Genetic characterization and phylogenetic study of Indonesian cuscuses from Maluku and Papua Island based on 16S rRNA gene.

BACKGROUND AND AIM: Indonesian cuscuses are now becoming scarce because of the reduction of habitat and poaching. Further, molecular characterization of Indonesian cuscuses is still very lacking. This study aimed to determine genetic markers and phylogenetic relationships of Indonesian cuscuses based on 16S rRNA gene sequences. MATERIALS AND METHODS: This study used 21 cuscuses caught from two provinces and 16 islands: 13 from Maluku and eight from Papua. Cuscus samples were taken by biopsy following ethics guidelines for animals. The genome isolation was done using gSYNC DNA Mini Kit (Geneaid Biotech Ltd., Taiwan). The 16S rRNA gene was amplified by primers (16SKUSAF and 16SKUSAR), and the polymerase chain reaction product obtained was 1875 base pair (bp). The analysis of genetic characterization and the phylogenetic relationship was performed usingMEGA version X software (https://www.megasoftware.net/). RESULTS: 16S rRNA gene sequencing attained 1598 bp for all samples. Based on the 16S rRNA nucleotide sequences, cuscuses from Papua and Maluku belong to the genus Phalanger and Spilocuscus. Phalanger spp. and Spilocuscus spp. from Papua can be distinguished from Phalanger and Spilocuscus from Maluku, except Spilocuscus from Ternate has a very close relationship with cuscus from Sentani, Papua. CONCLUSION: Indonesian cuscuses were derived into two clades based on 16S rRNA gene sequence, one group to genus Phalanger and another group to Spilocuscus.

Genetic characterization and phylogenetic study of Lakor goat from Southwest Maluku Regency based on mitochondrial COI gene.

AIM: This study is aimed at characterizing the genetic and phylogenetic structure of Lakor goats as indigenous livestock from the Southwest Maluku Regency based on mitochondrial COI gene sequences. MATERIALS AND METHODS: The genomes of 103 follicle samples from Lakor goats, collected from Lakor Island, were analyzed. The polymerase chain reaction was used to amplify 1548 bp of the mitochondrial COI gene using two primer pairs (COIA and COIB). Following sequencing, genetic variation and phylogenetic relationship were established using MEGA version X software. RESULTS: The results of multiple COI gene alignment of the total sequences identified four polymorphic nucleotides that function as genetic markers between individual animals within the Lakor goat population. These correspond to positions 228 (A-G), 519 (G-A), 900 (C-T), and 1266 (T-C). Phylogenetic signals based on the COI gene showed that Lakor goat breed is a monophyletic group or single clade with a bootstrap value of 100% by the neighbor-joining (NJ) and maximum likelihood (ML) evolutionary models. This data indicated that evolutionarily, the Lakor goat breed has a very close kinship with three goat breeds from China: The Meigu goat (KM 244714.1), Chinese Tibet (Capra hircus) (KJ 940969.1), and C. hircus (KP 677510.1). Phylogenetic information based on the cladistics system classified the Lakor goat as a single clade (monophyletic group). The low-genetic diversity within populations indicates that there has been an inbreeding depression occurring at a very high frequency. CONCLUSION: We conclude that the Lakor goat may be divided into a single clade or monophyletic group based on the COI gene sequence. Four nucleotides were identified that can be used as genetic markers among individual animals within the Lakor goat population, as well as C. hircus and others as derived from GenBank data. The Lakor goat population has a high level of inbreeding depression as a result of geographical isolation, which supports the formation of a monophyletic group with different genetic characteristics, and does not allow the introduction of males from other breeds. Phylogenetic signals indicated that Capra aegagrus (bezoar) is the ancestor of the native goats in Indonesia, including the Lakor goats.

Study of Genetic Marker of Cuscuses (Marsupialia: Phalangeridae) from Maluku and Papua Based on Cytochrome b Gene Sequences.

Cuscuses is marsupials animal (Phalangeridae) which has limited spread in eastern Indonesia (Sulawesi, Maluku, Papua and Timor islands), Australia and Papua New Guinea. The ex-situ and in-situ conservation of cuscuses under captivating condition is an alternative solution to protect from extinction. This study aimed to determine nucleotide sequences and genetic marker on cyt b gene with sequencing method of each species on two provinces. Whole genome DNA was extracted from 22 samples of cuscuses obtained from different habitats, Maluku (13 individuals) and Papua (8 individuals) according to the protocol of Qiamp DNA Blood Mini Kit (Qiagen) and then it was used as template for amplification of cyt b gene by using PCR method. The PCR product were then purified using column chromatography and were used as template for sequencing reaction. Results sequencing of cyt b gene were analyzed using MEGA program versions 6.0. The PCR product gives results nucleotides of 982 bp according to database GeneBank and sequencing product gives results nucleotides of 771 bp. Nucleotides alignment of Phalanger members was found 24 nucleotides distinguishing and Spilocuscus members was found 11 nucleotides distinguishing, which can be used as genetic marker between Phalanger and Spilocuscus members from Papua and Maluku.