Abstract

The objective of the present study was to develop a Real Time-Sequence Characterized Amplified Region (RT-SCAR) of Catharanthus roseus, an effective molecular marker for authentication and quantification. Here, we made a comparative study of SCAR and RT-SCAR marker. These newer RT-SCAR methods are useful for the correct identification and quantification of C. roseus. In particular, this technique is very attractive and can be used as a novel tool for quantification of C. roseus when the SCAR results cannot be quantified and the RAPD results are not consistent. RAPD analysis of collected samples from different geographical locations in India was carried out with 25 random primers. The RAPD polymorphic band was cloned, sequenced and from the sequence information, primers pair for SCAR was developed. The same primers was used for RT-SCAR marker by using SYBR green fluorescence-based detection in Real Time PCR. Utilizing this newly specific RT-SCAR marker, we detected the presence and quantity of C. roseus in all the analyzed several samples, by aiding 10% its substitute and adulterants at time of DNA isolation. No difference was found in pure sample and 10% adulterant sample with the help of SCAR but in the case of RT-SCAR difference was found in the DNA sample. This analytical strategy could be used further for large scale quantification of C. roseus’s market samples.

Key words: Catharanthus roseus, RAPD, Sequence-Characterized-Amplified-Region (SCAR) Marker, Real Time Sequence-Characterized-Amplified-Region (RT-SCAR) Marker, Real Time PCR.