Določevanje raznolikosti rodu Viola z analizo ribosomalne DNA in merjenjem velikosti genoma

Jamnik, Polona (author), Rudolf Pilih, Katarina (mentor)

Rod Viola, ki spada v družino Violaceae, vključuje več kot petsto vrst, razširjenih po svetu, od zmernega pasu severne poloble do tropov in subtropov. Ta rod, prvič opisan v 18. stoletju, vključuje tako enoletne kot trajne zelnate rastline. V nalogi smo bili osredotočeni na morfološke in genetske značilnosti vrst Viola sp., z namenom, da bi bolje razumeli njihovo biotsko raznovrstnost in pripomogli k njihovemu ohranjanju. Morfološke značilnosti, kot so oblika in velikost listov, cvetov in stebel, ter barva cvetov, so ključne za razlikovanje med vrstami. Pri zbranih petdesetih vzorcih različnih vrst smo opravili meritve velikosti genoma s pretočno citometrijo, ki so pokazale, da se velikost genoma med vrstami razlikuje. Najmanjšo povprečno izmerjeno vrednost 2C je imela Viola reichenbachiana Jord ex. Boreau, medtem ko je največjo povprečno vrednost 2C dosegla Viola x wittrockiana. Za analizo ribosomalne DNA je ena glavnih težav prisotnost sekundarnih metabolitov, ki otežujejo izolacijo DNA in nadaljnje analize. Ugotovili smo, da CTAB metoda ni učinkovita pri vseh vrstah, zato so potrebne prilagoditve ali uporaba alternativnih metod. Uspešne PCR reakcije so omogočile določitev nukleotidnih zaporedij ribosomalne DNA 26 vzorcev različnih vrst, ki smo jih primerjali z zbirkami NCBI BLAST. Sekvenciranje ribosomalne DNA je pokazalo, da čeprav je odstotek genetske identitete med vrstami relativno visok, obstajajo značilne variacije, ki omogočajo diferenciacijo med posameznimi vrstami.

• Language: Slovenian
• Keywords: Viola, velikost genoma, vrednost 2C, ribosomalna DNA, sekvenciranje
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: BF - Biotechnical Faculty
• Year: 2024
• PID: 20.500.12556/RUL-162839
• COBISS.SI-ID: 210409987
• Publication date in RUL: 28.09.2024

Secondary language

• Language: English
• Title: Determination of the diversity of the genus Viola by analyzing ribosomal DNA and measuring genome size

Abstract

The genus Viola, which belongs to the family Violaceae, includes more than 500 species distributed worldwide, ranging from the temperate zones of the Northern Hemisphere to the tropics and subtropics. First described in the 18th century, this genus encompasses both annual and perennial herbaceous plants. In this study, we focused on the morphological and genetic characteristics of Viola species to better understand their biodiversity and contribute to their conservation. Morphological traits, such as the shape and size of leaves, flowers, and stems, as well as flower color, are key for distinguishing between species. For the 50 collected samples of various species, we measured genome size using flow cytometry, revealing that genome size varies among species. The smallest average 2C value was recorded for Viola reichenbachiana Jord ex. Boreau, while the largest average 2C value was found in Viola x wittrockiana. One of the main challenges in ribosomal DNA analysis is the presence of secondary metabolites, which complicate DNA isolation and subsequent analyses. We found that the CTAB method was not effective for all species, necessitating adjustments or the use of alternative methods. Successful PCR reactions allowed for the determination of ribosomal DNA nucleotide sequences in 26 samples of different species, which we compared with NCBI BLAST databases. Ribosomal DNA sequencing showed that although the percentage of genetic identity among species is relatively high, there are distinct variations that allow differentiation between individual species.

• Keywords: Viola, genom size, 2C value, ribosomal DNA, sequencing