Starch accumulates in plants as granules in chloroplasts of source organs such as leaves (transitory starch) or in amyloplasts of sink organs such as seeds, tubers and roots (storage starch). Starch is composed of two types of glucose polymers: the essentially linear polymer amylose and highly branched amylopectin. The amylose content of wheat and rice seeds is an important quality trait, affecting the nutritional and sensory quality of two of the world's most important crops. In this review, we focus on the relationship between amylose biosynthesis and the structure, physical behaviour and functionality of wheat and rice grains. We briefly describe the structure and composition of starch and then in more detail describe what is known about the mechanism of amylose synthesis and how the amount of amylose in starch might be controlled. This more specifically includes analysis of GBSS alleles, the relationship between waxy allelic forms and amylose, and related quantitative trait loci. Finally, different methods for increasing or lowering amylose content are evaluated.
This study reports on the detection of additional expressed sequence tags (EST) derived simple sequence repeat (SSR) markers for the oil palm. A large collection of 19243 Elaeis guineensis ESTs were assembled to give 10258 unique sequences, of which 629 ESTs were found to contain 722 SSRs with a variety of motifs. Dinucleotide repeats formed the largest group (45.6%) consisting of 66.9% AG/CT, 21.9% AT/AT, 10.9% AC/GT and 0.3% CG/CG motifs. This was followed by trinucleotide repeats, which is the second most abundant repeat types (34.5%) consisting of AAG/CTT (23.3%), AGG/CCT (13.7%), CCG/CGG (11.2%), AAT/ATT (10.8%), AGC/GCT (10.0%), ACT/AGT (8.8%), ACG/CGT (7.6%), ACC/GGT (7.2%), AAC/GTT (3.6%) and AGT/ACT (3.6%) motifs. Primer pairs were designed for 405 unique EST-SSRs and 15 of these were used to genotype 105 E. guineensis and 30 E. oleifera accessions. Fourteen SSRs were polymorphic in at least one germplasm revealing a total of 101 alleles. The high percentage (78.0%) of alleles found to be specific for either E. guineensis or E. oleifera has increased the power for discriminating the two species. The estimates of genetic differentiation detected by EST-SSRs were compared to those reported previously. The transferability across palm taxa to two Cocos nucifera and six exotic palms is also presented. The polymerase chain reaction (PCR) products of three primer-pairs detected in E. guineensis, E. oleifera, C. nucifera and Jessinia bataua were cloned and sequenced. Sequence alignments showed mutations within the SSR site and the flanking regions. Phenetic analysis based on the sequence data revealed that C. nucifera is closer to oil palm compared to J. bataua; consistent with the taxanomic classification.
A total of 19 polymorphic microsatellite loci were used to analyse levels of genetic variation for 10 populations of Perna viridis L. collected from all over peninsular Malaysia. The populations involved in this study included Pulau Aman in Penang, Tanjung Rhu in Kedah, Bagan Tiang in Perak, Pulau Ketam in Selangor, Muar, Parit Jawa, Pantai Lido and Kampung Pasir Puteh in Johore, and Kuala Pontian and Nenasi in Pahang state. The number of alleles per locus ranged from two to seven, with an average of 3.1. Heterozygote deficiencies were observed across all the 10 populations. Characterization of the populations revealed that local populations of P. viridis in peninsular Malaysia were genetically similar enough to be used as a biomonitoring agent for heavy metal contamination in the Straits of Malacca. Cluster analysis grouped the P. viridis populations according to their geographical distributions with the exception of Parit Jawa. The analysis also revealed that P. viridis from the northern parts of peninsular Malaysia were found to be the most distant populations among the populations of mussels investigated and P. viridis from the eastern part of peninsular Malaysia were closer to the central and southern populations than to the northern populations.
The mating system and seed variation of Acacia hybrid (A. mangium x A. auriculiformis) were studied using allozymes and random amplified polymorphic DNA (RAPD) markers, respectively. Multi-locus outcrossing rate estimations indicated that the hybrid was predominantly outcrossed (mean+/- s.e. t(m) = 0.86+/-0.01). Seed variation was investigated using 35 polymorphic RAPD fragments. An analysis of molecular variance (AMOVA) revealed the highest genetic variation among seeds within a pod (66%-70%), followed by among pods within inflorescence (29%-37%), and the least variation among inflorescences within tree (1%). In addition, two to four RAPD profiles could be detected among seeds within pod. Therefore, the results suggest that a maximum of four seeds per pod could be sampled for the establishment of a mapping population for further studies.
The apical membrane antigen-1 (AMA-1) of Plasmodium spp. is a merozoite surface antigen that is essential for the recognition and invasion of erythrocytes. Polymorphisms occurring in this surface antigen will cause major obstacles in developing effective malaria vaccines based on AMA-1. The objective of this study was to characterize ama1 gene in Plasmodium knowlesi isolates from Sabah. DNA was extracted from blood samples collected from Keningau, Kota Kinabalu and Kudat. The Pkama1 gene was amplified using nested PCR and subjected to bidirectional sequencing. Analysis of DNA sequence revealed that most of the nucleotide polymorphisms were synonymous and concentrated in domain I of PkAMA-1. Forteen haplotypes were identified based on amino acid variations and haplotype K5 was the most common haplotype. dN/dS ratios implied that purifying selection was prevalent in Pkama1 gene. Fu and Li's D and F values further provided evidence of negative selection acting on domain II of Pkama1. Lownucleotide diversitywas also detected for the Pkama1 sequences,which is similar to reports on Pkama1 from Peninsular Malaysia and Sarawak. The presence of purifying selection and low nucleotide diversity indicated that domain II of Pkama1 can be used as a target for vaccine development.
Triple negative breast cancer (TNBC) is typically associated with poor and interindividual variability in treatment response. Cytochrome P450 family 1 subfamily B1 (CYP1B1) is a metabolizing enzyme, involved in the biotransformation of xenobiotics and anticancer drugs. We hypothesized that, single-nucleotide polymorphisms (SNPs), CYP1B1 142 C>G, 4326 C>G and 4360 A>G, and CYP1B1 mRNA expression might be potential biomarkers for prediction of treatment response in TNBC patients. CYP1B1 SNPs genotyping (76 TNBC patients) was performed using allele-specific polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism methods and mRNA expression of CYP1B1 (41 formalin-fixed paraffin embeddedblocks) was quantified using quantitative reverse transcription PCR. Homozygous variant genotype (GG) and variant allele (G) of CYP1B1 4326C>G polymorphism showed significantly higher risk for development of resistance to chemotherapy with adjusted odds ratio (OR): 6.802 and 3.010, respectively. Whereas, CYP1B1 142 CG heterozygous genotype showed significant association with goodtreatment response with adjusted OR: 0.199. CYP1B1 142C-4326G haplotype was associated with higher risk for chemoresistance with OR: 2.579. Expression analysis revealed that the relative expression of CYP1B1 was downregulated (0.592) in cancerous tissue compared with normal adjacent tissues. When analysed for association with chemotherapy response, CYP1B1 expression was found to be significantly upregulated (3.256) in cancerous tissues of patients who did not respond as opposed to those of patients who showed response to chemotherapy. Our findings suggest that SNPs together with mRNA expression of CYP1B1 may be useful biomarkers to predict chemotherapy response in TNBC patients.
The tiger frog Hoplobatrachus rugulosus (Wiegmann 1834) is a large robust dicroglossid frog widely distributed in southern China, Malaysia, Myanmar, Vietnam and Thailand. The escaped bred tiger frog introduced from Thailand hybridized with Chinese native population may have affected the genetic diversity of local Chinese tiger frogs. However, previous microsatellite loci of this species do not offer enough information to construct the genetic map. Here, we reported 33 new microsatellite loci from transcriptome sequencing for H. rugulosus. Alleles ranged between 1 and 10 per locus and only one locus (HRT001) was monomorphic. The polymorphic information content, observed and expected heterozygosity were 0-0.794, 0-0.969 and 0-0.831, respectively. None of the loci was observed in linkage disequilibrium and two loci (HRT023 and HRT068) deviated from Hardy-Weinberg equilibrium after Bonferroni correction for multiple tests. These transcriptome-derived microsatellite markers will be usedto study the genetic divergence and construct the genetic map in H. rugulosus.
Plasmodium knowlesi contributes to the majority of human malaria incidences in Malaysia. Its uncontrollable passage among the natural monkey hosts can potentially lead to zoonotic outbreaks. The merozoite of this parasite invades host erythrocytes through interaction between its erythrocyte-binding proteins (EBPs) and their respective receptor on the erythrocytes. The regionII of P. knowlesi EBP, P. knowlesi beta (PkβII) protein is found to be mediating merozoite invasion into monkey erythrocytes by interacting with sialic acid receptors. Hence, the objective of this study was to investigate the genetic diversity, natural selection and haplotype grouping of PkβII of P. knowlesi isolates in Malaysia. Polymerase chain reaction amplifications of PkβII were performed on archived blood samples from Malaysia and 64 PkβII sequences were obtained. Sequence analysis revealed length polymorphism, and its amino acids at critical residues indicate the ability of PkβII to mediate P. knowlesi invasion into monkey erythrocytes. Low genetic diversity (π = 0.007) was observed in the PkβII of Malaysia Borneo compared to Peninsular Malaysia (π = 0.015). The PkβII was found to be under strong purifying selection to retain infectivity in monkeys and it plays a limited role in the zoonotic potential of P. knowlesi. Its haplotypes could be clustered into Peninsular Malaysia and Malaysia Borneo groups, indicating the existence of two distinct P. knowlesi parasites in Malaysia as reported in an earlier study.
The detoxifying activity of glutathione S-transferases (GST) enzymes not only protect cells from the adverse effects of xenobiotics, but also alters the effectiveness of drugs in cancer cells, resulting in toxicity or drug resistance. In this study, we aimed to evaluate the association of GSTM1, GSTT1 and GSTP1 Ile105Val polymorphisms with treatment response among Malaysian chronic myeloid leukaemia (CML) patients who everyday undergo 400 mg of imatinib mesylate (IM) therapy. Multiplex polymerase chain reaction (multiplex-PCR) was performed to detect GSTM1 and GSTT1 polymorphisms simultaneously and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was conducted to detect the GSTP1 Ile195Val polymorphism. On evaluating the association of the variant genotype with treatment outcome, heterozygous variant (AG) and homozygous variant (GG) of GSTP1 Ile105Val showed significantly a higher risk for the development of resistance to IM with OR: 1.951 (95% CI: 1.186-3.209, P = 0.009) and OR: 3.540 (95% CI: 1.305-9.606, P = 0.013), respectively. Likewise, GSTT1 null genotype was also associated with a significantly higher risk for the development of resistance to IM with OR = 1.664 (95% CI: 1.011-2.739, P = 0.045). Our results indicate the potential usefulness of GST polymorphism genotyping in predicting the IM treatment response among CML patients.
Imatinib mesylate (IM), a well-established gold standard drug in the treatment of chronic myeloid leukaemia (CML), is a synthetic tyrosine kinase inhibitor. Despite excellent efficacy, a significant number of patients on IM therapy develop resistance to IM. Currently, great focus has been laid on the effect of interindividual pharmacogenetic variability on IM treatment responses. IM uptake is mediated by the hOCT1 protein encoded by the solute carrier 22 gene (SLC22A1). The current study investigated the impact of few single-nucleotide polymorphisms (SNPs) of SLC22A1 on mediating resistance and/or good response to IM among 278 Malaysian CML patients (146 IM-resistant group and 132 IM good response group) undergoing IM therapy on 400 mg daily. Our results showed that the allelic frequencies of heterozygous (CG) and homozygous variant (GG) genotypes of SLC22A1 C480G were significantly higher in the IM-resistant group compared with the IM good response group (41.8% versus 30.3% and 10.9% versus 4.5% with P values of 0.047 and 0.048, respectively). On evaluating the association of genotypes with risk of IM resistance development, heterozygous (CG) and homozygous (GG) variant genotypes showed significantly higher risk for developing resistance to IM treatment with odds ratio (OR): 1.901 (95% confidence interval (CI): 1.142-3.163, P = 0.013) and 3.324 (95% CI: 1.235-8.947, P = 0.017), respectively. Two SNPs and two insertions/deletions were detected in exon 7 of SLC22A1. For exon 7, 1222AA carriers together with the presence of both the 8-bp insertion and 3-bp deletion, and M420del alleles showed higher possibility of developing resistance towards IMtreatment. Our results warrant the need of genotyping this SNP in terms of modulating IM treatment in CML patients.
Msb069 primer pairs encompassed region is believed to be associated with a quantitative trait loci (QTL) of dorsal fin length in subgenus Poecilia. However, detailed investigation on Msb069 which originated from Xiphophorus on subgenus Poecilia remains unexplored. In this study, full sequence of Msb069 was characterized by sequencing bioinformatics analysis and gene expression. The sequence analysis of Msb069 primer pairs encompassed region on three species of Poecilia revealed higher number of microsatellite tandem repeats in Poecilia latipinna (ATG 16 ) compared to P. sphenops (ATG 13-14 ). There is no notable pattern of ATGtandem repeats discovered in the hybrids. The full sequence of Msb069 is 734 bp in length and showed a 233 bp conserved region between Xiphophorus and Poecilia. BLAST search performed on this sequence revealed no significant similarities. Nonquantitative RT-PCR exhibited the presence of Msb069 transcripts in three different tissues in subgenus Poecilia. Meanwhile, quantitative RTPCR expression on two different tissues showed relatively higher expression of Msb069 transcript in P. latipinna dorsal fin tissues in both male and female fishes, suggesting a repressive function of this transcript with respect to dorsal fin length. However the exact gene expression event of Msb069 is still unknown and requires further investigation.
The yellowtail rasbora (Rasbora tornieri) is a miniature ray-finned fish categorized under the genus Rasbora in the family of Cyprinidae. In this study, a complete mitogenome sequence of R. tornieri was sequenced using four primers targeting two halves of the mitogenome with overlapping flanking regions. The size of mitogenome was 16,573 bp, housing 22 transfer RNA genes, 13 protein-coding genes, two ribosomal RNA genes and a putative control region. Identical gene organization was detected between this species and other members of Rasbora genus. The heavy strand encompassed 28 genes while the light strand accommodated the other nine genes. Most protein-coding genes execute ATG as start codon, excluding COI and ND3 genes, which utilized GTG instead. The central conserved sequence blocks (CSB-E, CSB-F and CSB-D), variable sequence blocks (CSB-1, CSB-3 and CSB-2) as well as the terminal associated sequence (TAS) were conserved within the control region. The maximum likelihood phylogenetic family tree revealed the divergence of R. tornieri from the basal region of the Rasbora clade, where its evolutionary relationships with other Rasbora members are poorly resolved as indicated by the low bootstrap values. This work acts as window for further population genetics and molecular evolution studies of Rasbora genus in future.
The serotonin transporter 5-HTT is encoded by a single gene SLC6A4. Polymorphisms in SLC6A4 has been associated with a wide variety of neurological and psychiatric disorders including increased risk of posttraumatic stress disorder, higher likelihood for depression, obsessive-compulsive disorder (OCD), increased hostility and criminal behaviour. Genes associated with complex diseases often exhibit strong signatures of purifying selection compared to others. Further, discernible population specific variation in the signature of natural selection have been observed for several complex disease-related genes. In this project we aimed to investigate the population genetic variation of the serotonin transporter gene (SLC6A4), focussing on the single nucleotide polymorphisms (SNPs). To this end, we employed 2504 individuals around the globe available in 1000 Genome project Phase III data and classified them into five ethnic groups: Americans (AMR), Europeans (EUR), Africans (AFR), East Asians (EAS) and South Asians (SAS). Principal component analysis (PCA) performed on all annotated SNPs of SLC6A4 depicted clear clustering between Africans and the rest of the world along PC1, and East Asians and other non-African populations along PC2. Further, these SNPs were found to be under strong selection pressure especially among East Asian populations with significantly high positive cross-population extended haplotype homozygosity scores compared to Africans, indicating that SLC6A4 has likely undergone a strong selective sweep among the East Asians in the recent past. Our study can potentially explain the association between polymorphisms in SLC6A4, and major depression and suicidal tendencies among people of East Asian ancestry and the absence of such associations among people of European ancestry.