Genetic identification of selected woody plant using amplification of Chloroplast DNA sequences and DNA sequencing

Abstract

Gaharu or Aquilaria species is one of the well-known and expensive fragrance wood in the world. The study is carried out for genetic identification of selected samples of agarwood (Aquilaria spp) using amplification of chloroplast deoxyribonucleic acid (DNA) sequences based on primer pairs used and combination of DNA sequencing. Genetic identification of Aquilaria species was carried out through molecular approach as the genetic identity and diversity was not widely characterized and discovered in Gaharu Tea Valley plantation. A total of nine samples of leaves were obtained from block A, C and G for DNA extraction by using conventional cetyltrimethylammonium bromide (CTAB) method. Amplification of target region of chloroplast DNA by using specific primers followed by DNA sequencing and phylogenetic analysis was performed. The results showed that CTAB extraction method able to produce good quality of DNA from the samples. Single, bright band for each samples during visualization of agarose gel proved the amplification potential of designed primer pairs cp104 and cp105. The average length of nucleotides that being amplified by cp104 primer pair was 450 bp while 412 bp to 440 bp for cp105 primer pair. It revealed the potential of primer pairs on species discrimination of Aquilaria species. The high genetic similarity after comparison of database in NCBI though Basic Local Alignment Tool Search (BLAST) proved that the gaharu trees from block A, C and G belong to the same species Aquilaria sinensis.