Elevating Thyme Species Identification: Exploiting Key Chloroplast Genes (matK, rbcL, and psbA-trnH) through DNA Barcoding and Phylogenetic Analysis

Öz

Understanding genetic relationships and diversity among species is crucial for unraveling evolutionary processes, ecological interactions, and conservation strategies. DNA sequence analysis serves as a powerful tool in this endeavor. This study focuses on the Thymus genus, a collection of notable species, to investigate its genetic framework. Leveraging DNA sequences from key regions (matK, rbcL, and psbA-trnH), we aim to elucidate genetic connections within the Thymus genus and uncover mechanisms driving its diversity. The Thymus genus, with its diverse species and ecological characteristics, provides a captivating platform for genetic exploration. Through DNA sequence analysis, we aim to unveil genetic interconnections, biodiversity patterns, and the factors shaping the genus's evolution. Our findings are aligned with previous studies, and this consistency highlights the presence of polymorphism within potential sequences. Employing coding loci and spacer regions, our study contributes to Lamiaceae family barcoding research. Despite variations across gene regions, the concatenation of sequences enhances result reliability. We analyzed the suitability of matK, rbcL, and psbA sequences for Thymus identification, observing rbcL and psbA outperforming matK. Our novel approach, rooted in chloroplast DNA, presents a promising method for species discernment. By analyzing multiple chloroplast gene regions, this technique offers a fresh perspective on genetic affinity assessment using DNA barcodes. In conclusion, this study not only contributes to Thymus germplasm resource preservation but also exemplifies a novel approach to discerning Thymus species through DNA analysis. This methodology carries the potential for broader application, enriching our understanding of genetic relationships and diversity in the plant kingdom.

Anahtar Kelimeler

• Thymus
• DNA barcoding
• Choloroplast genes
• Phylogenetic analysis

Kaynakça

1. Abdulrahman SS, Daştan SD, Shahbaz SE, Selamoglu Z. 2023. Phylogenetic analysis of Prunus genus using nuclear and chloroplast gene markers as a bioorganic structure profiling. J Mol Struct, 1284: 135300. Doı: 10.1016/j.molstruc.2023.135300 .
2. Alissandrakis E, Tarantilis PA, Harizanis PC, Polissiou M. 2007. Comparison of the volatile composition in thyme honeys from several origins in Greece. J Agri Food Chem, 55(20): 8152-8157. DOI: 10.1021/jf071442y.
3. Amiteye S. 2021. Basic concepts and methodologies of DNA marker systems in plant molecular breeding. Heliyon, 7(10): 202. DOI: 10.1016/j.heliyon.2021.e08093.
4. Aneva I, Zhelev P, Bonchev G, Boycheva I, Simeonova S, Kancheva D. 2022. DNA Barcoding Study of Representative Thymus Species in Bulgaria. Plants, 11(3): 270. DOI: 10.3390/plants11030270.
5. Ansarifar E, Moradinezhad F. 2022. Encapsulation of thyme essential oil using electrospun zein fiber for strawberry preservation, Chem Biol Techn Agri, 9: 2. DOI: 10.1186/s40538-021-00267-y.
6. Chen S, Yin X, Han J, Sun W, Yao H, Song J, Li X. 2023. DNA barcoding in herbal medicine: retrospective and prospective. J Pharm Anal, 13(5): 431–441. DOI: 10. 1016/j.jpha.2023.03.008.
7. Chinnkar M, Jadhav P. 2023. Evaluating DNA barcoding using five loci (matK, ITS, trnH- psbA, rpoB, and rbcL) for species identification and phylogenetic analysis of Capsicum frutescens. J Appl Biol Biotech, 11(3): 97103. DOI: 10.7324/JABB.2023.57526.
8. Coissac E, Hollingsworth PM, Lavergne S. 2016. From barcodes to genomes: extending the concept of DNA barcoding. Mol Ecol, 25: 423-1428. DOI: 10.1111/mec.13549.

9. Cutillas AB, Carrasco A, Gutierrez RM, Tomas V, Tudela J. 2018. Thymus mastichina L. essential oils from Murcia (Spain): Composition and antioxidant, antienzymatic and antimicrobial bioactivities. PLoS ONE, 13(1): e0190790. DOI: 10.1371/journal.pone.0190790.
10. Debrah I, Ochwedo KO, Otambo WO, Machani MG, Mago- mere EO, Onyango SA, Zhong D, Amoah LE, Githeko AK, Afrane YA, Yan G. 2023. Genetic diversity and population structure of anopheles funestus in Western Kenya based on mitochondrial DNA marker COII. InSects, 14: 273. DOI: 10.3390/insects14030273.
11. El Ouariachi EM, Paolini J, Bouyanzer A, Tomi P, Hammouti B, Salghi R, Majidi L, Costa J. 2011. Chemical composition and antioxidant activity of essential oils and solvent extracts of Thymus capitatus (L.) Hoffmanns and link from Morocco. J Medic Plants Res, 5: 5773-5778.
12. Furan MA. 2023. Species identification and germplasm conservation of origanum based on chloroplast genes. Genet Resour Crop Evol, DOI: 10.1007/s10722-023-01679-5.
13. Hammoudi Halat D, Krayem M, Khaled S, Younes S. 2022. A Focused Insight into Thyme: Biological, Chemical, and Therapeutic Properties of an Indigenous Mediterranean Herb Nutrients, 14(10): 2104. DOI: 10.3390/nu14102104.
14. Hollingworth PM, Forrest LL, Little DP. 2009 A DNA barcode for land plants. PNAS, 106: 12794-12797. DOI: 10.1073/pnas.0905845106.
15. Honorato AC, Nohara GA, Assis RMA, Maciel JFA, Carvalho AA, Pinto JEBP, Bertolucci SKV. 2023. Colored shade nets and different harvest times alter the growth, antioxidant status, and quantitative attributes of glandular trichomes and essential oil of Thymus vulgaris L. J Appl Res Medic Arom Plants, 35: 100474. DOI: 10.1016/j.jarmap.2023.100474.
16. Hua Z, Jiang C, Song S, Tian D, Chen Z, Jin Y, Zhao Y, Zhou J, Zhang Z, Huang L, Yuan Y. 2023. Accurate identifica- tion of taxon-specific molecular markers in plants based on DNA signature sequence. Mol Ecol Resour, 23: 106-117. DOI: 10.1111/1755-0998.13697.
17. Jiang Y, Zhu C, Wang S, Wang F, Sun Z. 2023. Identification of three cultivated varieties of Scutellaria baicalensis using the complete chloroplast genome as a super-barcode. Sci Rep, 13: 5602. DOI: 10.1038/s41598-023-32493-9.
18. Johnson MS, Venkataram S, Kryazhimskiy S. 2023. Best practices in designing, sequencing, and identifying random DNA barcodes. J Mol Evol, 91: 283-280. DOI: 10.1007/s00239-022-10083-z.
19. Joshi K, Chavan P, Warude D, Patwardhan B. 2004. Molecular markers in herbal drug technology. Current Sci, 159-165.
20. Kim M, Sowndhararajan K, Kim S. 2022. The chemical composition and biological activities of essential oil from Korean native thyme Bak-Ri-Hyang (Thymus quinquecostatus Celak.). Molecules, 27(13): 4251. DOI: 10.3390/molecules27134251.
21. Kryvtsova M, Hrytsyna M, Salamon I, Skybitska M, Novykevuch O. 2022. Chemotypes of Species of the Genus Thymus L. in Carpathians region of Ukraine their essential oil qualitative and quantitative characteristics and antimicrobial activity. Horticulturae, 8(12): 1218. DOI: 10.3390/horticulturae8121218.
22. Lawrence BM, Tucker AO. 2002 The genus Thymus as a source of commercial products. Taylor & Francis Group, Hamburg Germany, pp: 11.
23. Li H, Xiao W, Tong T, Li Y, Zhang M, Lin X, Zou X, Wu Q, Guo X. 2021. The specific DNA barcodes based on chloroplast genes for species identification of Orchidaceae plants. Sci Rep, 11: 1424. DOI: 10.1038/s41598-021-81087-w.
24. Michel CI, Meyer RS, Taveras Y, Molina J. 2016. The nuclear internal transcribed spacer (ITS2) as a practical plant DNA barcode for herbal medicines. J Appl Res Med Aromat Plants, 3: 94–100. DOI: 10.1016/j.jarmap. 2016.02.002.
25. Mohammadi A, Mahjoub S, Ghafarzadegan K, Nouri HR. 2018. Immunomodulatory effects of Thymol through modulation of redox status and trace element content in experimental model of asthma. Biomed Pharmacother, 105: 856-861, DOI: 10.1016/j.biopha.2018.05.154.
26. Morshdy AEMA, El-tahlawy AS, Qari SH, Qumsani AT, Bay DH, Sami R, Althubaiti EH, Mansour AMA, Aljahani AH, Hafez AE-SE, Mohmoud AFA, Bayomi RME, Hussein MA. 2022. Anti-biofilms’ activity of garlic and thyme essential oils against Salmonella typhimurium. Molecules, 27(7): 2182. DOI: 10.3390/molecules27072182.
27. Mosavat N, Golkar P, Yousefifard M, Javed R. 2019. Modulation of callus growth and secondary metabolites in different Thymus species and Zataria multiflora micropropagated under ZnO nanoparticles stress, Biotechnol Appl Biochem, 66: 316-322. DOI: 10.1002/bab.1727.
28. Nabissi M, Marinelli O, Morelli MB, Nicotra G, Iannarelli R, Amantini C, Santoni G, Maggi F. 2018. Thyme extract increases mucociliary-beating frequency in primary cell lines from chronic obstructive pulmonary disease patients, Biomedic Pharmacother, 105: 1248-1253. DOI: 10.1016/j.biopha.2018.06.004.
29. Nair KP. 2023. Thyme. In: a compendium of unique and rare spices. Springer Cham, pp: 85-92. DOI: 10.1007/978-3-031-20249-0_8.
30. Nongbet A, Chrungoo NK. 2023. Distribution mapping and diversity assessment of ilex venulosa from meghalaya using internal transcribed spacer regions, matK and rbcL. In: Uppaluri RVS, Rangan L (eds) Conservation of biodiversity in the North Eastern States of India. Springer, Singapore, pp: 83–87. DOI: 10.1007/978-981-99-0945-2_5.
31. Polaiah AC, Damor PR, Reddy RN, Manivel P, Shivakumara KT, Suthar MK, Thondaiman V, Manjesh GN, Bindu KH, Kumar J. 2023. Development of genomic SSR markers in Gymnema sylvestre (Retz.) R.Br. ex Sm. using next generation DNA sequencing and their application in genetic diversity analysis. J Appl Res Med Aromat Plants, 34: 100455. DOI: 10.1016/j.jarmap.2022.100455.
32. Radi FZ, Bouhrim M, Mechchate H, Al-Zahrani M, Qurtam AA, Aleissa AM, Drioiche A, Handaq N, Zair T. 2021. Phytochemical analysis, antimicrobial and antioxidant properties of Thymus zygis L. and Thymus willdenowii Boiss. essential oils. Plants, 11(1): 15. DOI: 10.3390/plants11010015.
33. Rahimi A, Mohammadi MM, Moghaddam SS, Heydarzadeh S, Gitari H. 2022. Effects of stress modifier biostimulants on vegetative growth, nutrients, and antioxidants contents of garden thyme (Thymus vulgaris L.) under water deficit conditions. J Plant Growth Regul, 41: 2059-2072. DOI: 10.1007/s00344-022-10604-6.
34. Raskoti BB, Ale R. 2021. DNA barcoding of medicinal orchids in Asia. Sci Rep, 11: 23651. DOI: 10. 1038/s41598-021-03025-0.
35. Rodríguez I, Tananaki C, Galán-Soldevilla H, Pérez-Cacho PR, Serrano S. 2021. Sensory profile of greek islands thyme honey. Appl Sci, 11(20): 9548. DOI: 10.3390/app11209548.
36. Salehi B, Mishra AP, Shukla I, Rad MS, Contreras MM, Carretero AS, Fathi H, Nasrabadi NN, Kobarfard F, Rad JS. 2018. Thymol, thyme, and other plant sources: Health and potential uses. Phytother Res, 32: 1688-1706. DOI: 10.1002/ptr.6109.
37. Sardar SK, Ghosal A, Haldar T, Maruf M, Das K, Saito- Nakano Y, Kobayashi S, Dutta S, Nozaki T, Ganguly S. 2023. Prevalence and molecular characterization of Enta- moeba moshkovskii in diarrheal patients from Eastern India. PLoS Negl Trop Dis, 17(5): e0011287. DOI: 10.1371/journ al.pntd.0011287.
38. Singh RB, Mahenderakar MD, Jugran AK, Singh RK, Srivas- tava RK. 2020. Assessing genetic diversity and population structure of sugarcane cultivars, progenitor species and genera using microsatellite (SSR) markers. Gene, 753: DOI: 10.1016/j.gene.2020.144800.
39. Taher MS, Salloom YF, Hussein Al-Asadi RAU, Al-Mousswi ZJ, Alamrani HA. 2021. The medicinal importance of Thyme plant (Thymus vulgaris). Biomedicine, 41(3): 531-534.
40. Tamura K, Stecher G, Kumar S. 2021. MEGA 11: molecular evolutionary genetics analysis version 11. Mol Biol Evol, 38(7): 3022-3027. DOI: 10.1093/molbev/msab120.
41. Techen N, Parveen I, Pan Z, Khan IA. 2014. DNA barcoding of medicinal plant material for identification. Current Opin Biotechnol, 25: 103–110. DOI: 10.1016/j. copbio.2013.09.010.
42. Thakur VV, Tripathi N, Tiwari S. 2021. DNA barcoding of some medicinally important plant species of lamiaceae family in India. Mol Biol Rep, 48: 3097-3106. DOI: 10.1007/s11033-021-06356-3.
43. Tomanić D, Božin B, Kladar N, Stanojević J, Čabarkapa I, Stilinović N, Apić J, Božić DD, Kovačević Z. 2022. Environmental bovine mastitis pathogens: Prevalence, antimicrobial susceptibility, and sensitivity to Thymus vulgaris L., Thymus serpyllum L., and Origanum vulgare L. essential oils. Antibiotics, 11(8): 1077. DOI: 10.3390/antibiotics11081077.
44. Trindade H. 2010 Molecular biology of aromatic plants and spices. A review. Flavour Fragr J, 25(5): 272-281. DOI: 10.1002/ffj.1974.
45. Uritu CM, Mihai CT, Stanciu GD, Dodi G, Stratulat TA, Luca A, Constantin MML, Stefanescu R, Bild V, Melnic S, Tamba BI. 2018. Review article medicinal plants of the family Lamiaceae in pain therapy. Pain Res Manag, 2018: 1-45 DOI: 10.1155/2018/7801543.
46. Wang Y, Dietrich CH, Zhang Y. 2017. Phylogeny and historical biogeography of leafhopper subfamily Evacanthinae (Hemiptera: Cicadellidae) based on morphological and molecular data. Sci Rep, 7: 45387. DOI: 10.1038/srep45387.
47. Yu J, Wu X, Liu C, Newmaster S, Ragupathy S, Kress WJ. 2021. Progress in the use of DNA barcodes in the identification and classification of medicinal plants. Ecotoxicol Environ Saf, 208: 111691. DOI: 10.1016/j.Ecoenv.2020.111691.