNAand omics‐based technology in Food testing and authentication: A review

“…(CP) syn and Evolvulus alsinoides of ayurvedic medicine Shankhpushpi (Sharma & Shrivastava, 2016a). Similarly, the identity of another ayurvedic medicine Terminalia arjuna is determined through ITS regions, which are also used in the detection of adulteration from other three Terminalia species (Sharma & Shrivastava, 2016b).…”