The Detection of Anaphalis spp. Genetic Diversity Based on Molecular Character (using ITS, ETS, and EST-SSR markers)

Abstract

Anaphalis is the natural vegetation component in mountainous areas and is found in volcanic soils. Anaphalis is one of the species which existence is currently presumed to decrease and is feared to be extinct in nature. The purpose of this study was to detect genetic diversity of Anaphalis spp. in Bromo Tengger Semeru National Park (BTSNP) based on molecular characters by using ITS, ETS, and EST-SSR markers in supporting the conservation aspects of Anaphalis genetically. This research method was carried out by exploring the existence of Anaphalis populations, their coordinates marked by GPS, and leaf samples collected as a source of molecular analysis material. PCR products from ITS and ETS markers were sequenced, and the results’ phylogenetic were analyzed using the MEGA6 program. PCR products from EST-SSR markers were performed by scoring DNA bands (allel) and both variations and genetics population were analyzed by using the POPGENE 1.32 program. Anaphalis populations found in BTSNP are Anaphalis javanica, A. longifolia, and A. viscida. The genetic diversity of Anaphalis spp. in BTSNP has polymorphism potential that is high enough, which is 57% (ITS) and 31% (ETS-partial), with a maximum likelihood phylogenetic tree topology which is monophyletic separated into four clusters consisting one cluster outgroup and three others being an ingroup. Whereas based on the genetic diversity value of the EST-SSR sequences in Anaphalis spp. in BTSNP shows that only A. longifolia populations in the Ranu Kumbolo area that have a high genetic diversity value (0.024) compared to the other two Anaphalis species. The highest genetic distance of Anaphalis spp. BTSNP in A. longifolia is found in the population of Penanjakan and Mt. Batok areas (0.040) with the smallest gene flow rate (0.428). Further research is needed to obtain a more complete picture of Anaphalis genetic diversity by using more molecular markers, bigger population numbers, more individuals and in bigger populations in other conservation areas in supporting the Anaphalis conservation strategy program.