Geographically isolated populations of endemic orchids have evolved and adapted to an existence within specifi c ecological niches. These populations are highly susceptible to anthropogenic
infl uences on their microhabitats. The primary objective of conservation programs is the restoration of endangered populations to their ecologically sustainable levels, and the fi rst stage in the process of conservation involves estimation of molecular diversity at the level of the population. The approach described in this article involves the application of RAPD, Microsatellites and Chloroplast DNA markers for the characterization of the genetic structure of Paphiopedilum rothschildianum and Phalaenopsis gigantea, two endangered and endemic orchids of Sabah. This study has isolated a total of 96 microsatellite loci in P. rothschildianum and P. gigantea, 42 specifi c primer pairs have been designed for amplifi cation of microsatellite loci and are currently being applied to screen the breeding pools. The Chloroplast DNA regions amplifi ed by the primer pairs trnH-psbA and trnL-trnF exhibit distinct polymorphisms and can be used to establish phylogenetic
relationships. The ability of microsatellite loci to cross-amplify selected varieties of orchids has been determined. The molecular markers developed will be applied to estimate population diversity
levels and to formulate long-term management strategies for the conservation of endangered species of orchids of Sabah.
Pineapple (Ananas comosus var. comosus) is known as the king of fruits for its crown and is the third most important tropical fruit after banana and citrus. The plant, which is indigenous to South America, is the most important species in the Bromeliaceae family and is largely traded for fresh fruit consumption. Here, we report the complete chloroplast sequence of the MD-2 pineapple that was sequenced using the PacBio sequencing technology.
Improving the quality of the non-climacteric fruit, pineapple, is possible with information on the expression of genes that occur during the process of fruit ripening. This can be made known though the generation of partial mRNA transcript sequences known as expressed sequence tags (ESTs). ESTs are useful not only for gene discovery but also function as a resource for the identification of molecular markers, such as simple sequence repeats (SSRs). This paper reports on firstly, the construction of a normalized library of the mature green pineapple fruit and secondly, the mining of EST-SSRs markers using the newly obtained pineapple ESTs as well as publically available pineapple ESTs deposited in GenBank. Sequencing of the clones from the EST library resulted in 282 good sequences. Assembly of sequences generated 168 unique transcripts (UTs) consisting of 34 contigs and 134 singletons with an average length of ≈500 bp. Annotation of the UTs categorized the known proteins transcripts into the three ontologies as: molecular function (34.88%), biological process (38.43%), and cellular component (26.69%). Approximately 7% (416) of the pineapple ESTs contained SSRs with an abundance of trinucleotide SSRs (48.3%) being identified. This was followed by dinucleotide and tetranucleotide SSRs with frequency of 46 and 57%, respectively. From these EST-containing SSRs, 355 (85.3%) matched to known proteins while 133 contained flanking regions for primer design. Both the ESTs were sequenced and the mined EST-SSRs will be useful in the understanding of non-climacteric ripening and the screening of biomarkers linked to fruit quality traits.
Epinephelus fuscoguttatus is a commercially important marine fish species in southeast Asia. Due to overfishing and water pollution, this species has been declared as near-threatened. Thus, to provide information to help maintain and preserve the species, microsatellites were developed, using an enriched genomic library method. Thirty individuals were collected from the hatchery of the Fishery Research Institute, Terengganu, Malaysia. These individuals, from four to six years old, originated from Sabah and are maintained in captive culture as broodstock. Genomic DNA was extracted from the fins of selected individuals that weighed 3-8 kg. Ten microsatellite loci were found to be polymorphic in this population, with 5 to 21 alleles per locus. Observed and expected heterozygosities ranged from 0.53 to 0.97 and 0.59 to 0.95, respectively. Only one locus deviated significantly from Hardy-Weinberg equilibrium and no significant linkage disequilibrium was found among the pairs of loci. These polymorphic microsatellite loci will be used by the Malaysian Fishery Research Institute for investigating genetic diversity and for developing breeding strategies.