Seven (6.1%) of 115 strains of Salmonella typhi isolated from Malaysian patients harbored a single large plasmid of 71 to 166 mD. Two of the seven plasmid-bearing strains were resistant to chloramphenicol (Cm) and tetracycline (Tc) and they transferred Cm and Tc resistance traits to Escherichia coli K12 at frequencies from 1.6 x 10(-7) to 1.9 x 10(-6). Agarose gel electrophoresis provided evidence that the resistance traits were cotransferred on a conjugative plasmid. The significance and importance of these results are discussed.
Glucose phosphate isomerase of subperiodic Brugia malayi was studied by horizontal starch-gel electrophoresis. Two heterophenotypes, each represented by 3 bands of enzyme activity, were found among 38 parasites studied. This finding is attributed to the occurrence of 2 Gpi gene loci.
1. A total of 8 samples from three natural populations and a laboratory strain of Aedes albopictus were analysed for glycerol-3-phosphate dehydrogenase phenotypes by means of horizontal starch-gel electrophoresis. 2. The electrophoretic phenotypes were governed by three codominant Gpd alleles. 3. There was low variability, with the heterozygosity in the variable samples ranging from 0.02 to 0.12. 4. The commonest allele in all the population samples was GpdB which encoded an electrophoretic band with intermediate mobility. 5. There was no temporal or spatial variation.
The 'Dextran' and the 'Buffy-coat' methods for isolation of human leucocytes for DNA extraction were compared on the basis of DNA yield from the same amounts (10 ml) of blood. Human leucocytes from a total of 11 samples were isolated using both methods for each sample after which DNA was extracted. Extracted DNA samples were treated with ribonucleases and proteinase K after which the yields were quantitated by measuring absorbance at 260 nm. The 'Buffy-coat' method yielded a mean concentration of DNA of 476.7 micrograms/ml (range: 212 to 700 micrograms/ml) while the 'Dextran' method yielded 188.4 micrograms/ml (range: 64 to 340 micrograms/ml). The difference was confirmed by subjecting the extracted DNA samples to agarose gel electrophoresis.
1. Three natural populations and a laboratory strain of Aedes albopictus were analysed for glucose phosphate isomerase by means of horizontal starch-gel electrophoresis. 2. The electrophoretic phenotypes were governed by five codominant Gpi alleles. 3. The commonest allele in all the four population samples was GpiC which encoded an electrophoretic band with intermediate mobility. 4. The distributions of GPI phenotypes were in accordance with Hardy-Weinberg expectations. 5. The four population samples could be differentiated by the presence of a unique Gpi allele or the absence of a particular Gpi allele.
Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Purified preparations of Getah virus strains have been analysed by sodium-dodecyl-sulphate polyacrylamide gel electrophoresis (SDS-PAGE) to reveal their structural proteins. Two envelope proteins (E1 and E2) and core protein (C) were found with the prototype AMM2021 strain both under reducing and nonreducing conditions, while separation of E1 and E2 was observed only under nonreducing conditions for 3 strains isolated in Japan. Limited digestion by Staphylococcus aureus V8 protease revealed difference in the peptide patterns of E1 between AMM2021 and Japanese isolates. Mobility of E1 and E2 was slower for the virus grown in BHK21 cells compared with the virus grown in Aedes albopictus cells, indicating host-controlled modification on the envelope glycoproteins.
Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
A chiral separation method coupled with capillary electrophoresis (CE) analysis for ketoconazole and miconazole enantiomers using chiral selectors such as β-cyclodextrin (β-CD) and hydroxypropyl-β-CD (HP-β-CD) was developed in this study, which included the optimisation, validation and application of the method on the antifungal cream samples. The formation of inclusion complex between the hosts (β-CD and HP-β-CD) and guests (ketoconazole and miconazole) were compared and analysed using ultraviolet-visible spectrophotometry, nuclear magnetic resonance (NMR) spectroscopy and molecular docking methods. Results from the study showed that in a concentration that ranged between 0.25 and 50 mg L-1 , the linear calibration curves of each enantiomer had a high coefficient of regression (R2 > 0.999), low limit of detection (0.075 mg L-1 ) and low limit of quantification (0.25 mg L-1 ). The relative standard deviation (RSD) of the intraday and interday analyses ranged from 0.79% to 8.01% and 3.30% to 11.43%, respectively, while the recoveries ranged from 82.0% to 105.7% (RSD < 7%, n = 3). The most probable structure of the inclusion complexes was proposed based on the findings from the molecular docking studies conducted using the PatchDock server.
Malaysian Mahseer (Tor tambroides) is considered as a good prospect for aquaculture in Malaysia. However, knowledge about Malaysian Mahseer-associated sperm microbiota is still limited, although some studies reported that sperm-related bacteria are a factor in the decline of sperm quality, as sperm may become the carrier of pathogenic bacteria to the egg. The goal of this study was to evaluate the sperm microbiota associated with Malaysian Mahseer from 3 different locations (Universiti Malaysia Terengganu [UMT], Ajil, and Pahang) using polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE) fingerprinting and to compare location differences by cluster analysis. Our results showed that the UMT sample had different sperm microbiota composition and a different trend in its relationship with sperm quality. Correlation analysis showed a relationship between bacterial diversity and sperm quality. Phylogenetic analysis indicated that sperm microbiota was composed of diverse phyla, including Proteobacteria, Firmicutes, and Actinobacteria. Interestingly, bacteria such as Salinisphaera sp., Pelomonas sp., and Staphylococcus spp. were detected in all the locations, suggesting that these bacteria are indigenous bacterial members of the Malaysian Mahseer sperm microbiota, although their function is still unclear.
Matched MeSH terms: Denaturing Gradient Gel Electrophoresis
This study focused on the identification of pathogenic bacteria in raw water intake and after sand filtration for drinking water treatment plant during flood event in 2014. The samples was collected from the Lubok Buntar Water Treatment Plant (WTP) and processed through bacterial isolation using chocolate agar as a media. The isolation process conducted based on serial samples dilution and streaking method prior to DNA extraction. Deoxyribonucleic acid (DNA) extraction kit was used to get selected bacteria DNA and further analysis using Polymerase Chain Reaction (PCR) test and electrophoresis to get DNA sequences. The Basic Local Alignment Search Tool (BLAST) analysis was employed to identify the species of the isolated bacteria. As a result, Pantoeaagglomerans and Enterobacter sp. were found in raw and filtered water sample and indicating the same family types. It was concluded that bacteria of the same species were found before and after sand filtration and need to be removed by disinfectant process. The findings also indicated that all the physicochemical parameters measured were within the values prescribed by the Interim National Water Quality Standard (INWQS).
Oil palm is the major crop grown and cultivated in various Asian countries such as Malaysia, Indonesia and Thailand.
The core of oil palm trunk (COPT) consists of high sugar content, hence suitable for synthesis of fine chemicals and
biofuels. Increase of sugar content was reported previously during prolonged COPT storage. However, until now, there
has been no report on protein profiles during storage. Therefore, in this study, protein expression of the COPT during the
storage period of one to six weeks was investigated using sodium dodecyl sulphate polyacrylamide gel electrophoresis
(SDS-PAGE) coupled with optical density quantification and multivariate analyses for measuring differentially expressed
proteins. Accordingly, protein bands were subjected to tryptic digestion followed by tandem mass spectrometry (nanoLCMS/MS)
protein identification. The results from SDS-PAGE showed consistent protein bands appearing across the biological
replicates ranging from 10.455 to 202.92 kDa molecular weight (MW) regions. The findings from the principal component
analysis (PCA) plot illustrated the separation pattern of the proteins at weeks 4 and 5 of storage, which was influenced
mainly by the molecular weights of 14.283, 25.543, 29.757, 30.549, 31.511, 34.585 and 84.395 kDa, respectively. The
majority of these proteins are identified as those involved in stress- and defense-related, disease resistance, as well
as gene/protein expression processes. Indeed, these proteins were mostly upregulated during the later storage period
suggesting that long-term storage may influence the molecular regulation of COPT sap.
Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Protein degradation can occur through Ubiquitin 26S-Proteosome System (UPS). The degradation can be mediated by
the SCF E3 ubiquitin ligase complex consisting of Skp1, Cullin, and F-box protein as the main components. The F-box
protein at the C-terminal domain functions to recognize the targeted protein to be ubiquitinated and degraded via UPS.
A stress-responsive F-box gene, PmF-box1 from Persicaria minor was categorized in the F-box containing kelch repeat
(FBK) family; a family that specific to plant kingdom. To identify the targeted protein of PmF-box1, yeast-two hybrid system
(Y2H) was used. In the Y2H screening process, mating efficiency is very important to fish out the interacting proteins.
Therefore, one modification was conducted to increase the mating efficiency. In this screening, PmF-box1 was used as a
bait to screen for the Y2H library which was constructed using RNA from plant samples treated with abscisic acid (ABA)
and polyethylene glycol (PEG)-8000 and control sample. Autoactivation and toxicity tests of bait were performed before
the Y2H screening. Tests on PmF-box1 showed that it is not toxic to the yeast and cannot autoactivate the yeast reporter
genes. Mating efficiency was improved from 2.07% to 9.15% after addition of PEG-4000 in the mating culture compared
to the original protocol, which it also increased the colony number in the screening step afterward. Additionally, bands
of gene with different sizes were observed on electrophoresis gel after colony PCR analysis from the improved technique.
Those genes may code for potential interacting proteins that needs further identification and confirmation.
The amplification of the highly conserved chromo-helicase-DNA binding region found in both the Z and W chromosome was evaluated with three sets of primers (P8/P2, 1237L/1272H and 2550F/2718R). DNA extracted from feathers through a simple boiling method was used to address its reliability in generating the sex-linked bands. All the bird samples, including the seven bird families that have not been reported previously, were successfully amplified with the primer set 2550F/2718R. The resulting polymerase chain reaction products showed clearly resolved fragments on a conventional agarose gel electrophoresis with size differences ranging from 80 to 540 bp between the two respective ZW gene copies. Although the P8/P2 primer was not as effective under the same conditions, it was able to produce well-resolved Z and W bands from bird species under the Antidea family, whereas the 2250F/2718R primer set only produced a single amplified fragment of a different size between the male and the female. Zoo Biol 27:62-69, 2008. (c) 2007 Wiley-Liss, Inc.
Introduction: Drug-resistance is a major hindrance to successful treatment of AML. Current predictive biomarkers are mainly genetic aberrations and insufficient in foretelling treatment outcome in all acute myeloid leukaemia (AML) due to its heterogeneous and aggressive nature. Proteins are stable and reliable. Secreted proteins in AML may have predictive or prognostic values for early intervention. Proteomic studies on AML are few and further investigations will benefit in selection of best markers. The aim of the study was to identify differentially expressed plasma proteins in AML with different treatment outcome. Methods: Two-dimensional electrophoresis (2-DE) technique was utilised to identify proteins differentially expressed in chemo-sensitive/chemo-resistant AML. Plasma and peripheral blood mononuclear cell (PBMC) lysate proteome analysis were performed on six chemo-resistant, four chemo-sensitive and six healthy controls and seven chemo-resistant, three chemo-sensitive and six healthy controls, respectively. Each experiment was conducted in duplicate or triplicate. Images were captured and protein spots detected by software. Differentially expressed protein spots were excised from gel and proteins were identified using LC/MS/MS. Proteins spots that were also detected in healthy controls were excluded. Results: Comparing mean % volume of each spot demonstrated significantly enhanced expression of apoliprotein-E (APO-E) and haptoglobin (HP) (p
Background: The low yield and quality of buccal-derived genomic DNA have reduced its applicability in various genetic research. The aim of this study was to assess the quantity, purity and genotyping efficiency of genomic DNA isolated from neonatal buccal swabs. Methods: Paired buccal swabs and whole blood samples were collected from 60 neonates with the mean age 5 days (SD=1.57). The genomic DNA quantity and purity were measured by using Infinite® 200 PRO NanoQuant reader and agarose gel electrophoresis. High-resolution melting (HRM) analysis was used to analyse the sequence variants present in uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1 c.211G>A) and nuclear receptor subfamily 1, group I, member 3 (NR1I3 IVS8+116T>G) genes. Results: Buccal swabs provided lower mean genomic DNA concentration (18.78 ± 8.39 ng/μl versus 40.02 ± 13.03 ng/μl), yield (2.63 ± 1.17 μgversus8.00 ± 2.61 μg). The purity of buccal samples however were inconsistent with 16 samples (26.7%) having A260/280 ratios below 1.8 which indicated protein contamination. Genomic DNA purity for all blood samples were within the ideal range with average absorbance ratios of 1.8−2.0. However, all buccal genomic DNA demonstrated 100% genotype call rates for all variants. A complete genotype concordance was also observed between paired genomic DNA samples. Conclusion: Despite related to a reduced quantity and purity, neonatal buccal genomic DNA could generate reliable HRM genotyping results. Therefore, buccal swab collection is a promising alternative to the invasive blood sampling to provide genomic DNA for genetic analysis involving paediatric population.
Introduction: Dengue is caused by dengue virus (DENV) which is a member of the genus Flavivirus of the family Flaviviridae. The prevalence of dengue has been increasing all over the world especially in Southeast Asia and Western Pacific regions. In 2016 - 2017 dengue outbreaks were reported in Sandakan and Kudat of Sabah, Malay-sia. The aim of this study was to determine the serotypes of dengue viruses circulating in these two sites during the outbreaks. Methods: A total of 200 dengue patients’ sera tested positive with NS1 and IgM & IgG rapid test (PanBio) were collected from Hospital Duchess of Kent Sandakan and Hospital Kudat between June 2016 and December 2017. PCR was done at the Faculty of Medicine and Health Sciences, Universiti Malaysia Sabah. One-Step Reverse transcriptase PCR (RT-PCR) and nested PCR was performed using C-prM amplimers designed by Lanciotti et al and later redesigned by Chien et al, followed by sequencing some of the PCR products. Results: Out of 200 sera tested 128 were PCR positive. All the four dengue serotypes were detected with PCR products with specific sizes in gel electrophoresis. However, in four samples, no serotype-specific band was amplified by the nested PCR, while they were dengue-positive in RT-PCR showing 511 base pair amplicon. Sequencing results revealed all four samples were found to belong to DENV4. The sequences of these samples were aligned with that of DENV 4 reverse primer rTS4. The DENV4 specific primer rTS4 was found to have four mismatched nucleotides to the DENV4 sequences. Conclusion: There was a co-circulation of DENV1 to 4 in Sandakan and Kudat in the study period. DENV1 was the predominant serotype. DENV4 specific C-prM primer rTS4 should be redesigned for the local DENV4 strain in Sabah in future research.
Haemoglobin Bart's hydrops fetalis syndrome is totally lethal. Globin chain electrophoresis on mylar backed cellulose acetate strips, by a method modified from Ueda and Schneider has been established to demonstrate total absence of alpha chains in this syndrome. This simple test can identify fetuses, stillbirths and newborns with homozygous αo-thalassaemia. In this region where DNA studies are limited, and prenatal diagnosis is unavailable, this test which describes the phenotypic expression of Hb Bart's syndrome will improve genetic counselling of women at risk of homozygous αo-thalassaemia.
Arterial/venous thrombosis is the major cardiovascular disorder accountable for substantial mortality; and the current demand for antithrombotic agents is extensive. Heparinases depolymerize unfractionated heparin (UFH) for the production of low molecular-weight heparins (LMWHs; used as anticoagulants against thrombosis). A microbial strain of Streptomyces sp. showing antithrombotic activity was isolated from the soil sample collected from north India. The strain was characterized by using 16S rRNA homology technique and identified as Streptomyces variabilis MTCC 12266 capable of producing heparinase enzyme. This is the very first communication reporting Streptomyces genus as the producer of heparinase. It was observed that the production of intracellular heparinase was [63.8 U/mg protein (specific activity)] 1.58 folds higher compared to extracellular heparinase [40.28 U/mg protein]. DEAE-Sephadex A-50 column followed by Sepharose-6B column purification of the crude protein resulted 19.18 folds purified heparinase. SDS-PAGE analysis of heparinase resulted an estimated molecular-weight of 42 kDa. It was also found that intracellular heparinase has the ability to depolymerize heparin to generate LMWHs. Further studies related to the mechanistic action, structural details, and genomics involved in heparinase production from Streptomyces variabilis are warranted for large scale production/purification optimization of heparinase for antithrombotic applications.
Matched MeSH terms: Electrophoresis, Polyacrylamide Gel