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Tropentag, September 17 - 19, 2014 in Prague, Czech Republic

"Bridging the gap between increasing knowledge and decreasing resources"

Inter Simple Sequence Repeat (ISSR) Markers as Reproducible Tools for Genetic Diversity Analysis of an Agroforestry Tree Species Guazuma crinita from Peruvian Amazon

Lady Laura Tuisima Coral, Petra Hlásná Cepková, Bohdan Lojka

Czech University of Life Sciences Prague, Fac. of Tropical AgriSciences, Dept. of Crop Sciences and Agroforestry, Czech Republic


Agroforestry system is an efficient method for sustainable use and conservation of tropical trees. Guazuma crinita is an important fast-growing timber tree species for agroforestry systems in the Peruvian Amazon and little is known about its genetic variation. A molecular approach using inter-simple sequence repeat (ISSR) markers was applied to 45 genotypes of G. crinita from a collection of clones from a small and representative watershed in Peruvian Amazon. To obtain clear and reproducible bands, 30 ISSR primers and 2 parameters (annealing temperature, DNA concentrations) were screened. The optimal conditions for ISSR system for each 20 µl PCR reaction mixture was composed of 10 µl of 2× PPP Master Mix, 0.5 µl of respective ISSR primer, 2 µl of DNA (50 ng/µl), 0.2 µl of BSA and 7.3 µl PCR H2O. Of the total 30 ISSR markers, seven primers produced well reproducible bands with optimal annealing temperature (Ta) varying from 47 to 52°C. A total of 47 were amplified of which 44 were polymorphic (93.62%). The range of DNA amplification varied from 260 to 1700 bp. The general genetic differentiation (Gst) was estimated to be 0.40 and the gen flow (Nm) 0.75 alleles per generation. The analysis of molecular variation (AMOVA) assigned 100% of genetic variability to within provenances diversity. The UPGMA analysis did not distinguish provenances relationship suggesting a common gene pool. The genetic assessment suggests the collection of plant material for the establishment of genebank and in situ conservation strategies program for this tree species. ISSR markers were chosen because the technique is very simple, fast and it also requires small quantity of sample DNA.

Keywords: Genetic diversity, Guazuma crinita, inter-simple sequence repeat marker, sustainable conservation

Contact Address: Bohdan Lojka, Czech University of Life Sciences Prague, Fac. of Tropical AgriSciences, Dept. of Crop Sciences and Agroforestry, Kamýcká 129, 169 21 Prague, Czech Republic, e-mail: lojka@ftz.czu.cz

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