In the present study, we employed microsatellite DNA markers to analyze the genetic diversity and differentiation between and within cultured stocks and wild populations of the orange-spotted grouper originating from the South China Sea and Southeast Asia. Compared to wild populations, genetic changes including reduced genetic diversity and significant differentiation have taken place in cultured grouper stocks, as shown by allele richness and heterozygosity studies, pairwise F(st), structure, molecular variance analysis, as well as multidimensional scaling analysis. Although two geographically adjacent orange-spotted grouper populations in China showed negligible genetic divergence, significant population differentiation was observed in wild grouper populations distributed in a wide geographical area from China, through Malaysia to Indonesia. However, the Mantel test rejected the isolation-by-distance model of genetic structure, which indicated the genetic differentiation among the populations could result from the co-effects of various factors, such as historical dispersal, local environment, ocean currents, river flows and island blocks. Our results demonstrated that microsatellite markers could be suitable not only for genetic monitoring cultured stocks but also for revealing the population structuring of wild orange-spotted grouper populations. Meanwhile, our study provided important information for breeding programs, management of cultured stocks and conservation of wild populations of the orange-spotted grouper.
Oil palm is an important perennial oil crop with an extremely long selection cycle of 10 to 12 years. As such, any tool that speeds up its genetic improvement process, such as marker-assisted breeding is invaluable. Previously, genetic linkage maps based on AFLP, RFLP and SSR markers were developed and QTLs for fatty acid composition and yield components identified. High density genetic maps of crosses of different genetic backgrounds are indispensable tools for investigating oil palm genetics. They are also useful for comparative mapping analyses to identify markers closely linked to traits of interest.
Over the last few decades, the use of molecular markers has played an increasing role in rice breeding and genetics. Of the different types of molecular markers, microsatellites have been utilized most extensively, because they can be readily amplified by PCR and the large amount of allelic variation at each locus. Microsatellites are also known as simple sequence repeats (SSR), and they are typically composed of 1-6 nucleotide repeats. These markers are abundant, distributed throughout the genome and are highly polymorphic compared with other genetic markers, as well as being species-specific and co-dominant. For these reasons, they have become increasingly important genetic markers in rice breeding programs. The evolution of new biotypes of pests and diseases as well as the pressures of climate change pose serious challenges to rice breeders, who would like to increase rice production by introducing resistance to multiple biotic and abiotic stresses. Recent advances in rice genomics have now made it possible to identify and map a number of genes through linkage to existing DNA markers. Among the more noteworthy examples of genes that have been tightly linked to molecular markers in rice are those that confer resistance or tolerance to blast. Therefore, in combination with conventional breeding approaches, marker-assisted selection (MAS) can be used to monitor the presence or lack of these genes in breeding populations. For example, marker-assisted backcross breeding has been used to integrate important genes with significant biological effects into a number of commonly grown rice varieties. The use of cost-effective, finely mapped microsatellite markers and MAS strategies should provide opportunities for breeders to develop high-yield, blast resistance rice cultivars. The aim of this review is to summarize the current knowledge concerning the linkage of microsatellite markers to rice blast resistance genes, as well as to explore the use of MAS in rice breeding programs aimed at improving blast resistance in this species. We also discuss the various advantages, disadvantages and uses of microsatellite markers relative to other molecular marker types.
Aggressive collections and trade activities in recent decades have resulted in heavy pressure on the natural stands of Aquilaria malaccensis and concerns over its long-term survival potential. To aid DNA profiling and assessment of its genetic diversity, microsatellite markers were developed for the species.
Species-specific simple sequence repeat (SSR) markers are favored for genetic studies and marker-assisted selection (MAS) breeding for oil palm genetic improvement. This report characterizes 20 SSR markers from an Elaeis oleifera genomic library (gSSR). Characterization of the repeat type in 2000 sequences revealed a high percentage of di-nucleotides (63.6%), followed by tri-nucleotides (24.2%). Primer pairs were successfully designed for 394 of the E. oleifera gSSRs. Subsequent analysis showed the ability of the 20 selected E. oleifera gSSR markers to reveal genetic diversity in the genus Elaeis. The average Polymorphism Information Content (PIC) value for the SSRs was 0.402, with the tri-repeats showing the highest average PIC (0.626). Low values of observed heterozygosity (H(o)) (0.164) and highly positive fixation indices (F(is)) in the E. oleifera germplasm collection, compared to the E. guineensis, indicated an excess of homozygosity in E. oleifera. The transferability of the markers to closely related palms, Elaeis guineensis, Cocos nucifera and ornamental palms is also reported. Sequencing the amplicons of three selected E. oleifera gSSRs across both species and palm taxa revealed variations in the repeat-units. The study showed the potential of E. oleifera gSSR markers to reveal genetic diversity in the genus Elaeis. The markers are also a valuable genetic resource for studying E. oleifera and other genus in the Arecaceae family.
The taxonomy of the causal pathogen of basal stem rot of oil palms, Ganoderma is somewhat problematic at present. In order to determine the genetic distance relationship between G. boninense isolates and non-boninense isolates, a random amplified microsatellites DNA (RAMS) technique was carried out. The result was then compared with interfertility data of G. boninense that had been determined in previous mating studies to confirm the species of G. boninense. Dendrogram from cluster analysis based on UPGMA of RAMS data showed that two major clusters, I and II which separated at a genetic distance of 0.7935 were generated. Cluster I consisted of all the biological species G. boninense isolates namely CNLB, GSDK 3, PER 71, WD 814, GBL 3, GBL 6, OC, GH 02, 170 SL and 348781 while all non-boninense isolates namely G. ASAM, WRR, TFRI 129, G. RES, GJ, and CNLM were grouped together in cluster II. Although the RAMS markers showed polymorphisms in all the isolates tested, the results obtained were in agreement with the interfertility data. Therefore, the RAMS data could support the interfertility data for the identification of Ganoderma isolates.
Trichoderma species are commercially applied as biocontrol agents against numerous plant pathogenic fungi due to their production of antifungal metabolites, competition for nutrients and space, and mycoparasitism. However, currently the identification of Trichoderma species from throughout the world based on micro-morphological descriptions is tedious and prone to error. The correct identification of Trichoderma species is important as several traits are species-specific. The Random Amplified Microsatellites (RAMS) analysis done using five primers in this study showed different degrees of the genetic similarity among 42 isolates of this genus. The genetic similarity values were found to be in the range of 12.50-85.11% based on a total of 76 bands scored in the Trichoderma isolates. Of these 76 bands, 96.05% were polymorphic, 3.95% were monomorphic and 16% were exclusive bands. Two bands (250 bp and 200 bp) produced by primer LR-5 and one band (250 bp) by primer P1A were present in all the Trichoderma isolates collected from healthy and infected oil palm plantation soils. Cluster analysis based on UPGMA of the RAMS marker data showed that T. harzianum, T. virens and T. longibrachiatum isolates were grouped into different clades and lineages. In this study we found that although T. aureoviride isolates were morphologically different when compared to T. harzianum isolates, the UPGMA cluster analysis showed that the majority isolates of T. aureoviride (seven from nine) were closely related to the isolates of T. harzianum.
Data from three published studies of genetic variation at 18 microsatellite loci in water buffalo populations in China (18 swamp type, two river type), Nepal (one wild, one domestic river, one hybrid) and south-east Asia (eight swamp, three river) were combined so as to gain a broader understanding of genetic relationships among the populations and their demographic history. Mean numbers of alleles and expected heterozygosities were significantly different among populations. Estimates of θ (a measure of population differentiation) were significant among the swamp populations for all loci and among the river populations for most loci. Differentiation among the Chinese swamp populations (which was due primarily to just one population) was much less than among the south-east Asian. The Nepal wild animals, phenotypically swamp type but genetically like river type, are significantly different from all the domestic river populations and presumably represent the ancestral Bubalus arnee (possibly with some river-type introgression). Relationships among the swamp populations (D(A) genetic distances, principal component analysis and structure analyses) show the south-east Asian populations separated into two groups by the Chinese populations. Given these relationships and the patterns of genetic variability, we postulate that the swamp buffalo was domesticated in the region of the far south of China, northern Thailand and Indochina. Following domestication, it spread south through peninsular Malaysia to Sumatra, Java and Sulawesi, and north through China, and then to Taiwan, the Philippines and Borneo.
Horseshoe crabs are said to be declining worldwide. However, there is still no published report on the status of horseshoe crabs in Malaysia. Thus, we report here eight informative microsatellite markers that were developed using the 5'-anchored ISSR-PCR enrichment procedure to diagnose the population genetic structure of the mangrove horseshoe crab, Carcinoscorpius rotundicauda from Peninsular Malaysia. This set of markers was tested on 127 samples and showed polymorphism in this species. Hence they should be useful in future essential population genetic studies of these living fossils in the Southeast Asian region.
Analysis of variable number tandem repeats (VNTRs) by polymerase chain reaction (PCR) is a common method used to predict engraftment status in post-allogeneic haematopoeitic stem cell transplantation (HSCT) patients. Different populations have different copies of repeated DNA sequence and hence, different percentage of informativeness between patient and donor.
We describe a chromosome 6 uniparental disomy (UPD6) in a boy, discovered during a screening for the genetic cause of cleft lip and palate. In the medical literature, almost all documented cases of UPD6 are paternal in origin, and only four were maternal. We present here a report of complete maternal chromosome 6 uniparental heterodisomy. Haplotype analysis was performed using highly polymorphic short tandem repeat (STR) markers that span both arms of chromosome 6. Analysis of these markers revealed the presence of two maternal alleles but no paternal allele, indicating an instance of maternal uniparental heterodisomy. Chromosome analysis of peripheral blood lymphocytes confirmed a normal male karyotype. Advanced maternal age at the time of the infant's birth and heterodisomy of markers around the centromere favors a meiosis-I error. No specific phenotype has been reported for maternal UPD6. Therefore, the cleft lip and palate in the present case probably occurred due to other risk factors. This report provides further evidence that maternal UPD6 has no specific clinical consequences and adds to the collective knowledge of this rare chromosomal finding.
Invasive species are one of the main sources of the ongoing global loss of biodiversity. Invasive ants are known as particularly damaging invaders and their introductions are often accompanied by population-level behavioural and genetic changes that may contribute to their success. Anoplolepis gracilipes is an invasive ant that has just recently received increased attention due to its negative impact on native ecosystems. We examined the behaviour and population structure of A. gracilipes in Sabah, Malaysia. A total of 475 individuals from 24 colonies were genotyped with eight microsatellite markers. Intracolonial relatedness was high, ranging from 0.37 to 1 (mean +/- SD: 0.82 +/- 0.04), while intercolonial relatedness was low (0.0 +/- 0.02, range -0.5-0.76). We compared five distinct sampling regions in Sabah and Brunei. A three-level hierarchical F-analysis revealed high genetic differentiation among colonies within the same region, but low genetic differentiation within colonies or across regions. Overall levels of heterozygosity were unusually high (mean H(O) = 0.95, mean H(E) = 0.71) with two loci being entirely heterozygous, indicating an unusual reproductive system in this species. Bioassays revealed a negative correlation between relatedness and aggression, suggesting kinship as one factor facilitating supercolony formation in this species. Furthermore, we genotyped one individual per nest from Sabah (22 nests), Sarawak (one nest), Brunei (three nests) and the Philippines (two nests) using two mitochondrial DNA markers. We found six haplotypes, two of which included 82.1% of all sequences. Our study shows that the sampled area in Sabah consists of a mosaic of differently interrelated nests in different stages of colony establishment. While some of the sampled colonies may belong to large supercolonies, others are more likely to represent recently introduced or dispersed propagules that are just beginning to expand.
The diamondback moth, Plutella xylostella, is renowned for developing resistance to insecticides and causing significant economic damage to Brassica vegetable crops throughout the world. Yet despite its economic importance, little is known about the population structure and movement patterns of this pest both at local and regional scales. In Australia, the movement patterns and insecticide resistance status of P. xylostella infesting canola, vegetables, forage brassicas and weeds have fundamental implications for the management of this pest. Here we use six polymorphic microsatellite loci to investigate population structure and gene flow in Australian populations of P. xylostella. Samples of P. xylostella from New Zealand, Malaysia, Indonesia and Kenya were also scored at these loci. We found no evidence of population structure within Australia, with most populations having low inbreeding coefficients and in Hardy-Weinberg equilibrium. In addition, a sample from the North Island of New Zealand was indistinguishable from the Australian samples. However, large genetic differences were found between the Australia/New Zealand samples and samples from Kenya, Malaysia and Indonesia. There was no relationship between genetic distance and geographic distance among Australian and New Zealand samples. Two of the loci were found to have null alleles, the frequency of which was increased in the populations outside the Australia/New Zealand region. We discuss these results with reference to insecticide resistance management strategies for P. xylostella in Australia.
The extent to which response to environmental change is mediated by species-specific ecology is an important aspect of the population histories of tropical taxa. During the Pleistocene glacial cycles and associated sea level fluctuations, the Sunda region in Southeast Asia experienced concurrent changes in landmass area and the ratio of forest to open habitat, providing an ideal setting to test the expectation that habitat associations played an important role in determining species' response to the opportunity for geographic expansion. We used mitochondrial control region sequences and six microsatellite loci to compare the phylogeographic structure and demographic histories of four broadly sympatric species of Old World fruit bats in the genus, Cynopterus. Two forest-associated species and two open-habitat generalists were sampled along a latitudinal transect in Singapore, peninsular Malaysia, and southern Thailand. Contrary to expectations based on habitat associations, the geographic scale of population structure was not concordant across ecologically similar species. We found evidence for long and relatively stable demographic history in one forest and one open-habitat species, and inferred non-coincident demographic expansions in the second forest and open-habitat species. Thus, while these results indicate that Pleistocene climate change did not have a single effect on population structure across species, a correlation between habitat association and response to environmental change was supported in only two of four species. We conclude that interactions between multiple factors, including historical and contemporary environmental change, species-specific ecology and interspecific interactions, have shaped the recent evolutionary histories of Cynopterus fruit bats in Southeast Asia.
This study was carried out to evaluate the genetic effect of quantitative trait loci (QTLs) conferring drought tolerance in wheat. A population of 120 F(2) individuals from the cross between the drought-tolerant S-78-11 and drought-sensitive Tajan cultivars were analyzed for their segregation under drought stress conditions. The relative water content under drought stress conditions exhibited continuous variation, indicating the minor gene effects on the trait. Single-marker analysis (SMA) was carried out to detect the main QTL association with drought tolerance. The SMA results revealed that the simple sequence repeat markers GWM182 and GWM292 on chromosome 5D and GWM410 on chromosome 5A exhibited significant association with drought tolerance, accounting for 30, 22, and 21% of the total variation, respectively. The 3 genetic loci, especially GWM182, can be used in marker-assisted selection methods in drought tolerance breeding in wheat.
We investigated the genetic structure within and among Bornean orang-utans (Pongo pygmaeus) in forest fragments of the Lower Kinabatangan flood plain in Sabah, Malaysia. DNA was extracted from hair and faecal samples for 200 wild individuals collected during boat surveys on the Kinabatangan River. Fourteen microsatellite loci were used to characterize patterns of genetic diversity. We found that genetic diversity was high in the set of samples (mean H(E) = 0.74) and that genetic differentiation was significant between the samples (average F(ST) = 0.04, P < 0.001) with F(ST) values ranging from low (0.01) to moderately large (0.12) values. Pairwise F(ST) values were significantly higher across the Kinabatangan River than between samples from the same river side, thereby confirming the role of the river as a natural barrier to gene flow. The correlation between genetic and geographical distance was tested by means of a series of Mantel tests based on different measures of geographical distance. We used a Bayesian method to estimate immigration rates. The results indicate that migration is unlikely across the river but cannot be completely ruled out because of the limited F(ST) values. Assignment tests confirm the overall picture that gene flow is limited across the river. We found that migration between samples from the same side of the river had a high probability indicating that orang-utans used to move relatively freely between neighbouring areas. This strongly suggests that there is a need to maintain migration between isolated forest fragments. This could be done by restoring forest corridors alongside the river banks and between patches.
Clonal reproduction of oil palm by means of tissue culture is a very inefficient process. Tissue culturability is known to be genotype dependent with some genotypes being more amenable to tissue culture than others. In this study, genetic linkage maps enriched with simple sequence repeat (SSR) markers were developed for dura (ENL48) and pisifera (ML161), the two fruit forms of oil palm, Elaeis guineensis. The SSR markers were mapped onto earlier reported parental maps based on amplified fragment length polymorphism (AFLP) and restriction fragment length polymorphism (RFLP) markers. The new linkage map of ENL48 contains 148 markers (33 AFLPs, 38 RFLPs and 77 SSRs) in 23 linkage groups (LGs), covering a total map length of 798.0 cM. The ML161 map contains 240 markers (50 AFLPs, 71 RFLPs and 119 SSRs) in 24 LGs covering a total of 1,328.1 cM. Using the improved maps, two quantitative trait loci (QTLs) associated with tissue culturability were identified each for callusing rate and embryogenesis rate. A QTL for callogenesis was identified in LGD4b of ENL48 and explained 17.5% of the phenotypic variation. For embryogenesis rate, a QTL was detected on LGP16b in ML161 and explained 20.1% of the variation. This study is the first attempt to identify QTL associated with tissue culture amenity in oil palm which is an important step towards understanding the molecular processes underlying clonal regeneration of oil palm.
Malaysian rice, Pongsu Seribu 2, has wide-spectrum resistance against blast disease. Chromosomal locations conferring quantitative resistance were detected by linkage mapping with SSRs and quantitative trait locus (QTL) analysis. For the mapping population, 188 F3 families were derived from a cross between the susceptible cultivar, Mahsuri, and a resistant variety, Pongsu Seribu 2. Partial resistance to leaf blast in the mapping population was assessed. A linkage map covering ten chromosomes and consisting of 63 SSR markers was constructed. 13 QTLs, including 6 putative and 7 putative QTLs, were detected on chromosomes 1, 2, 3, 5, 6, 10, 11 and 12. The resulting phenotypic variation due to a single QTL ranged from 2 to 13 %. These QTLs accounted for approx. 80 % of the total phenotypic variation within the F3 population. Therefore, partial resistance to blast in Pongsu Seribu 2 is due to combined effects of multiple loci with major and minor effects.
Tomistoma schlegelii, also referred to as the "false gharial", is one of the most exclusive and least known of the world's fresh water crocodilians, limited to Southeast Asia. Indeed, lack of economic value for its skin has led to neglect the biodiversity of the species. The current study aimed to investigate the mentioned case using 40 simple sequence repeat (SSR) primer pairs and 45 inter-simple sequence repeat (ISSR) primers. DNA analysis of 17 T. schlegelii samples using the SSR and ISSR markers resulted in producing a total of 49 and 108 polymorphic bands, respectively. Furthermore, the SSR- and ISSR-based cluster analyses both generated two main clusters. However, the SSR based results were found to be more in line with the geographical distributions of the crocodile samples collected across the country as compared with the ISSR-based results. The observed heterozygosity (HO) and expected heterozygosity (HE) of the polymorphic SSRs ranged between 0.588-1 and 0.470-0.891, respectively. The present results suggest that the Malaysian T. schlegelii populations had originated from a core population of crocodiles. In cooperation with the SSR markers, the ISSRs showed high potential for studying the genetic variation of T. schlegelii, and these markers are suitable to be employed in conservation genetic programs of this endangered species. Both SSR- and ISSR-based STRUCTURE analyses suggested that all the individuals of T. schlegelii are genetically similar with each other.
The maintenance of mixed mating was studied in Shorea curtisii, a dominant and widely distributed dipterocarp species in Southeast Asia. Paternity and hierarchical Bayesian analyses were used to estimate the parameters of pollen dispersal kernel, male fecundity and self-pollen affinity. We hypothesized that partial self incompatibility and/or inbreeding depression reduce the number of selfed seeds if the mother trees receive sufficient pollen, whereas reproductive assurance increases the numbers of selfed seeds under low amounts of pollen. Comparison of estimated parameters of self-pollen affinity between high density undisturbed and low density selectively logged forests indicated that self-pollen was selectively excluded from mating in the former, probably due to partial self incompatibility or inbreeding depression until seed maturation. By estimating the self-pollen affinity of each mother tree in both forests, mother trees with higher amount of self-pollen indicated significance of self-pollen affinity with negative estimated value. The exclusion of self-fertilization and/or inbreeding depression during seed maturation occurred in the mother trees with large female fecundity, whereas reproductive assurance increased self-fertilization in the mother trees with lower female fecundity.