Phylogenetic analysis and DNA barcoding of Luisia tristis (Orchidaceae): Implications for species identification and biodiversity preservation

Abstract

Background: Understanding and preserving biodiversity requires effective methods for plant identification. The study aimed to analyze the genetic relatedness of Luisia tristis, a species of the Orchidaceae family, and assess the potential of DNA barcoding as a reliable identification technique. Methods: In this study, DNA was isolated from leaf samples, amplified, and sequenced for the chloroplast gene matK (maturase K). The sequence was uploaded to the NCBI database and subjected to multiple sequence alignment and BLAST analysis to assess its similarity to other close taxa. Using the Maximum Likelihood (ML) and Neighbor-Joining (NJ) methods, their phylogenetic relationships were investigated using MEGA X version 7.0. To verify the statistical robustness, 1000 bootstrap repetitions were used. Results: The results were further validated by constructing a phylogenetic tree utilizing sequence data from other orchid species. With sequence similarity values of nearly 98.5%, the analysis revealed that the species clustered into two distinct clades, indicating a close genetic relationship. Based on matK divergence patterns, the derived phylogenetic trees indicate that the taxa most likely have a monophyletic origin. A matK-specific QR code was generated using Bio-Rad’s barcode generator, providing a convenient method for encoding and retrieving genetic information. This study shows the effectiveness of DNA barcoding as a powerful tool for identifying species and preserving orchid taxa that face serious threats. Additionally, the molecular function of the matK protein from L. tristis was clarified by utilizing EMBOSS software to predict its 3D structure models. Conclusion: The result of this study was essential for the easy identification and protection of such orchid species for the future.