RbcLa marker based identification and phylogenetic analysis of Kasuri methi (Trigonella foenum-graecum L.): a native plant of Kasur district of Punjab (Pakistan)

Trigonella foenum-graecum (L.) is one of the potent medicinal plants and is a well-known spice for its specific aroma. It is a native plant of Kasur District of Punjab Pakistan commonly known as Kasuri methi. Kasur district is a semi-arid subtropical area, famous for its production worldwide. The exact plant identification up to the species level is hard for plant taxonomists and herbalists, due to its resemblance with its close relatives. DNA barcoding is the most modern approach for plant identification based upon a highly conserved chloroplast genome. Rbcla marker is considered one of the most conserved regions used for highly complex and confusing species-level identification. The current study is the first effort for molecular identification of Trigonella foenum-graecum (L.) based upon DNA barcoding in South Asia. Chloroplast DNA was isolated by applying the modified CTAB method. Specific primers for rbcLa were used for polymerase chain reaction (PCR) amplification. Purified PCR products were then subjected to DNA sequence for rbcla DNA barcode. After DNA sequencing, the chromatogram file was analyzed by using Chromas. Then align sequences by multiple sequence alignment by using clustalW in Geneious. Afterward, BLAST the sequence from the NCBI database and align this query sequence. The rbcLa barcode showed 99.80% precision for species identification. The cataloging derivative from the DNA barcode tree was then compared with archaic morphology-based taxonomy. This will help identification of Kasuri methi from its closely related species for better business opportunities and safer usage by the public as spice and medicine.