Oil palm is continually being improved via controlled crossing of selected palms to ensure sustainable yields and productivity. As such, correct parental assignment is important as the presence of illegitimates will compromise the progress of improvement. In the present study, we determined the optimal number of microsatellite (SSR) markers for detection of illegitimates in selected oil palm crosses with high confidence. Determining the optimal number of markers to assign parentage will ensure that the DNA fingerprinting will be cost effective for routine use as a quality control tool in oil palm improvement programs. Here, we evaluated a wide range of crosses that included a cross derived from wild germplasm palm. The results revealed that markers with high PIC are informative and detect most of the alleles present in a cross, including those exhibited by the illegitimates. A larger number of optimum sets of markers are needed to detect all illegitimates for crosses with higher levels of genetic diversity. The optimal number of polymorphic SSR markers determined in the present study can ensure that appropriate quality control is implemented for oil palm improvement programs.
The bioavailability of a generic preparation of acyclovir (Avorax) was compared with the innovator product, Zovirax. Twelve healthy volunteers participated in the study, conducted according to a randomized, two-way crossover design. The preparations were compared using the parameters area under the plasma concentration time curve (AUC(0-infinity), peak plasma concentration (Cmax), and time to reach peak plasma concentration (Tmax). No statistically significant difference was observed between the Tmax or the logarithmic transformed AUC(0-infinity) and C(max) values of the two preparations. In addition, the 90% confidence interval for the ratio of the logarithmic transformed AUC(0-infinity) values of Avorax over those of Zovirax was found to lie between 0.85 and 1.06, while that of the logarithmic transformed Cmax values was between 0.95 and 1.25, being within the bioequivalence limit of 0.80-1.25. Moreover, the elimination rate constant (k(e)), elimination half-life (t(1/2)), and apparent volume of distribution (Vd) values obtained with the two preparations were comparable and not significantly different statistically.
A study was conducted to compare the in vivo bioavailability of a generic metoprolol tablet preparation (Metoprolol) with that of the innovator product, Betaloc. Both preparations have a labeled dose of 100 mg metoprolol tartrate. Twelve healthy adult male volunteers participated in the study, which was conducted according to a standard two-way crossover design with a washout period of 1 week. The bioavailability was compared using the total area under the plasma level versus time curve (AUC0-infinity), peak plasma concentration (Cmax), and time to reach peak plasma concentration (Tmax). No statistically significant difference was observed between the logarithmically transformed AUC0-infinity values or the logarithmically transformed Cmax values of the two preparations. However, a statistically significant difference was observed between the Tmax values, but may not be therapeutically significant or important. Moreover, the 90% confidence interval (CI) for the ratio of the logarithmically transformed AUC0-infinity values of Metoprolol over those of Betaloc was calculated to be between 0.94 and 1.02, while that of Cmax was between 0.98 and 1.01, both of which are within the acceptable limit of 0.80-1.25. From the data obtained, it was also observed that a high proportion of our volunteers of Asian origin appeared to be poor metabolizers of metoprolol, which was consistent with what had been observed in our previous study of another preparation of metoprolol.
Glucose phosphate isomerase (E.C. 5.3.1.9) and phosphoglucomutase (E.C. 2.7.5.1) were found to be polymorphic in a laboratory colony of Aedes albopictus. The glucose phosphate isomerase locus is represented by two alleles resulting in three genotypes, while the phosphoglucomutase locus is represented by at least five alleles giving rise to a total of 15 genotypes. The inheritance of these two enzymes is of the Mendelian type with codominant alleles. Present data indicate that these genes are not linked.Of 105 mosquitoes analysed for these two gene-enzyme systems, the frequencies for glucose phosphate isomerase alleles are Gpi (S)=0.68 and Gpi (F)=0.32, while the frequencies for phosphoglucomutase alleles are Pgm (A)=0.16, Pgm (B)=0.11, Pgm (C)=0.19, Pgm (D)=0.30 and Pgm (F)= 0.24. The frequencies of the three glucose phosphate isomerase genotypes are in accord with Hardy-Weinberg expectations (X 1 (2) =2.74). Similarly, the frequencies of the 15 phosphoglucomutase genotypes probably do not differ significantly from Hardy-Weinberg expectations (X 10 (2) = 18.45).
The genetics of glucosephosphate isomerase (E.C. 5.3.1.9) in two strains (Malaysian and Taiwan) of Aedes togoi is reported. Three electrophoretic phenotypes were presented in both sexes. The zymogram patterns were identical in both strains of A. togoi. The phenotypes were governed by a pair of codominant alleles. The allele frequency of the slow-moving band was 0.63 in the Malaysian strain adn was 0,86 and 0.82 in F161 and F169 generations, respectively, of the Taiwan strain. The sample studied was in good accord with Hardy-Weinberg expectation.
Bioactive compounds from the medicinal plant, Eurycoma longifolia Jack have been shown to promote anti-proliferative effects on various cancer cell lines. Here we examined the effects of purified eurycomanone, a quassinoid found in Eurycoma longifolia Jack extract, on the expression of selected genes of the A549 lung cancer cells. Eurycomanone inhibited A549 lung cancer cell proliferation in a dose-dependent manner at concentrations ranging from 5 to 20 μg/ml. The concentration that inhibited 50% of cell growth (GI(50)) was 5.1 μg/ml. The anti-proliferative effects were not fully reversible following the removal of eurycomanone, in which 30% of cell inhibition still remained (p<0.0001, T-test). At 8 μg/ml (GI(70)), eurycomanone suppressed anchorage-independent growth of A549 cells by >25% (p<0.05, T-test, n=8) as determined using soft agar colony formation assay. Cisplatin, a chemotherapy drug used for the treatment of non small cell lung cancer on the other hand, inhibited A549 cells proliferation at concentrations ranging from 0.2 μg/ml to 15 μg/ml with a GI(50) of 0.58 μg/ml. The treatment with eurycomanone reduced the abundance expression of the lung cancer markers, heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1, p53 tumor suppressor protein and other cancer-associated genes including prohibitin (PHB), annexin 1 (ANX1) and endoplasmic reticulum protein 28 (ERp28) but not the house keeping genes. The mRNA expressions of all genes with the exception of PHB were significantly downregulated, 72 h after treatment (p<0.05, T-test, n=9). These findings suggest that eurycomanone at viable therapeutic concentrations of 5-20 μg/ml exhibited significant anti-proliferative and anti-clonogenic cell growth effects on A549 lung cancer cells. The treatment also resulted in suppression of the lung cancer cell tumor markers and several known cancer cell growth-associated genes.
The habitats of Eurycoma longifolia Jack, a slender tree, are jungles in Malaysia and Indonesia. It belongs to the family Simaroubaceae and is a source of quassinoids with anabolic, antimalarial and cytostatic activity. In this study, conducted during 2008 in Mae Sot, Thailand, a standardized extract of E. longifolia containing three major quassinoids, eurycomanone (1), 13,21-dihydroeurycomanone (2) and 13alpha(21)-epoxyeurycomanone (3) was evaluated for antiplasmodial activity against Plasmodium falciparum and its activity has been compared with that of artemisinin, using 38 fresh parasite isolates and assessment of inhibition of schizont maturation. The IC(50), IC(90) and IC(99) values for artemisinin were 4.30, 45.48 and 310.97 microg/l, and those for the root extract from E. longifolia 14.72, 139.65 and 874.15 microg/l respectively. The GMCOC for artemisinin was 337.81 mug/l, and for the plant extract it was 807.41 microg/l. The log-concentration probit regressions were parallel. The inhibitory activity of the E. longifolia extract was higher than that expected from the three quassinoids isolated from the plant, suggesting synergism between the quassinoids or the presence of other unidentified compounds.
Quassinoids are a group of diterpenoids found in plants from the Simaroubaceae family. They are also the major bioactive compounds found in Eurycoma longifolia which is commonly used as traditional medicine in South East Asia to treat various ailments including sexual dysfunction and infertility. These uses are attributed to its ability to improve testosterone level in men. Chronic consumption of E. longifolia extracts has been reported to increase testosterone level in men and animal model but its effect on prostate growth remains unknown. Therefore, the present study investigates the effects of a standardized total quassinoids composition (SQ40) containing 40% of the total quassinoids found in E. longifolia on LNCaP human prostate cancer cell line. SQ40 inhibited LNCaP cell growth at IC50 value of 5.97 μg/mL while the IC50 on RWPE-1 human prostate normal cells was 59.26 μg/mL. SQ40 also inhibited 5α-dihydrotestosterone-stimulated growth in LNCaP cells dose-dependently. The inhibitory effect of SQ40 in anchorage-independent growth of LNCaP cells was also demonstrated using soft agar assay. SQ40 suppressed LNCaP cell growth via G0/G1 phase arrest which was accompanied by the down-regulation of CDK4, CDK2, Cyclin D1 and Cyclin D3 and up-regulation of p21Waf1/Cip1 protein levels. SQ40 at higher concentrations or longer treatment duration can cause G2M growth arrest leading to apoptotic cell death as demonstrated by the detection of poly(ADP-ribose) polymerase cleavage in LNCaP cells. Moreover, SQ40 also inhibited androgen receptor translocation to nucleus which is important for the transactivation of its target gene, prostate-specific antigen (PSA) and resulted in a significant reduction of PSA secretion after the treatment. In addition, intraperitoneal injection of 5 and 10 mg/kg of SQ40 also significantly suppressed the LNCaP tumor growth on mouse xenograft model. Results from the present study suggest that the standardized total quassinoids composition from E. longifolia promotes anti-prostate cancer activities in LNCaP human prostate cancer cells.
The commercial oil palm (Elaeis guineensis Jacq.) produces a mesocarp oil (commonly called 'palm oil') with approximately equal proportions of saturated and unsaturated fatty acids (FAs). An increase in unsaturated FAs content or iodine value (IV) as a measure of the degree of unsaturation would help to open up new markets for the oil. One way to manipulate the fatty acid composition (FAC) in palm oil is through introgression of favourable alleles from the American oil palm, E. oleifera, which has a more unsaturated oil.
Evaluation of transcriptome data in combination with QTL information has been applied in many crops to study the expression of genes responsible for specific phenotypes. In oil palm, the mesocarp oil extracted from E. oleifera × E. guineensis interspecific hybrids is known to have lower palmitic acid (C16:0) content compared to pure African palms. The present study demonstrates the effectiveness of transcriptome data in revealing the expression profiles of genes in the fatty acid (FA) and triacylglycerol (TAG) biosynthesis processes in interspecific hybrids. The transcriptome assembly yielded 43,920 putative genes of which a large proportion were homologous to known genes in the public databases. Most of the genes encoding key enzymes involved in the FA and TAG synthesis pathways were identified. Of these, 27, including two candidate genes located within the QTL associated with C16:0 content, showed differential expression between developmental stages, populations and/or palms with contrasting C16:0 content. Further evaluation using quantitative real-time PCR revealed that differentially expressed patterns are generally consistent with those observed in the transcriptome data. Our results also suggest that different isoforms are likely to be responsible for some of the variation observed in FA composition of interspecific hybrids.
An extensive comparative study on the electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) mass spectrometry using automated flow injection analysis (FIA), was performed on eurycomanone (1), 13α(21)-epoxyeurycomanone (2), eurycomanol (3), eurycomanol-2-O-β-d-glucopyranoside (4), and 13,21-dihydroeurycomanone (5), the bioactive markers isolated from Eurycoma longifolia. The effects of eluent mixture (methanol or acetonitrile in water) and acidic modifiers (acetic acid, formic acid and trifluoroacetic acid) on the ionization efficiency of the markers were also investigated. The ESI in the positive ion mode with methanol containing 0.1% (v/v) acetic acid was selected for the subsequent optimization of nebulizer pressure, dry gas flow, dry gas temperature and capillary voltage to improve the sensitivity of the total ion chromatogram (TIC). Fragmentation of the analytes was further investigated by varying the capillary exit offset voltage and fragmentation amplitude in positive mode of ESI. The detection limits (LODs) were determined in isolation mode (selected ion monitoring, SIM). Their limits of detection (LODs) ranged between 0.03 and 0.1μgmL(-1) while the intra-day and inter-day precisions were less than 5.72% and 4.82%, respectively. The method was next applied for the simultaneous analysis of the markers to standardize various batches of manufactured extracts of E. longifolia for potential use as antimalarial products. Multiple Reaction Monitoring (MRM) mode was used for the quantification of analytes which gave protonated molecular ion, [M+H](+). For those without pseudo-molecular ions, SIM mode was used to quantify the analytes. The batches contained 5.65-9.95% of eurycomanone (1), 5.21-19.75% of eurycomanol (3) and 7.59-19.95% of eurycomanol-2-O-β-d-glucopyranoside (4) as major quassinoids whereas, 13α(21)-epoxyeurycomanone (2), and 13,21-dihydroeurycomanone (5) were much lower in concentrations of 0.78-3.90% and 0.47-1.76%, respectively.
13 α,21-Dihydroeurycomanone (1), a known quassinoid of Eurycoma longifolia Jack was recrystallized from chloroform into a novel crystal structure in space group P2 (1). Its X-ray data were compared with those of eurycomanone ( 2). Following intraperioneal injections at similar doses of 2.44 µmol/kg/day for 3 consecutive days, 2 displayed comparable potency with tamoxifen but was more potent than 1 in the anti-estrogenic effect against 17 α-ethynylestradiol (EE)-induced uterotrophy of immature rats.
A new and simple HPLC method using fluorescence detection was developed to determine 9-methoxycanthin-6-one, an active compound of Eurycoma longifolia Jack in rat and human plasma. The method entailed direct injection of plasma sample after deproteinization using acetonitrile. The mobile phase comprised acetonitrile and distilled water (55 : 45, v/v). Analysis was run at a flow rate of 1.0 ml/min with the detector operating at an excitation wavelength of 371 nm and emission wavelength of 504 nm. The method was specific and sensitive with a detection limit of 0.6 ng/ml and a quantification limit of approximately 1.6 ng/ml. The method was applied in a pilot pharmacokinetic/bioavailability study of the compound in rats. Less than 1 % of the compound was found to be absorbed orally.
Oil palm, a plantation crop of major economic importance in Southeast Asia, is the predominant source of edible oil worldwide. We report the identification of the virescens (VIR) gene, which controls fruit exocarp colour and is an indicator of ripeness. VIR is a R2R3-MYB transcription factor with homology to Lilium LhMYB12 and similarity to Arabidopsis production of anthocyanin pigment1 (PAP1). We identify five independent mutant alleles of VIR in over 400 accessions from sub-Saharan Africa that account for the dominant-negative virescens phenotype. Each mutation results in premature termination of the carboxy-terminal domain of VIR, resembling McClintock's C1-I allele in maize. The abundance of alleles likely reflects cultural practices, by which fruits were venerated for magical and medicinal properties. The identification of VIR will allow selection of the trait at the seed or early-nursery stage, 3-6 years before fruits are produced, greatly advancing introgression into elite breeding material.
Oil palm is the most productive oil-bearing crop. Although it is planted on only 5% of the total world vegetable oil acreage, palm oil accounts for 33% of vegetable oil and 45% of edible oil worldwide, but increased cultivation competes with dwindling rainforest reserves. We report the 1.8-gigabase (Gb) genome sequence of the African oil palm Elaeis guineensis, the predominant source of worldwide oil production. A total of 1.535 Gb of assembled sequence and transcriptome data from 30 tissue types were used to predict at least 34,802 genes, including oil biosynthesis genes and homologues of WRINKLED1 (WRI1), and other transcriptional regulators, which are highly expressed in the kernel. We also report the draft sequence of the South American oil palm Elaeis oleifera, which has the same number of chromosomes (2n = 32) and produces fertile interspecific hybrids with E. guineensis but seems to have diverged in the New World. Segmental duplications of chromosome arms define the palaeotetraploid origin of palm trees. The oil palm sequence enables the discovery of genes for important traits as well as somaclonal epigenetic alterations that restrict the use of clones in commercial plantings, and should therefore help to achieve sustainability for biofuels and edible oils, reducing the rainforest footprint of this tropical plantation crop.
A key event in the domestication and breeding of the oil palm Elaeis guineensis was loss of the thick coconut-like shell surrounding the kernel. Modern E. guineensis has three fruit forms, dura (thick-shelled), pisifera (shell-less) and tenera (thin-shelled), a hybrid between dura and pisifera. The pisifera palm is usually female-sterile. The tenera palm yields far more oil than dura, and is the basis for commercial palm oil production in all of southeast Asia. Here we describe the mapping and identification of the SHELL gene responsible for the different fruit forms. Using homozygosity mapping by sequencing, we found two independent mutations in the DNA-binding domain of a homologue of the MADS-box gene SEEDSTICK (STK, also known as AGAMOUS-LIKE 11), which controls ovule identity and seed development in Arabidopsis. The SHELL gene is responsible for the tenera phenotype in both cultivated and wild palms from sub-Saharan Africa, and our findings provide a genetic explanation for the single gene hybrid vigour (or heterosis) attributed to SHELL, via heterodimerization. This gene mutation explains the single most important economic trait in oil palm, and has implications for the competing interests of global edible oil production, biofuels and rainforest conservation.
Oil palm breeding involves crossing dura and pisifera palms to produce tenera progeny with greatly improved oil yield. Oil yield is controlled by variant alleles of a type II MADS-box gene, SHELL, that impact the presence and thickness of the endocarp, or shell, surrounding the fruit kernel. We identified six novel SHELL alleles in noncommercial African germplasm populations from the Malaysian Palm Oil Board. These populations provide extensive diversity to harness genetic, mechanistic and phenotypic variation associated with oil yield in a globally critical crop. We investigated phenotypes in heteroallelic combinations, as well as SHELL heterodimerization and subcellular localization by yeast two-hybrid, bimolecular fluorescence complementation and gene expression analyses. Four novel SHELL alleles were associated with fruit form phenotype. Candidate heterodimerization partners were identified, and interactions with EgSEP3 and subcellular localization were SHELL allele-specific. Our findings reveal allele-specific mechanisms by which variant SHELL alleles impact yield, as well as speculative insights into the potential role of SHELL in single-gene oil yield heterosis. Future field trials for combinability and introgression may further optimize yield and improve sustainability.
A set of Elaeis guineensis genes had been generated by combining two gene prediction pipelines: Fgenesh++ developed by Softberry and Seqping by the Malaysian Palm Oil Board. PalmXplore was developed to provide a scalable data repository and a user-friendly search engine system to efficiently store, manage and retrieve the oil palm gene sequences and annotations. Information deposited in PalmXplore includes predicted genes, their genomic coordinates, as well as the annotations derived from external databases, such as Pfam, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes. Information about genes related to important traits, such as those involved in fatty acid biosynthesis (FAB) and disease resistance, is also provided. The system offers Basic Local Alignment Search Tool homology search, where the results can be downloaded or visualized in the oil palm genome browser (MYPalmViewer). PalmXplore is regularly updated offering new features, improvements to genome annotation and new genomic sequences. The system is freely accessible at http://palmxplore.mpob.gov.my.
Comparative genomics and transcriptomic analyses were performed on two agronomically important groups of genes from oil palm versus other major crop species and the model organism, Arabidopsis thaliana. The first analysis was of two gene families with key roles in regulation of oil quality and in particular the accumulation of oleic acid, namely stearoyl ACP desaturases (SAD) and acyl-acyl carrier protein (ACP) thioesterases (FAT). In both cases, these were found to be large gene families with complex expression profiles across a wide range of tissue types and developmental stages. The detailed classification of the oil palm SAD and FAT genes has enabled the updating of the latest version of the oil palm gene model. The second analysis focused on disease resistance (R) genes in order to elucidate possible candidates for breeding of pathogen tolerance/resistance. Ortholog analysis showed that 141 out of the 210 putative oil palm R genes had homologs in banana and rice. These genes formed 37 clusters with 634 orthologous genes. Classification of the 141 oil palm R genes showed that the genes belong to the Kinase (7), CNL (95), MLO-like (8), RLK (3) and Others (28) categories. The CNL R genes formed eight clusters. Expression data for selected R genes also identified potential candidates for breeding of disease resistance traits. Furthermore, these findings can provide information about the species evolution as well as the identification of agronomically important genes in oil palm and other major crops.