A number of studies have pointed to the association of BDNF (brain-derived neurotrophic factor) and DARPP-32 (dopamine- and cAMP-regulated phosphoprotein, 32 kDa) with schizophrenia. The purpose of this study was to determine whether these two genes are involved in the pathogenesis of schizophrenia in the Malay population. Two single nucleotide polymorphisms Val66Met of BDNF, -2036C>G and g.1238delG of DARPP-32 were genotyped in the Malay population in 200 patients with schizophrenia and 256 healthy controls. Analysis of allele and genotype frequencies in these two groups revealed no significant association of BDNF or DARPP-32 polymorphisms with schizophrenia in Malays. This is the first such association study in the Malay population.
Ursodeoxycholic acid (UDCA) is used to treat liver diseases and demonstrates cardioprotective effects. Accumulation of the plasma membrane sphingolipid sphingomyelin in the heart can lead to atherosclerosis and coronary artery disease. Sphingomyelinases (SMases) break down sphingomyelin, producing ceramide, and inhibition of SMases activity can promote cell survival. We hypothesized that UDCA regulates activation of ERK and Akt survival signaling pathways and SMases in protecting cardiac cells against hypoxia. Neonatal cardiomyocytes were isolated from 0- to 2-day-old Sprague Dawley rats, and given 100 μM CoCl2, 150 μM H2O2, or placed in a hypoxia chamber for 24 h. The ameliorative effects of 100-μM UDCA treatment for 12 h were then assessed using MTS, QuantiGene Plex (for Smpd1 and Smpd2), and SMase assays, beating rate assessment, and western blotting (for ERK and Akt). Data were analyzed by the paired Student t-tests and one-way analyses of variance. Cell viability decreased significantly after H2O2 (85%), CoCl2 (50%), and hypoxia chamber (52%) treatments compared to the untreated control (100%). UDCA significantly counteracted the effects of chamber- and CoCl2- induced hypoxia on viability and beating rate. However, no significant differences were observed in acid SMase gene and protein expression between the untreated, CoCl2, and UDCA-CoCl2 groups. In contrast, neutral SMase gene and protein expression did significantly differ between the latter two groups. ERK and Akt phosphorylation was higher in hypoxic cardiomyocytes treated with UDCA than those given CoCl2 alone. In conclusion, UDCA regulates the activation of survival signaling proteins and SMases in neonatal rat cardiomyocytes during hypoxia.
The genus Curcuma is a member of the ginger family (Zingiberaceae) that has recently become popular for use as flowering pot plants, both indoors and as patio and landscape plants. We used PCR-based molecular markers (SSRs) to elucidate genetic variation and relationships between five varieties of Curcuma (Curcuma alismatifolia) cultivated in Malaysia. Of the primers tested, 8 (of 17) SSR primers were selected for their reproducibility and high rates of polymorphism. The number of presumed alleles revealed by the SSR analysis ranged from two to six alleles, with a mean value of 3.25 alleles per locus. The values of HO and HE ranged from 0 to 0.8 (mean value of 0.2) and 0.1837 to 0.7755 (mean value of 0.5102), respectively. Eight SSR primers yielded 26 total amplified fragments and revealed high rates of polymorphism among the varieties studied. The polymorphic information content varied from 0.26 to 0.73. Dice's similarity coefficient was calculated for all pairwise comparisons and used to construct an unweighted pair group method with arithmetic average (UPGMA) dendrogram. Similarity coefficient values from 0.2105 to 0.6667 (with an average of 0.4386) were found among the five varieties examined. A cluster analysis of data using a UPGMA algorithm divided the five varieties/hybrids into 2 groups.
The identification of new biomarkers for early detection of highly recurrent head and neck cancer is urgently needed. MicroRNAs (miRNAs) are small and non-coding RNAs that regulate cancer-related gene expression, such as tumor protein 53 (TP53) gene expression. This study was carried out to analyze TP53 gene expression using real-time PCR and to determine changes in intracellular p53 level by flow cytometry after downregulation of miRNA-181a miRNA inhibitor in the FaDu cell line. TP53 gene expression showed a 3-fold increment and the p53 protein level was also increased in the miRNA-181a-treated cells. In conclusion, miRNA-181a binds to the TP53 gene and inhibits its expression, decreasing the synthesis of p53.
Gastrointestinal cancers are malignant diseases with high mortality rate. Early diagnosis of patients could improve the results of treatment. Many studies used dermatoglyphics as a biomarker to predict the incidence of genetic diseases and cancers. This study assessed the association between gastrointestinal cancers and particular fingerprint patterns, which could be useful in early diagnosis of these malignancies. The study was conducted on 153 histopathologically confirmed gastrointestinal cancer patients and 299 healthy individuals. The fingerprints were taken by a specific method of rolling the subject's fingers or thumbs in ink. The data were analyzed for the significance using the chi-square test and the t-test. Odds ratio with 95% confidence intervals were calculated. Dermatoglyphic analysis showed that whorl and loop patterns significantly changed in the case group as compared to control. However, the odds ratio suggested that whorl pattern in 6 or more fingers might be a risk factor for developing gastrointestinal cancers. Our results showed that there is an association between fingerprint patterns and gastrointestinal cancers, and so, the dermatoglyphic analysis may aid in the early diagnosis of these cancers.
DNA extraction was carried out on 32 medicinal plant samples available in Malaysia using the TriOmic(TM) extraction kit. Amounts of 0.1 g flowers or young leaves were ground with liquid nitrogen, lysed at 65°C in RY1(plus) buffer and followed by RNAse treatment. Then, RY2 buffer was added to the samples and mixed completely by vortexing before removal of cell debris by centrifugation. Supernatants were transferred to fresh microcentrifuge tubes and 0.1 volume RY3 buffer was added to each of the transferred supernatant. The mixtures were applied to spin columns followed by a centrifugation step to remove buffers and other residues. Washing step was carried out twice by applying 70% ethanol to the spin columns. Genomic DNA of the samples was recovered by applying 50 μL TE buffer to the membrane of each spin column, followed by a centrifugation step at room temperature. A modification of the TriOmic(TM) extraction procedure was carried out by adding chloroform:isoamyl alcohol (24:1) steps in the extraction procedure. The genomic DNA extracted from most of the 32 samples showed an increase of total yield when chloroform:isoamyl alcohol (24:1) steps were applied in the TriOmicTM extraction procedure. This preliminary study is very important for molecular studies of medicinal plants available in Malaysia since the DNA extraction can be completed in a shorter period of time (within 1 h) compared to manual extraction, which entails applying phenol, chloroform and ethanol precipitation, and requires 1-2 days to complete.
Contamination of insect DNA for RAPD-PCR analysis can be a problem because many primers are non-specific and DNA from parasites or gut contents may be simultaneously extracted along with that of the insect. We measured the quantity of food ingested and assimilated by two sympatric populations of brown planthopper (BPH), Nilaparvata lugens, one from rice and the other from Leersia hexandra (Poaceae), a wetland forage grass, and we also investigated whether host plant DNA contaminates that of herbivore insects in extractions of whole insects. Ingestion and assimilation of food were reduced significantly when individuals derived from one host plant were caged on the other species. The bands, OPA3 (1.25), OPD3 (1.10), OPD3 (0.80), OPD3 (0.60), pUC/M13F (0.35), pUC/M13F (0.20), BOXAIR (0.50), peh#3 (0.50), and peh#3 (0.17) were found in both rice-infesting populations of brown planthopper and its host plant (rice). Similarly, the bands, OPA4 (1.00), OPB10 (0.70), OPD3 (0.90), OPD3 (0.80), OPD3 (0.60), pUC/ M13F (0.35), pUC/M13F (0.20), and BOXAIR (0.50) were found in both Leersia-infesting populations of brown planthopper and the host plant. So, it is clear that the DNA bands amplified in the host plants were also found in the extracts from the insects feeding on them.
Variable number of tandem repeats (VNTR) polymorphism in the interleukin 4 (IL-4) gene has been associated with end-stage renal disease (ESRD) subjects in many different populations, although with conflicting results. We determined the 70 bp of VNTR polymorphism at intron 3 of the IL-4 gene in Malaysian ESRD subjects. Buccal cells were collected from 160 case and 160 control subjects; genomic DNA was amplified using PCR, followed by agarose gel electrophoresis. There were significant differences in genotypes and alleles of the IL-4 gene. We conclude that VNTR polymorphism of the IL-4 gene is a risk factor for the development of ESRD among Malaysians.
Plants have been used throughout human history for food and medicine. However, many plants are toxic, and cannot easily be morphologically distinguished from non-toxic plants. DNA identification solves this problem and is widely used. Nonetheless, plant DNA barcode identification faces a number of challenges, and many studies have been conducted to find suitable barcodes. The present study was conducted to test the efficiency of commonly used primers, namely ITS2, rpoC1, and trnH-psbA, in order to find the best DNA barcode markers for the identification of medicinal plants in Malaysia. Fresh leaves from 12 medicinal plants that are commonly used by Malay traditional healers were collected from the Tropical Spice Garden, Pulau Pinang, and subjected to polymerase chain reaction amplification using ITS2, rpoC1, and trnH-psbA DNA markers. We found that trnH-psbA is the best DNA marker for the species-level identification of medicinal plants in Malaysia.
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.
A genome-wide association study showed that the tagging single nucleotide polymorphism (SNP) rs7566605 in the insulin-induced gene 2 (INSIG2) was associated with obesity. Attempts to replicate this result in different populations have produced inconsistent findings. We aimed to study the association between the rs7566605 SNP with obesity and other metabolic parameters in Malaysian Malays. Anthropometric and obesity-related metabolic parameters and DNA samples were collected. We genotyped the rs7566605 polymorphism in 672 subjects using real-time polymerase chain reaction. No significant associations were found between the rs7566605 tagging SNP of INSIG2 with obesity or other metabolic parameters in the Malaysian Malay population. The INSIG2 rs7566605 SNP may not play a role in the development of obesity-related metabolic traits in Malaysian Malays.
Carbamazepine (CBZ) is used as the first line of treatment of complex partial seizures (CPS) in Malaysia. While this drug is known to be effective for the treatment of CPS, more than 30% of patients remain drug resistant to CBZ mono-therapy. We examined a possible relationship between patients' response to CBZ mono-therapy and the G2677T SNP of the ABCB1 gene. Three hundred and fourteen patients with CPS were recruited from the Neurology Department of the Kuala Lumpur Hospital, of whom 152 were responders and the other 162 were non-responders to CBZ mono-therapy. DNA was extracted from blood samples and real-time PCR was performed to detect the G2677T SNP of the ABCB1 gene. Results were described as genotype frequencies and compared by logistic regression analysis. Among the 152 responders, 74% had the GG genotype. However, among the 162 non-responders, 26.5% had the GT genotype and 39% had the TT genotype. There was a significant difference in genotype frequency (TT vs GG; odds ratio 4.70; 95% confidence interval, 2.70-8.20) between responders and non-responders. The presence of the T allele of the G2677T SNP appears to be a useful screening marker to determine if a patient is going to be resistant to CBZ as a single drug therapy in the treatment of CPS.
Malaysia remains as a crossroad of different cultures and peoples, and it has long been recognized that studying its population history can provide crucial insight into the prehistory of Southeast Asia as a whole. The earliest inhabitants were the Orang Asli in Peninsular Malaysia and the indigenous groups in Sabah and Sarawak. Although they were the earliest migrants in this region, these tribes are divided geographically by the South China Sea. We analyzed DNA sequences of 18 Orang Asli using mitochondrial DNA extracted from blood samples, each representing one sub-tribe, and from five Sarawakian Iban. Mitochondrial DNA was extracted from hair samples in order to examine relationships with the main ethnic groups in Malaysia. The D-loop region and cytochrome b genes were used as the candidate loci. Phylogenetic relationships were investigated using maximum parsimony and neighbor joining algorithms, and each tree was subjected to bootstrap analysis with 1000 replicates. Analyses of the HVS I region showed that the Iban are not a distinct group from the Orang Asli; they form a sub-clade within the Orang Asli. Based on the cytochrome b gene, the Iban clustered with the Orang Asli in the same clade. We found evidence for considerable gene flow between Orang Asli and Iban. We concluded that the Orang Asli, Iban and the main ethnic groups of Malaysia are probably derived from a common ancestor. This is in agreement with a single-route migration theory, but it does not dismiss a two-route migration theory.
The Malayan gaur (Bos gaurus hubbacki) is one of the three subspecies of gaurs that can be found in Malaysia. We examined the phylogenetic relationships of this subspecies with other species of the genus Bos (B. javanicus, B. indicus, B. taurus, and B. grunniens). The sequence of a key gene, cytochrome b, was compared among 20 Bos species and the bongo antelope, used as an outgroup. Phylogenetic reconstruction was employed using neighbor joining and maximum parsimony in PAUP and Bayesian inference in MrBayes 3.1. All tree topologies indicated that the Malayan gaur is in its own monophyletic clade, distinct from other species of the genus Bos. We also found significant branching differences in the tree topologies between wild and domestic cattle.
PCR has been extensively used for amplification of DNA sequences. We conducted a study to obtain the best amplification conditions for cytochrome b (Cyt b), cytochrome c oxidase I (COI) and 12S rRNA (12S) gene fragments of Malayan gaur mtDNA. DNA from seven Malayan gaur samples were extracted for PCR amplification. Various trials and combinations were tested to determine the best conditions of PCR mixture and profile to obtain the best PCR products for sequencing purposes. Four selected target factors for enhancing PCR, annealing temperature, concentration of primer pairs, amount of Taq polymerase, and PCR cycle duration, were optimized by keeping the amount of DNA template (50 ng/μL) and concentration of PCR buffer (1X), MgCl(2) (2.5 mM) and dNTP mixture (200 μM each) constant. All genes were successfully amplified, giving the correct fragment lengths, as assigned for both forward and reverse primers. The optimal conditions were determined to be: 0.1 μM primers for Cyt b and COI, 0.3 μM primers for 12S, 1 U Taq polymerase for all genes, 30 s of both denaturation and annealing cycles for Cyt b, 1 min of both stages for 12S and COI and annealing temperature of 58.4 ° C for Cyt b, 56.1 ° C for 12S and 51.3 ° C for COI. PCR products obtained under these conditions produced excellent DNA sequences.
Little is known about the classification and phylogenetic relationships of the leaf monkeys (Presbytis). We analyzed mitochondrial DNA sequences of cytochrome b (Cyt b) and 12S rRNA to determine the phylogenetic relationships of the genus Presbytis. Gene fragments of 388 and 371 bp of Cyt b and 12S rRNA, respectively, were sequenced from samples of Presbytis melalophos (subspecies femoralis, siamensis, robinsoni, and chrysomelas), P. rubicunda and P. hosei. The genus Trachypithecus (Cercopithecidae) was used as an outgroup. The Cyt b NJ and MP phylogeny trees showed P. m. chrysomelas to be the most primitive, followed by P. hosei, whereas 12S rRNA tree topology only indicated that these two species have close relationships with the other members of the genus. In our analysis, chrysomelas, previously classified as a subspecies of P. melalophos, was not included in either the P. m. femoralis clade or the P. m. siamensis clade. Whether or not there should be a separation at the species level remains to be clarified. The tree topologies also showed that P. m. siamensis is paraphyletic with P. m. robinsoni, and P. m. femoralis with P. rubicunda, in two different clades. Cyt b and 12S rRNA are good gene candidates for the study of phylogenetic relationships at the species level. However, the systematic relationships of some subspecies in this genus remain unclear.
Twelve introgressed oil palm (Elaeis guineensis) progenies of Nigerian dura x Deli dura were evaluated for bunch yield, yield attributes, bunch quality components and vegetative characters at the Malaysian Palm Oil Board Research Station, in Keratong, Pahang, Malaysia. Analysis of variance revealed significant to highly significant genotypic differences, indicating sufficient genetic variability among the progenies for bunch yield and its attributes, vegetative characters and bunch quality components, except fruit to bunch ratio. Fresh fruit bunch yield ranged from 167 kg·palm(-1)·year(-1) in PK1330 to 212 kg·palm(-1)·year(-1) in PK1351, with a mean yield of 192 kg·palm(-1)·year(-1). Among the progeny, PK1313 had the highest oil to bunch ratio (19.36%), due to its high mesocarp to fruit ratio, fruit to bunch ratio and low shell to fruit ratio. Among the progenies, PK1313 produced the highest oil yield of 31.4 kg·palm(-1)·year(-1), due to a high mesocarp to fruit ratio (61.2%) and a low shell to fruit ratio (30.7%), coupled with high fruit to bunch ratio (65.6%). PK1330 was found promising for selection, as it had desirable vegetative characters, including smaller petiole cross section (27.15 cm2), short rachis length (4.83 m), short palm height (1.85 m), and the lowest leaf number (164.6), as these vegetative characters are prerequisites for selecting palms for high density planting and high yield per hectare. The genetic variability among the progenies was found to be high, indicating ample scope for further breeding, followed by selection.
Mitochondrial DNA cytochrome c oxidase II (COII) gene sequences of Malaysian Cercopithecidae were examined to ascertain their phylogenetic relationships. Colobinae were represented by the genera Presbytis, Trachypithecus and Nasalis, while the genus Macaca represented Cercopithecinae. DNA amplification and sequencing of the COII gene was performed on 16 samples. Symphalangus syndactylus (Hylobatidae) was used as the outgroup. Data were analyzed using both character (maximum parsimony) and distance (neighbor-joining) methods. Tree topologies indicated that Colobinae and Cercopithecinae have their own distinct monophyletic clade. This result was well supported by bootstrap values and genetic distances derived from the Kimura-2-parameter algorithm. Separation of Macaca nemestrina from M. fascicularis was also well supported by bootstrap values. In addition, tree topologies indicate a good resolution of the Colobinae phylogenetic relationships at the intergeneric level, but with low bootstrap support. The position of Nasalis remained problematic in both trees. Overall, COII is a good gene candidate for portraying the phylogenetic relationships of Malaysian primates at the inter- and intra-subfamily levels.
Molecular components of the dopamine D3 receptor (DRD3) may play an important role in the pathophysiology of schizophrenia. Previous studies have demonstrated an association between DRD3 Ser9Gly and cathechol-o-methyltransferase (COMT, SNP = rs165656) polymorphisms and schizophrenia but the results were inconclusive. We investigated this apparent association between Ser9Gly (A/G) polymorphism and an intronic SNP (dbSNP or rs165656) in 261 Malay patients diagnosed with schizophrenia and 216 controls, using PCR-RFLP. The genotype distribution of the polymorphism DRD3 Ser9Gly was in Hardy-Weinberg equilibrium (HWE) for patients (P = 0.1251) and out of HWE for controls (P = 0.0137). However, both healthy controls and schizophrenia patients were out of HWE for the polymorphism COMT rs165656. Based on allele and genotype frequencies in both groups, we found no significant association of DRD3 Ser9Gly polymorphisms and COMT (rs165656) with schizophrenia in Malays. Further studies should examine the association between other dopamine-related genes and the behavioral phenotypes of schizophrenia.
The serotoninergic system has been implicated in the etiology of schizophrenia and other behavioral disorders. Association studies have focused on the tryptophan hydroxylase 2 gene (TPH2) and the 5-hydroxytryptamine receptor 2A gene (5-HTR2A). We genotyped two single-nucleotide polymorphisms, A1438G of 5-HTR2A and intronic rs1386494 of TPH2 in the Malay population, using a sample size of 289 schizophrenic patients and 130 healthy controls. We found a significant association of A1438G of 5-HTR2A with schizophrenia in Malays. On the other hand, TPH2 polymorphism was not associated with schizophrenia. This is the first genetic association study concerning schizophrenia in the Malay population.