The use of DNA barcoding methods for identification of tropical trees

Bc. Barbora Legezová

Master's thesis

Abstract:

DNA barcoding allows fast, accurate, automatable and relatively cheap species identification by using a short, standardized DNA region. This method has proven effective mainly among animal species, however, the method is quickly evolving to be suitable for plant species as well. DNA barcoding of tropical trees is a very specific case, due to the often-inaccessible canopy, there is a need to barcode other plant tissues than the leaf, which may bring several complications. The aim of this thesis was to evaluate the DNA barcoding method for identification of tropical trees by both traditional taxonomic and molecular methods. The reliability of both species recognition methods were compared as well as the results among different tissue types (leave, young wood, old wood). 11 tree species, indigenous to Central and West Africa, were sampled. In total 150 samples were collected both for the morphological and genetic analyses. The DNA extraction was done using the CTAB method and the leaf material yielded the best nucleic acid concentration with the highest purity. For PCR amplification the trnL approach was used, the samples were amplified in three different regions of various length. The ‘long’ sequence (primer pair c-d) appeared fragmented when visualized by electrophoresis. When sent for Sanger sequencing the short region (primer pair g-h) returned with low taxonomic discrimination and the sequencing success rate was low as well. Therefore, for the means of sequencing the middle region (primer pair c-h) was used. The sequencing success rate was 92%. All the tree species were correctly assigned to their family, 48% to the correct genus and 23% to their species. However, the low recognition to higher taxa may be also given by the small number of listed sequences in the GenBank database. Therefore, the best sequence per species was chosen and submitted to the GenBank database, which now can be found there under their accession numbers and used for future barcode identifications.

Keywords

Cameroon, plant barcode, tree molecular identification, trnL intron, trnL approach, wood barcoding

Language used: English

Date on which the thesis was submitted / produced: 14. 8. 2020

Thesis defence

• Supervisor: doc. Ing. Bohdan Lojka, Ph.D.

Czech University of Life Sciences Prague

Faculty of Tropical AgriSciences

Master programme / field:
Forestry Engineering / Tropical Forestry and Agroforestry