ABSTRACT
Objective To assess the feasibility of the rbcL sequence of chloroplast DNA as a genetic marker to identify Cannabis sativa L. Methods The rbcL sequences in 62 Cannabis sativa L. samples, 10 Humulus lupulus samples and 10 Humulus scandens DNA samples were detected, and 96 rbcL sequences of the Cannabaceae family were downloaded from Genbank. Sequence alignment was performed by MEGA X software, the intraspecific and interspecific Kimura-2-Parameter （K2P） genetic distances were calculated, and the system clustering tree was constructed. Results The rbcL sequence length acquired by sequencing of Cannabis sativa L. and Humulus scandens were 617 bp and 649 bp, respectively, and two haplotypes of Cannabis sativa L. were observed in the samples. The BLAST similarity search results showed that the highest similarity between the sequences acquired by sequencing and Cannabis sativa L. rbcL sequences available from Genbank was 100%. The genetic distance analysis showed that the maximum intraspecific genetic distance （0.004 9） of Cannabis sativa L. was less than the minimum interspecific genetic distance （0.012 9）. The results of median-joining network and system clustering tree analysis showed that Cannabis sativa L. and other members of the Cannabaceae family were located in different branches. Conclusion The rbcL sequence could be used as a DNA barcode for identifying Cannabis sativa L., and combined with comparative analysis of the rbcL sequence and system cluster analysis could be a reliable and effective detection method for Cannabis sativa L. identification in forensic investigation.
Subject(s)
Cannabis/genetics , Genetic Markers , Sequence Analysis, DNAABSTRACT
Objective: To establish the fingerprint of Althaeae Roseae Flos by HPLC and the molecular identification method of DNA barcode of rbcL sequence. Methods: The fingerprint establishment of Althaeae Roseae Flos was performed on Welchrom Column C18 (300 mm × 4.6 mm, 5 μm) with acetonitrile - 0.1% formic acid solution as mobile phase for gradient elution, with flow rate of 1.0 mL/min, column temperature of 35 ℃, detection wavelength of 365 nm, injection volume of 10 μL. DNA barcode molecular identification method was used for PCR amplification and determination of rbcL sequence. Results: The fingerprints of 11 samples were established, 21 common peaks were obtained, their similarities were calculated, and four components (hyperoside, quercetin, apigenin and kaempferide) were determined. The total length of rbcL sequence of 11 samples was measured, and the G + C content was 44.10%-44.40% and genetic distance (K2P) was 0.001 4. There were 10 ectopic points and the similarity was 99.00%. Conclusion: The two methods are stable and reliable, which can provide basis for the identification and quality control of A. rosea.