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Establishing the taxonomic rank of the endemic Romanian species Onobrychis transsilvanica
through molecular methods
CNDI-UEFISCDI, project number: PU-II-RU-PD-2012-3-0005-15/26.04.2013
Project summary
A major problem which appears in taxonomic studies in our country and beyond is represented by the problematic species, with overlapping characters and serious problems in their classification. This is the case for the species Onobrychis transilvanica. Our aim is to establish its taxonomical relationship with the taxon O. montana. Based on previous studies and because the interpretation of taxonomic position of O. transsilvanica is supported solely on morphological characters that proved to be inadequate, we believe the application of current molecular methods such as: microsatellites and DNA sequencing data from the plastid genom (cpDNA) is important in order to elucidate the taxonomic status of the species O. transsilvanica. The main goal of this project is to investigate whether there is a genetically sound basis for supporting Onobrychis transsilvanica as a distinct taxonomic unit. We also aim to identify molecular markers which are polymorphic enough to discriminate between Onobrychis transsilvanica and O. montana and to investigate taxonomic boundaries within the O. montana-O. transsilvanica group in the SE Carpathians. The molecular studies proposed in this project fit into the present approach which is currently used in the modern studies of taxonomy and phylogeny.
Project team
Postdoc, project manager: dr. Ioan Bacila, ioan.bacila@icbcluj.ro
Mentor, dr. Halmagyi Adela, adela.halmagyi@icbcluj.ro
Project structure
2013
Phase 1: Molecular characterization of Onobrychis transsilvanica and O. montana using chloroplast DNA sequence
The main objective of the phase 1
O1: Identification of the chloroplastic sequences able to distinguish between Onobrychis transsilvanica and Onobrychis montana
The main steps:
1. - DNA extraction from leaves stored in silicagel.
2. - Testing multiple chloroplastic regions in order to obtain the amplicons.
3. - Purification and sequencing of the selected amplicons.
4. - Data analysis using bioinformatics tools and results interpretation.
5. - Scientific progress report.
The expected results:
1. - list with the chloroplast regions selected.
2. -cpDNA amplicons for all the analysed populations.
3. -cpDNA sequences for all the analysed populations.
4. -phylogenetic trees and diversity indexes.
2014
Phase 2: Molecular characterization of Onobrychis transsilvanica and O. montana using microsatellites (SSR) markers.
The main objectives of the phase
O1: Testing multiple sets of SSR markers taken from literature to select optimal SSR molecular markers able to distinguish between Onobrychis transsilvanica and O. montana.
The main steps:
1. - Scientific literature consultation in order to select SSR markers potentially usable for the analysed species.
2. - PCR amplification of SSR markers selected in the previous step and capillary electrophoresis of the obtained amplicons.
3. - Selecting optimal SSR markers for the analysed species.
The expected results:
1.- SSR marker list selected from the scientific literature used for testing.
2.- Amplicons and genetic profiles for the samples used in testing.
3.The optimal SSR marker list selected for the analysed species.
O2: Using optimal SSR markers selected in the previous objective to obtain data for the entire set of sampled populations.
The main steps:
1. - Applying PCR amplification of the selected SSR markers to the entire set of sampled populations.
2. - Capillary electrophoresis of the obtained amplicons.
3. - Scientific progress report.
The expected results:
1.- Amplicons for the entire set of populations.
2.- Genetic profiles for the entire set of populations.
3.- Scientific progress report.
2015
O1: Data analysis and processing of the results previously obtained.
The main steps:
1. - Alleles-scoring of the obtained genetic profiles.
2. - Data analysis using specific programs and results interpretation.
3. - Final scientific report.
4. - Scientific publications.
The expected results:
1.- 1/0 matrix obtained from scoring.
2.- Phylogenetic trees and diversity indexes.
3.- Final scientific report.
4.- Scientific articles based on the project results.