Displaying publications 41 - 60 of 1868 in total

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  1. A simple and rapid genotyping method for beta-2 receptor (beta2 AR) gene using allele specific multiplex PCR
    Romaino SM, Teh LK, Zilfalil BA, Thong CP, Ismail AA, Amir J, et al.
    J Clin Pharm Ther, 2004 Feb;29(1):47-52.
    PMID: 14748897 DOI: 10.1046/j.1365-2710.2003.00535.x
    Polymorphism of the beta2-adrenergic receptor (beta2 AR) gene is an important determinant of the function of this receptor. It affects receptor down-regulation and beta2-agonist responses. It has also been a focus of interest in attempts to elucidate the genetic basis of asthma, hypertension, obesity and cystic fibrosis. Several different techniques have been established to determine beta2 AR genotypes but none of these methods are simple enough to detect simultaneously all the five alleles of our research interest (Arg16/Gly16, -20T/C, Gln27/Glu27, -47T/C and Thr164/Ile164).
    Matched MeSH terms: Polymerase Chain Reaction/methods
  2. Fulltext Combined PCR and MAT improves the early diagnosis of the biphasic illness leptospirosis
    Philip N, Affendy NB, Masri SN, Yuhana MY, Than LTL, Sekawi Z, et al.
    PLoS One, 2020;15(9):e0239069.
    PMID: 32915919 DOI: 10.1371/journal.pone.0239069
    The diagnosis of leptospirosis remains a challenge due to its non-specific symptoms and the biphasic nature of the illness. A comprehensive diagnosis that includes both molecular (polymerase chain reaction (PCR)) and serology is vital for early detection of leptospirosis and to avoid misdiagnosis. However, not all samples could be subjected to both tests (serology and molecular) due to budget limitation, infrastructure, and technical expertise at least in resource-limited countries. We evaluated the usefulness of testing the clinically suspected leptospirosis cases with both techniques on all samples collected from the patients on the day of admission. Among the 165 patient's blood/serum samples tested (from three hospitals in Central Malaysia), 43 (26%) showed positivity by microscopic agglutination test (MAT), 63 (38%) by PCR, while 14 (8%) were positive by both MAT and PCR. For PCR, we tested two molecular targets (lipL32 by qPCR and 16S rDNA or rrs by nested PCR) and detected lipL32 in 47 (29%) and rrs gene in 63 (38%) patients. The use of more than one target gene for PCR increased the detection rates. Hence, a highly sensitive multiplex PCR targeting more than one diagnostic marker is recommended for the early detection of Leptospira in suspected patients. When the frequencies for positivity detected either by MAT or PCR combined, leptospirosis was diagnosed in a total of 92 (56%) patients, a higher frequency compared to when samples were only tested by a single method (MAT or PCR). The results from this study suggest the inclusion of both serology and molecular methods for every first sample irrespective of the days post-onset of symptoms (DPO) collected from patients for early diagnosis of leptospirosis.
    Matched MeSH terms: Multiplex Polymerase Chain Reaction; Real-Time Polymerase Chain Reaction
  3. Isolation by the 5' anchored PCR technique and characterization of eighteen microsatellite loci in horseshoe crab (Tachypleus gigas)
    Ling LP, Adibah AB, Tan SG, Christianus A, Faridah QZ
    J Genet, 2011 Dec;90(3):e101-4.
    PMID: 22232191
    Matched MeSH terms: Polymerase Chain Reaction/methods
  4. Development of single locus DNA microsatellite markers in Oryctes rhinoceros (Linnaeus) using 5' anchored RAMs-PCR method
    Manjeri G, Muhamad R, Faridah QZ, Tan SG
    J Genet, 2012 Nov 22;91(3):e92-6.
    PMID: 23257301
    Matched MeSH terms: Polymerase Chain Reaction/methods*
  5. Fulltext Detection of all four dengue virus serotypes and identification of four mismatched nucleotides between DENV4 specific primer rTS4 and DENV 4 sequences in Sandakan and Kudat (2016-2017)
    Tin Sabai Aung, Amalina Emran, Chua Tock Hing, Tin Tin Thein, Win Win Than, Aye Aye Wynn, et al.
    Malaysian Journal of Medicine and Health Sciences, 2019;15(106):49-49.
    MyJurnal
    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.
    Matched MeSH terms: Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction
  6. Fulltext Quantitative analysis of the expression of p53 gene in colorectal carcinoma by using real-time PCR
    Bong I, Lim P, Balraj P, Sim Ui Hang E, Zakaria Z
    Trop Biomed, 2006 Jun;23(1):53-9.
    PMID: 17041552 MyJurnal
    Colorectal carcinoma ranks third among ten leading causes of cancer in Malaysia. The colorectal carcinoma tumourigenesis involves the inactivation of tumour suppressor genes, and activation of proto-oncogenes. The p53 is one of the tumour suppressor genes that is involved in the colorectal carcinogenesis. The p53 gene is located on human chromosome 17p13.1 and comprises of 11 exons. Deficiencies in the p53 gene can cause the cancerous cells to spread to distant organs such as liver, lungs, lymph nodes, spine and bone. The most common p53 abnormalities that can lead to the metastasis of colorectal tumours are mutation and deregulation of the gene. In this study, nine colorectal carcinoma samples were used to establish a simple and sensitive strategy in the study on in vivo p53 expression by using realtime LightCycler SYBR Green I technology.
    Matched MeSH terms: Polymerase Chain Reaction/methods*
  7. Diagnostic accuracy of the Xpert® MTB/RIF cycle threshold level to predict smear positivity: a meta-analysis
    Lange B, Khan P, Kalmambetova G, Al-Darraji HA, Alland D, Antonenka U, et al.
    Int J Tuberc Lung Dis, 2017 05 01;21(5):493-502.
    PMID: 28399963 DOI: 10.5588/ijtld.16.0702
    SETTING: Xpert® MTB/RIF is the most widely used molecular assay for rapid diagnosis of tuberculosis (TB). The number of polymerase chain reaction cycles after which detectable product is generated (cycle threshold value, CT) correlates with the bacillary burden.OBJECTIVE To investigate the association between Xpert CT values and smear status through a systematic review and individual-level data meta-analysis.

    DESIGN: Studies on the association between CT values and smear status were included in a descriptive systematic review. Authors of studies including smear, culture and Xpert results were asked for individual-level data, and receiver operating characteristic curves were calculated.

    RESULTS: Of 918 citations, 10 were included in the descriptive systematic review. Fifteen data sets from studies potentially relevant for individual-level data meta-analysis provided individual-level data (7511 samples from 4447 patients); 1212 patients had positive Xpert results for at least one respiratory sample (1859 samples overall). ROC analysis revealed an area under the curve (AUC) of 0.85 (95%CI 0.82-0.87). Cut-off CT values of 27.7 and 31.8 yielded sensitivities of 85% (95%CI 83-87) and 95% (95%CI 94-96) and specificities of 67% (95%CI 66-77) and 35% (95%CI 30-41) for smear-positive samples.

    CONCLUSION: Xpert CT values and smear status were strongly associated. However, diagnostic accuracy at set cut-off CT values of 27.7 or 31.8 would not replace smear microscopy. How CT values compare with smear microscopy in predicting infectiousness remains to be seen.

    Matched MeSH terms: Polymerase Chain Reaction/methods*
  8. Fulltext A duplex endpoint PCR assay for rapid detection and differentiation of Leptospira strains
    Benacer D, Zain SNM, Lewis JW, Khalid MKNM, Thong KL
    Rev Soc Bras Med Trop, 2017 Mar-Apr;50(2):239-242.
    PMID: 28562762 DOI: 10.1590/0037-8682-0364-2016
    INTRODUCTION:: This study aimed to develop a duplex endpoint PCR assay for rapid detection and differentiation of Leptospira strains.

    METHODS:: Primers were designed to target the rrs (LG1/LG2) and ligB (LP1/LP2) genes to confirm the presence of the Leptospira genus and the pathogenic species, respectively.

    RESULTS:: The assay showed 100% specificity against 17 Leptospira strains with a limit of detection of 23.1pg/µl of leptospiral DNA and sensitivity of 103 leptospires/ml in both spiked urine and water.

    CONCLUSIONS:: Our duplex endpoint PCR assay is suitable for rapid early detection of Leptospira with high sensitivity and specificity.
    Matched MeSH terms: Polymerase Chain Reaction/methods*
  9. Fulltext HLA-B*15:02 screening in epileptic patients using a high resolution melting-real time PCR (HRM-QPCR) method
    Zam Zureena Mohd Rani, Nor Azian Abdul Murad, Saberi Saimun, Sri Noraima Othman, Rahman Jamal, Sue-Mian Then, et al.
    Neurology Asia, 2018;23(2):137-144.
    MyJurnal
    Background: The HLA-B*15:02 polymorphism in epileptic patients is known to be associated with carbamazepine-induced Stevens-Johnson syndrome (SJS). The prevalence of HLA-B*15:02 polymorphism seemed to be ethnic-specific with a higher frequency of HLA-B*15:02 in Asian compared to the Europeans. This study was performed to determine the frequency of the HLA-B*15:02 polymorphism in epileptic patients at the Chancellor Tuanku Muhriz Hospital-UKM Medical Centre (HCTM-UKMMC) using high resolution melting-real time PCR (HRM-QPCR) method.
    Methods: We performed a fast and effective in-house high resolution melting-real time polymerase chain reaction method and compared it with the conventional multiplex-PCR method. The specificity and sensitivity of each test were also determined using DNA from saliva.
    Results: Using the conventional multiplexPCR approach for screening, 25 out of 64 (39.1%) epileptic patients were positive for HLA-B*15:02. However, using the HRM-QPCR technique, 24/64 (37.5%) of the patients were positive. The one patient who tested positive by the multiplex-PCR but negative using the HRM-QPCR turned out to be negative by DNA sequencing. The HRM-QPCR and DNA sequencing showed 100% sensitivity and specificity. The multiplex-PCR showed 100% sensitivity and 98.4% specificity compared to both HRM-QPCR and DNA sequencing. The HRM-QPCR is also more cost-effective (chain reaction, high resolution melting-real time polymerase chain reaction (HRM-QPCR), DNA sequencing
    Matched MeSH terms: Multiplex Polymerase Chain Reaction; Real-Time Polymerase Chain Reaction
  10. Fulltext A simple and inexpensive physical lysis method for DNA and RNA extraction from freshwater microalgae
    Yee W, Abdul-Kadir R, Lee LM, Koh B, Lee YS, Chan HY
    3 Biotech, 2018 Aug;8(8):354.
    PMID: 30105179 DOI: 10.1007/s13205-018-1381-1
    In this work, a simple and inexpensive physical lysis method using a cordless drill fitted with a plastic pellet pestle and 150 mg of sterile sea sand was established for the extraction of DNA from six strains of freshwater microalgae. This lysis method was also tested for RNA extraction from two microalgal strains. Lysis duration between 15 and 120 s using the cetyltrimethyl ammonium bromide (CTAB) buffer significantly increased the yield of DNA from four microalgalstrains (Monoraphidium griffithii NS16, Scenedesmus sp. NS6, Scenedesmus sp. DPBC1 and Acutodesmus sp. DPBB10) compared to control. It was also found that grinding was not required to obtain DNA from two strains of microalgae (Choricystis sp. NPA14 and Chlamydomonas sp. BM3). The average DNA yield obtained using this lysis method was between 62.5 and 78.9 ng/mg for M. griffithii NS16, 42.2-247.0 ng/mg for Scenedesmus sp. NS6, 70.2-110.9 ng/mg for Scenedesmus sp. DPBC1 and 142.8-164.8 ng/mg for Acutodesmus sp. DPBB10. DNA obtained using this method was sufficiently pure for PCR amplification. Extraction of total RNA from M. griffithii NS16 and Mychonastes sp. NPD7 using this lysis method yielded high-quality RNA suitable for RT-PCR. This lysis method is simple, cheap and would enable rapid nucleic acid extraction from freshwater microalgae without requiring costly materials and equipment such as liquid nitrogen or beadbeaters, and would facilitate molecular studies on microalgae in general.
    Matched MeSH terms: Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction
  11. Fulltext Real-time reverse transcription loop-mediated isothermal amplification for rapid detection of SARS-CoV-2
    Lau YL, Ismail I, Mustapa NI, Lai MY, Tuan Soh TS, Hassan A, et al.
    PeerJ, 2020;8:e9278.
    PMID: 32547882 DOI: 10.7717/peerj.9278
    Background: Highly sensitive real-time reverse transcription polymerase chain reaction (RT-qPCR) methods have been developed for the detection of SARS-CoV-2. However, they are costly. Loop-mediated isothermal amplification (LAMP) assay has emerged as a novel alternative isothermal amplification method for the detection of nucleic acid.

    Methods: A rapid, sensitive and specific real-time reverse transcription LAMP (RT-LAMP) assay was developed for SARS-CoV-2 detection.

    Results: This assay detected one copy/reaction of SARS-CoV-2 RNA in 30 min. Both the clinical sensitivity and specificity of this assay were 100%. The RT-LAMP showed comparable performance with RT-qPCR. Combining simplicity and cost-effectiveness, this assay is therefore recommended for use in resource resource-limited settings.

    Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction; Real-Time Polymerase Chain Reaction
  12. Long Fragment Polymerase Chain Reaction
    Chua EW, Maggo S, Kennedy MA
    Methods Mol Biol, 2017;1620:65-74.
    PMID: 28540699 DOI: 10.1007/978-1-4939-7060-5_3
    Polymerase chain reaction (PCR) is an oft-used preparatory technique in amplifying specific DNA regions for downstream analysis. The size of an amplicon was initially limited by errors in nucleotide polymerization and template deterioration during thermal cycling. A variant of PCR, designated long-range PCR, was devised to counter these drawbacks and enable the amplification of large fragments exceeding a few kb. In this chapter we describe a protocol for long-range PCR, which we have adopted to obtain products of 6.6, 7.2, 13, and 20 kb from human genomic DNA samples.
    Matched MeSH terms: Polymerase Chain Reaction/methods*
  13. Fulltext Performance Evaluation of Commercial Dengue Diagnostic Tests for Early Detection of Dengue in Clinical Samples
    Mat Jusoh TNA, Shueb RH
    J Trop Med, 2017;2017:4687182.
    PMID: 29379526 DOI: 10.1155/2017/4687182
    The shattering rise in dengue virus infections globally has created a need for an accurate and validated rapid diagnostic test for this virus. Rapid diagnostic test (RDT) and reverse transcription-polymerase chain reaction (RT-PCR) diagnostic detection are useful tools for diagnosis of early dengue infection. We prospectively evaluated the diagnostic performance of nonstructural 1 (NS1) RDT and real-time RT-PCR diagnostic kits in 86 patient serum samples. Thirty-six samples were positive for dengue NS1 antigen while the remaining 50 were negative when tested with enzyme-linked immunosorbent assay (ELISA). Commercially available RDTs for NS1 detection, RTK ProDetect™, and SD Bioline showed high sensitivity of 94% and 89%, respectively, compared with ELISA. GenoAmp® Trioplex Real-Time RT-PCR and RealStar® Dengue RT-PCR tests presented a comparable kappa agreement with 0.722. The result obtained from GenoAmp® Real-Time RT-PCR Dengue test showed that 14 samples harbored dengue virus type 1 (DENV-1), 8 samples harbored DENV-2, 2 samples harbored DENV-3, and 1 sample harbored DENV-4. 1 sample had a double infection with DENV-1 and DENV-2. The NS1 RDTs and real-time RT-PCR tests were found to be a useful diagnostic for early and rapid diagnosis of acute dengue and an excellent surveillance tool in our battle against dengue.
    Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction; Real-Time Polymerase Chain Reaction
  14. Development of a TaqMan minor groove binding probe-based quantitative reverse transcription polymerase chain reaction for the detection and quantification of Zika virus
    Chin KL, Teoh BT, Sam SS, Loong SK, Tan KK, Azizan NS, et al.
    Trop Biomed, 2022 Dec 01;39(4):518-523.
    PMID: 36602210 DOI: 10.47665/tb.39.4.005
    Zika virus (ZIKV) infection has emerged as a global health concern following epidemic outbreaks of severe neurological disorders reported in Pacific and Americas since 2016. Therefore, a rapid, sensitive and specific diagnostic test for ZIKV infection is critical for the appropriate patient management and the control of disease spread. A TaqMan minor groove binding (MGB) probe-based quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assay was developed based on the conserved sequence regions of 463 ZIKV NS2B genes. The designed ZIKV qRT-PCR assay was evaluated for its detection limit, strain coverage and cross-reactivity. We further assessed the clinical applicability of qRT-PCR assay for ZIKV RNA detection using a total 18 simulated clinical specimens. The detection limit of the qRT-PCR assay was 11.276 ZIKV RNA copies at the 95% probability level (probit analysis, p<= 0.05). Both Asian and African ZIKV strains were detected by the qRT-PCR assay without cross-reacting with DENV-1, DENV-2, DENV-3, DENV-4, CHIKV, JEV, LGTV, GETV and SINV. The qRT-PCR assay demonstrated a perfect agreement (k = 1.000, P < 0.001) with the reference assay; the sensitivity and specificity of the qRT-PCR assay were 100% (95% CI= 79.6-100) and 100% (95% CI= 43.9-100) respectively. The qRT-PCR assay developed in this study is a useful diagnostic tool for the broad coverage detection and quantification of both the Asian and African ZIKV strains.
    Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction; Real-Time Polymerase Chain Reaction
  15. Droplet digital PCR application for the detection of SARS-CoV-2 in air sample
    Rashid SA, Nazakat R, Muhamad Robat R, Ismail R, Suppiah J, Rajendran K, et al.
    Front Public Health, 2023;11:1208348.
    PMID: 37965510 DOI: 10.3389/fpubh.2023.1208348
    Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) may transmit through airborne route particularly when the aerosol particles remain in enclosed spaces with inadequate ventilation. There has been no standard recommended method of determining the virus in air due to limitations in pre-analytical and technical aspects. Furthermore, the presence of low virus loads in air samples could result in false negatives. Our study aims to explore the feasibility of detecting SARS-CoV-2 ribonucleic acid (RNA) in air samples using droplet digital polymerase chain reaction (ddPCR). Active and passive air sampling was conducted between December 2021 and February 2022 with the presence of COVID-19 confirmed cases in two hospitals and a quarantine center in Klang Valley, Malaysia. SARS-CoV-2 RNA in air was detected and quantified using ddPCR and real-time reverse transcriptase-polymerase chain reaction (RT-PCR). The comparability of two different digital PCR platforms (QX200 and QIAcuity) to RT-PCR were also investigated. Additionally negative staining transmission electron microscopy was performed to visualize virus ultrastructure. Detection rates of SARS-CoV-2 in air samples using ddPCR were higher compared to RT-PCR, which were 15.2% (22/145) and 3.4% (5/145), respectively. The sensitivity and specificity of ddPCR was 100 and 87%, respectively. After excluding 17 negative samples (50%) by both QX200 and QIAcuity, 15% samples (5/34) were found to be positive both ddPCR and dPCR. There were 23.5% (8/34) samples that were detected positive by ddPCR but negative by dPCR. In contrast, there were 11.7% (4/34) samples that were detected positive by dPCR but negative by ddPCR. The SARS-CoV-2 detection method by ddPCR is precise and has a high sensitivity for viral RNA detection. It could provide advances in determining low viral titter in air samples to reduce false negative reports, which could complement detection by RT-PCR.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction/methods
  16. Fulltext Duplex TaqMan Hydrolysis Probe-Based Molecular Assay for Simultaneous Detection and Differentiation of Burkholderia pseudomallei and Leptospira spp. DNA
    Mohd Ali MR, Lih Huey L, Foo PC, Goay YX, Ismail AS, Mustaffa KMF, et al.
    Biomed Res Int, 2019;2019:9451791.
    PMID: 31355287 DOI: 10.1155/2019/9451791
    Melioidosis and leptospirosis, caused by two different bacteria, Burkholderia pseudomallei and Leptospira spp., are potentially fatal infections that share a very similar spectrum of clinical features and cause significant mortality and morbidity in humans and livestock. Early detection is important for better clinical consequences. To our knowledge, there is no diagnostic tool available to simultaneously detect and differentiate melioidosis and leptospirosis in humans and animals. In this study, we described a duplex TaqMan probe-based qPCR for the detection of B. pseudomallei and Leptospira spp. DNA. The performance of the assay was evaluated on 20 B. pseudomallei isolates, 23 Leptospira strains, and 39 other microorganisms, as well as two sets of serially diluted reference strains. The duplex qPCR assay was able to detect 0.02 pg (~ 4 copies) Leptospira spp. DNA and 0.2 pg (~ 25.6 copies) B. pseudomallei DNA. No undesired amplification was observed in other microorganisms. In conclusion, the duplex qPCR assay was sensitive and specific for the detection of B. pseudomallei & Leptospira spp. DNA and is suitable for further analytical and clinical evaluation.
    Matched MeSH terms: Real-Time Polymerase Chain Reaction*
  17. Fulltext Molecular characterization of Brazilian FeLV strains in São Luis, Maranhão Brazil
    Martins NDS, Rodrigues APS, Bicalho JM, Albuquerque JJ, Reis LL, Alves LL, et al.
    Virus Genes, 2023 Aug;59(4):562-571.
    PMID: 37195404 DOI: 10.1007/s11262-023-01997-x
    The feline leukemia virus (FeLV) belongs to the Retroviridae family and Gammaretrovirus genus, and causes a variety of neoplastic and non-neoplastic diseases in domestic cats (Felis catus), such as thymic and multicentric lymphomas, myelodysplastic syndromes, acute myeloid leukemia, aplastic anemia, and immunodeficiency. The aim of the present study was to carry out the molecular characterization of FeLV-positive samples and determine the circulating viral subtype in the city of São Luís, Maranhão, Brazil, as well as identify its phylogenetic relationship and genetic diversity. The FIV Ac/FeLV Ag Test Kit (Alere™) and the commercial immunoenzymatic assay kit (Alere™) were used to detect the positive samples, which were subsequently confirmed by ELISA (ELISA - SNAP® Combo FeLV/FIV). To confirm the presence of proviral DNA, a polymerase chain reaction (PCR) was performed to amplify the target fragments of 450, 235, and 166 bp of the FeLV gag gene. For the detection of FeLV subtypes, nested PCR was performed for FeLV-A, B, and C, with amplification of 2350-, 1072-, 866-, and 1755-bp fragments for the FeLV env gene. The results obtained by nested PCR showed that the four positive samples amplified the A and B subtypes. The C subtype was not amplified. There was an AB combination but no ABC combination. Phylogenetic analysis revealed similarities (78% bootstrap) between the subtype circulating in Brazil and FeLV-AB and with the subtypes of Eastern Asia (Japan) and Southeast Asia (Malaysia), demonstrating that this subtype possesses high genetic variability and a differentiated genotype.
    Matched MeSH terms: Polymerase Chain Reaction/veterinary
  18. The use of a molecular technique for the detection of porcine ingredients in the Malaysian food market
    Farouk AE, Batcha MF, Greiner R, Salleh HM, Salleh MR, Sirajudin AR
    Saudi Med J, 2006 Sep;27(9):1397-400.
    PMID: 16951781
    To develop a molecular technique that is fast and reliable in detecting porcine contamination or ingredients in foods.
    Matched MeSH terms: Polymerase Chain Reaction/methods
  19. Development of Molecular Diagnosis by PCR for the Detection of Infection and Gene Expression for Nipah Virus (NiV)
    Pasha F, Alatawi A, Amir M, Faridi U
    Pak J Biol Sci, 2020 Jan;23(8):1086-1095.
    PMID: 32700860 DOI: 10.3923/pjbs.2020.1086.1095
    BACKGROUND AND OBJECTIVE: The epidemiology of Nipah virus (NiV) was shortly seen in many Asian countries like Malaysia, Bangladesh and India most recently. Nipah virus also synonym as bat born virus is transmitted primarily by fruit bats. The 2 different strains transmitted are Hendra (highly pathogenic) and Cedar (non-pathogenic). The present study was attempt to develop recombinant protein based reagents for molecular diagnosis of Nipah.

    MATERIALS AND METHODS: The different primer sets were developed using bioinformatics software DNASTAR. The E. coli cells were used for recombinant protein expression.

    RESULTS: The NiV 'G' region primers were designed and amplified for 1 kb fragment and cloned. The NiV 'G' fragments were sub-cloned in pET-28(+) B and pGEX-5x-1. Recombinant protein thus obtained in soluble form in both the cases was essayed using western blot. The result showed the protein expression yield was more in pET-28(+) B with low stability and vice versa for pGEX-5x-1.

    CONCLUSION: The antibodies raised from the protein can be used as diagnostic reagent for detection of NiV. Thus, a new diagnostic technique can be industrialized.

    Matched MeSH terms: Polymerase Chain Reaction/methods*
  20. Nanoplate-based digital PCR for highly sensitive pork DNA detection targeting multi-copy nuclear and mitochondrial genes
    Mokhtar NFK, Shun YQ, Raja Nhari RMH, Mohamad NA, Shahidan NM, Warsanah IH, et al.
    Food Addit Contam Part A Chem Anal Control Expo Risk Assess, 2024 Feb;41(2):120-133.
    PMID: 38190283 DOI: 10.1080/19440049.2023.2298476
    The inclusion of ingredients derived from pigs in highly processed consumer products poses a significant challenge for DNA-targeted analytical enforcement, which could be overcome by using digital PCR. However, most species detection methods use digital PCR to target single-copy nuclear genes, which limits their sensitivity. In this work, we examined the performance of a nanoplate-based digital PCR method that targets multi-copy nuclear (MPRE42) and mitochondrial (Cytb) genes. Poor separation of positive and negative partitions, as well as a 'rain effect' were obtained in the porcine-specific MPRE42 assay. Among the optimization strategies examined, the inclusion of restriction enzymes slightly improved the separation of positive and negative partitions, but a more extensive 'rain effect' was observed. The high copy number of the MPRE42 amplicon is hypothesized to contribute to the saturation of the positive signal. In contrast, the porcine-specific Cytb assay achieved perfect separation of positive and negative partitions with no 'rain effect'. This assay can detect as little as 0.4 pg of pork DNA, with a sensitivity of 0.05% (w/w) in a pork-chicken mixture, proving its applicability for detecting pork in meat and meat-based products. For the MPRE42 assay, potential applications in highly degraded products such as gelatin and lard are anticipated.
    Matched MeSH terms: Polymerase Chain Reaction/methods
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