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Fulltext Role of the CYP1A2 Gene Polymorphism on Early Ageing from Occupational Exposure

Eshkoor S, Ismail P, Rahman S, Moin S, Adon M

Balkan J. Med. Genet., 2013 Dec;16(2):45-52.
PMID: 24778563 DOI: 10.2478/bjmg-2013-0031

The ageing process is influenced by many internal and external factors. The toxic substances in the environment can cause genomic damages to cells, which increase the risk of early ageing. Furthermore, the cytochrome P450 1A2 (CYP1A2) gene polymorphism is a susceptibility factor and may enhance the risk of DNA damage in cells. The current study was carried out to show whether occupational exposure could cause genotoxicity in cells carrying the CYP1A2 gene polymorphism, thus enhancing the likelihood of early ageing. This study was conducted on mechanical workshop workers and a control group by collecting buccal cells from their mouths. Restriction fragment length polymorphism-polymerase chain reaction (RFLP-PCR) was used to identify the CYP1A2 gene polymorphism in the cells. In addition, three extra methods including micronuclei (MN) test, comet assay and real-time PCR (RT-PCR) were applied to determine the effects of gene polymorphisms on DNA damage and ageing from occupational exposure. The results showed that DNA damage in the cells carrying the mutated genotype was higher than the wild genotype. In addition, the difference in MN frequency (p = 0.001) and relative telomere length (p = 0.002) between workers and controls was significant (p <0.05) in the mutated genotype. The findings indicated a possible protective effect of gene polymorphism against early ageing, which was characterized by lack of a significant influence of CYP1A2 gene polymorphism on genetic material in the subjects (p >0.05). It was concluded that the CYP1A2 gene could be a contributing factor to prevent early ageing from occupational exposure.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
p16 gene expression in basal cell carcinoma

Eshkoor SA, Ismail P, Rahman SA, Oshkour SA

Arch Med Res, 2008 Oct;39(7):668-73.
PMID: 18760195 DOI: 10.1016/j.arcmed.2008.06.003

Basal cell carcinoma (BCC) develops predominantly in sun-exposed skin in fair-skinned individuals prone to sunburn. BCC typically occurs in adults. High exposure to ultraviolet (UV) radiation increases rate of developing BCC, a slowly growing tumor that occurs in hair-growing squamous epithelium and rarely metastasizes. In genetic studies, BCC patients have cell-cycle abnormalities of different parts of the signaling pathway. Retinoblastoma regulatory pathway is important in cell cycle arrest. In this pathway, p16INK4a, an inhibitor of Rb pathway, binds to CDK4 and CDK6 competitively with cyclin D1 to prevent phosphorylation of tumor suppressor pRB gene. Alteration of this pathway contributes to development of human cancers and also is effective in skin cancers. In this study, we analyzed mRNA expression using in situ RT-PCR and the role of immunohistochemical expression of p16INK4a in BCC.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Fulltext Circulation of human respiratory syncytial virus strains among hospitalized children with acute lower respiratory infection in malaysia

Etemadi MR, Sekawi Z, Othman N, Lye MS, Moghaddam FY

Evol Bioinform Online, 2013;9:151-61.
PMID: 23641140 DOI: 10.4137/EBO.S10999

Human respiratory syncytial virus (RSV) is a major viral pathogen associated with acute lower respiratory tract infections (ALRTIs) among hospitalized children. In this study, the genetic diversity of the RSV strains was investigated among nasopharyngeal aspirates (NPA) taken from children less than 5 years of age hospitalized with ALRTIs in Hospital Serdang, Malaysia. A total of 165 NPA samples were tested for the presence of RSV and other respiratory viruses from June until December 2009. RSV was found positive in 83 (50%) of the samples using reverse transcription polymerase chain reaction (RT-PCR). Further classification of 67 RSV strains showed that subgroups A and B comprised 11/67 (16.4%) and 56/67 (83.6%) of the strains, respectively. The second hypervariable region at the carboxyl-terminal of the G gene was amplified and sequenced in order to do phylogenetic study. The phylogenetic relationships of the samples were determined separately for subgroups A and B using neighbor joining (NJ), maximum parsimony (MP), and Bayesian inference (BI). Phylogenetic analysis of the 32 sequenced samples showed that all 9 RSV-A strains were clustered within NA1 genotype while the remaining 23 strains of the RSV-B subgroup could be grouped into a clade consisted of strains with 60-nucleotide duplication region. They were further classified into newly discovered BA10 and BA9 genotypes. The present finding suggests the emergence of RSV genotypes of NA1 and BA. This is the first documentation of the phylogenetic relationship and genetic diversity of RSV strains among hospitalized children diagnosed with ALRTI in Serdang, Malaysia.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Gene expression characteristic in human auricular cartilage tissue engineering

Farah Wahida I, Aminuddin BS, Munirah S, Chua KH, Fuzina NH, Isa MR, et al.

Med J Malaysia, 2004 May;59 Suppl B:190-1.
PMID: 15468882

This study was to assess collagen type II and collagen type I gene expression in tissue-engineered human auricular: cartilage formed via tissue engineering technique. Large-scale culture expansions were transformed into 3D in vitro construct and were implanted subcutaneously on the dorsal of athymic mice. After 8 weeks, explanted construct was processed in the same manner of native cartilage to facilitate cells for gene expression analysis. Isolated cells from in vivo construct demonstrated expression of type II collagen gene comparable to native cartilage. This study verified that tissue-engineered auricular cartilage expressed cartilage specific gene, collagen type II after in vivo maturation.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Fulltext Expression of a key gene involved in the biosynthetic pathway of vitamin E in red pericarp and white rice grains

Fasahat, P., Abdullah, A., Muhammad, K., Wickneswari, R.

International Food Research Journal, 2013;20(6):3395-3401.
MyJurnal

Tocochromanols (tocopherols and tocotrienols) unitedly known as vitamin E, are the necessary antioxidant components of both human and animal diets. There is a considerable interest in plants with increased or customized vitamin E content, due to their potential health benefits. To quantify the tocochromanol content and determine the expression of a key tocotrienol biosynthesis gene among a set of contrasting red pericarp and light brown rice genotypes of advanced breeding lines together with their parents; expression pattern of homogentisate geranylgeranyl transferase (HGGT), the key gene was studied by semi-quantitative RT-PCR in milky and matured grain stages. Vitamin E analysis was carried out by high performance liquid chromatography (HPLC). The chloroform-methanolic extracts prepared from red pericarp and light brown rice advanced breeding lines showed significant differences for vitamin E content. Averaged across all samples, the content of γ-tocotrienol > α-tocopherol > α-tocotrienol > γ-tocopherol > δ-tocotrienol, and total E vitamin content ranged from 10.30 to 31.65 µg/g. Genotype G37 (red pericarp) was found to have higher expression than G7 (light brown) and G33 (red pericarp) at both grain development stages but lower than both parents whereas their transcript levels were comparatively lower in mature grain, which indicates their possible regulation by plant growth stage. HPLC results of γ-tocotrienol content supported gene expression results with the exception of the recurrent parent MR219.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Fulltext Ex-vivo differentiation of stem cells from human extracted deciduous teeth into bone forming cells

Fazliah, S.N., Jaafar, S., Shamsuddin, S., Zainudin, Z., Hilmi, A.B., Razila, A.R., et al.

ASM Science Journal, 2010;4(1):1-14.
MyJurnal

Stem cells from human extracted deciduous teeth (SHED) have the ability to multiply much faster and double their population in culture at a greater rate, indicating that it may be in a more immature state than other type of adult stem cells. Mesenchymal stem cells (MSC) from human primary molars were isolated and cultured in media supplemented with 20% fetal bovine serum. The MSCs were confirmed using CD 105 and CD 166 and the identification of the osteoblast cells were done using reverse transcriptase polymerase chain reaction (RT-PCR) analysis. Differentiated osteoblast cells (DOC) were characterized by alkaline phosphotase and von Kossa staining followed by immunocytochemistry staining using osteocalcin and osteonectin antibodies. Further validation of SHED was done by RT-PCR to detect the presence of insulin-like growth factor 2 (IGF-2) and discoidin domain tyrosine kinase-2 (DDTK-2) transcripts, while the presence of Runx-2 mRNA was used to characterize DOC. The results showed that SHED was found positive for CD 105 and CD 166 and could differentiate into osteoblast, bone forming cells. The findings revealed the presence of distinct MSC population which had the capability to generate living human cells that could be a possible source for tissue engineering in the future.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Labisia pumila extract down-regulates hydroxysteroid (11-beta) dehydrogenase 1 expression and corticosterone levels in ovariectomized rats

Fazliana M, Gu HF, Östenson CG, Yusoff MM, Wan Nazaimoon WM

J Nat Med, 2012 Apr;66(2):257-64.
PMID: 21833773 DOI: 10.1007/s11418-011-0575-1

We evaluated the effects of a standardized Labisia pumila var. alata (LPva) extract on body weight change, hydroxysteroid (11-beta) dehydrogenase 1 (HSD11B1) expressions and corticosterone (CORT) level in ovariectomized (OVX) rats. The decoction of LPva has been used for generations among Malay women in Malaysia to maintain a healthy reproductive system.Thirty-six Sprague-Dawley OVX rats were treated orally with LPva extract (10, 20 or 50 mg/kg/day) or estrogen replacement (ERT) for 30 days. Sham operated rats were used as controls. Compared to untreated OVX rats, LPva-treated rats showed less weight gain and had significantly down-regulated HSD11B1 mRNA in liver tissues. HSD11B1 mRNA in adipose tissues increased by 55% (p < 0.05) in OVX rats but normalized in rats treated with LPva. Similarly, there was significant down-regulation (p < 0.05) of protein levels of HSD11B1 in both liver and adipose tissue of LPva and ERT groups, and CORT levels were significantly reduced in both groups of rats. This is the first study ever conducted to evaluate the beneficial effects of LPva in relation to weight gain caused by estrogen insufficiency. Results implied that the bioactive components in LPva extract affect not only HSD11B1 expressions in both adipose and liver tissues but also decrease circulating CORT. The extract should be explored for its potential use as a natural remedy for weight management.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Panspecies molecular assays detect viral pathogens missed by real-time PCR/reverse-transcriptase PCR among pneumonia patients, Sarawak, Malaysia

Fieldhouse JK, Bailey ES, Toh TH, Hii KC, Mallinson KA, Ting J, et al.

Trop Dis Travel Med Vaccines, 2020;6:13.
PMID: 32817802 DOI: 10.1186/s40794-020-00114-2

Background: In a year-long pneumonia etiology study conducted June 2017 to May 2018 in Sarawak, Malaysia, 599 patients' nasopharyngeal swab specimens were studied with real-time polymerase chain reaction (rPCR)/ reverse-transcription (rRT-PCR) assays for respiratory pathogens known to contribute to the high burden of lower respiratory tract infections. The study team sought to compare real-time assay results with panspecies conventional molecular diagnostics to compare sensitivities and learn if novel viruses had been missed.
Methods: Specimens were studied for evidence of adenovirus (AdV), enterovirus (EV) and coronavirus (CoV) with panspecies gel-based nested PCR/RT-PCR assays. Gene sequences of specimens positive by panspecies assays were sequenced and studied with the NCBI Basic Local Alignment Search Tool software.
Results: There was considerable discordance between real-time and conventional molecular methods. The real-time AdV assay found a positivity of 10.4%; however, the AdV panspecies assay detected a positivity of 12.4% and the conventional AdV-Hexon assay detected a positivity of 19.6%. The CoV and EV panspecies assays similarly detected more positive specimens than the real-time assays, with a positivity of 7.8% by the CoV panspecies assay versus 4.2% by rRT-PCR, and 8.0% by the EV panspecies assay versus 1.0% by rRT-PCR. We were not able to ascertain virus viability in this setting. While most discordance was likely due to assay sensitivity for previously described human viruses, two novel, possible zoonotic AdV were detected.
Conclusions: The observed differences in the two modes of amplification suggest that where a problem with sensitivity is suspected, real-time assay results might be supplemented with panspecies conventional PCR/RT-PCR assays.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Gene expression analysis of osteoblasts seeded in coral scaffold

Foo LH, Suzina AH, Azlina A, Kannan TP

J Biomed Mater Res A, 2008 Oct;87(1):215-21.
PMID: 18085658

Coral matrix of Porites sp. has the suitable properties for bone cell growth. This study was aimed to study the gene expression levels of osteoblast specific genetic markers; RUNX2, osteopontin, alkaline phosphatase and osteocalcin from osteoblasts seeded in coral scaffold, which are important in determining the feasibility of osteoblasts. Human osteoblasts were inoculated onto the processed coral in Dulbecco's Minimum Essential Medium. The cells were trypsinized on day 1, 7, 14, 18, and 21 and added with RNALater for preservation of RNA in cells. The RNA was extracted using commercial RNA extraction kit and the respective genes were amplified using RT-PCR kit and analyzed qualitatively on 1.5% agarose gel. The expressions were evaluated with the Integrated Density Value based on the intensity of band for different periods of cell harvest. Increased expressions of the RUNX2, osteopontin, alkaline phosphatase and osteocalcin genes in the present study proved that coral is a favorable carrier for osteogenetically competent cells to attach and remain viable.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Genetical and immunological analysis of recent Asian type A and O foot-and-mouth disease virus isolates

Freiberg B, Rahman MM, Marquardt O

Virus Genes, 1999;19(3):167-82.
PMID: 10595408

This report extends the knowledge on the epizootical situation of foot-and-mouth disease in Asia. RNA from six samples of type A and five of type O virus, isolated between 1987 and 1997 in Bangladesh, Iran, Malaysia and Turkey, was subjected to reverse transcription-dependent polymerase chain reactions that amplify large parts of the capsid protein VP1 encoding genome region. The amplification products were sequenced, and the sequences aligned to each other and to published sequences. This showed the type O isolates of 1987-1997 from Bangladesh to be of same genotype and closely related to isolates of 1988 and later from Saudi Arabia, 1990 from India, 1996 from Greece and Bulgaria, and 1997 from Iran. Among the analyzed type A isolates, those of 1992 and 1996 from Turkey were of same genotype and related to previously described isolates of 1987 from Iran and of 1992 from Saudi Arabia. The isolate of 1997 from Malaysia was found to be related to isolates from Thailand of 1993 and 1996. The isolates of 1987 from Bangladesh and 1997 from Iran, however, represent different so far not described genotypes. Monoclonal antibodies, raised against the vaccine production strains A22 Iraq, Asial Shamir, O1 Kaufbeuren and O1 Manisa, and the recent type A field isolates Saudi Arabia/92 and Albania/96, were used in an ELISA to compare the reaction patterns of many of the field isolates. The monoclonal antibodies were further characterized for virus-neutralizing activity and binding to trypsinized homologous virus. The failure of neutralizing antibodies in binding to trypsinized homologous as well as to heterologous virus suggested the epitopes to reside at the major antigenic component of the virus, which is the capsid protein VP1. Two non-neutralizing antibodies that bind to trypsin-sensitive epitopes cross-reacted, however, with heterologous virus. This indicates the existence of a trypsin-sensitive antigenic site outside of VP1. In summary, the results obtained by ELISA confirm the observed sequence differences, but indicate further sequence differences at minor antigenic sites that do not reside on VP1.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Fulltext SARS-CoV-2 multiplex RT-PCR to detect variants of concern (VOCs) in Malaysia, between January to May 2021

Fu JYL, Chong YM, Sam IC, Chan YF

J Virol Methods, 2022 Mar;301:114462.
PMID: 35026305 DOI: 10.1016/j.jviromet.2022.114462

Emerging SARS-CoV-2 variants of concern (VOC) have been associated with enhanced transmissibility and immune escape. Next-generation sequencing (NGS) of the whole genome is the gold standard for variant identification for surveillance but is time-consuming and costly. Rapid and cost-effective assays that detect SARS-CoV-2 variants are needed. We evaluated Allplex SARS-CoV-2 Master Assay and Variants I Assay to detect HV69/70 deletion, Y144 deletion, E484K, N501Y, and P681H spike mutations in 248 positive samples collected in Kuala Lumpur, Malaysia, between January and May 2021. Spike variants were detected in 78/248 (31.5 %), comprising 60 VOC B.1.351 (beta) and 18 B.1.1.7 (alpha). With NGS as reference for 115 samples, the sensitivity for detecting the spike mutations was 98.7 % with the Master Assay and 100 % with the Variants I Assay. The emergence of beta variants correlated with increasing COVID-19 infections in Malaysia. The prevalence of alpha VOC and lineage B.1.466.2 was low. These assays detect mutations present in alpha, beta and gamma VOCs. Of the VOCs which have subsequently emerged, the assays should detect omicron (B.1.1.529) but not B.1.617.2 (delta). In conclusion, spike variant PCR assays can be used to rapidly monitor selected SARS-CoV-2 VOCs in resource-limited settings, but require updates as new variants emerge.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Fulltext Detection of novel respiratory viruses from influenza-like illness in the Philippines

Furuse Y, Suzuki A, Kishi M, Galang HO, Lupisan SP, Olveda RM, et al.

J Med Virol, 2010 May;82(6):1071-4.
PMID: 20419824 DOI: 10.1002/jmv.21763

Several novel viruses have been recently identified in respiratory samples. However, the epidemiology of these viruses in tropical countries remains unclear. The aim of the present study was to provide an overview of the epidemiology of novel respiratory viruses, including human metapneumovirus, human bocavirus, new subtypes of human coronavirus (NL63 and HKU1), KI virus, WU virus, and Melaka virus in the Philippines, a tropical country. Nasopharyngeal aspirates from 465 patients with influenza-like illness were collected in 2006 and 2007. Reverse transcription polymerase chain reaction (RT-PCR) and PCR were performed to detect viruses from culture-negative specimens. Human metapneumovirus, human bocavirus, human coronavirus HKU1, KI virus, and WU virus were detected for the first time in the Philippines; Melaka virus was not found.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Fulltext Ankrd11 is a chromatin regulator involved in autism that is essential for neural development

Gallagher D, Voronova A, Zander MA, Cancino GI, Bramall A, Krause MP, et al.

Dev. Cell, 2015 Jan 12;32(1):31-42.
PMID: 25556659 DOI: 10.1016/j.devcel.2014.11.031

Ankrd11 is a potential chromatin regulator implicated in neural development and autism spectrum disorder (ASD) with no known function in the brain. Here, we show that knockdown of Ankrd11 in developing murine or human cortical neural precursors caused decreased proliferation, reduced neurogenesis, and aberrant neuronal positioning. Similar cellular phenotypes and aberrant ASD-like behaviors were observed in Yoda mice carrying a point mutation in the Ankrd11 HDAC-binding domain. Consistent with a role for Ankrd11 in histone acetylation, Ankrd11 was associated with chromatin and colocalized with HDAC3, and expression and histone acetylation of Ankrd11 target genes were altered in Yoda neural precursors. Moreover, the Ankrd11 knockdown-mediated decrease in precursor proliferation was rescued by inhibiting histone acetyltransferase activity or expressing HDAC3. Thus, Ankrd11 is a crucial chromatin regulator that controls histone acetylation and gene expression during neural development, thereby providing a likely explanation for its association with cognitive dysfunction and ASD.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Standard CYP2D6 genotyping procedures fail for the CYP2D6*5 and duplication alleles when hair roots are used as a source of DNA

Gan SH, Ismail R, Wan Adnan WA, Wan Z

Clin Chim Acta, 2003 Mar;329(1-2):61-8.
PMID: 12589966 DOI: 10.1016/s0009-8981(03)00019-6

BACKGROUND: Hair roots provide a useful alternative to blood as a source of DNA for genotyping. Besides simple and non-invasive collections, the DNA extraction step is also easy to perform and is fast. The aim of our study is to determine if hair roots can be used to genotype all of the common CYP2D6 alleles for routine screening purposes.

METHOD: The study complies with the Declaration of Helsinki. After obtaining informed consents, both blood and hair samples were collected from 92 patients for genotyping of the CYP2D6 gene. PCR was used to detect the following mutations: CYP2D6*1, *3, *4, *5, *9, *10, *17 and duplication gene. The results were compared where hair roots and blood were used as templates for DNA respectively.

RESULTS: When blood was used as a source of DNA for genotyping, all of the investigated CYP2D6 alleles were successfully amplified. However, with hair roots, the genes with the larger fragment sizes: CYP2D6*5 and the duplication gene could not be amplified and the bands of other alleles investigated were faint when visualized under UV light.

CONCLUSIONS: DNA extraction from hair roots and leucocytes yielded similar results but the DNA extracted from hair roots did not allow successful amplification of the longer genes such as the CYP2D6*5 and the duplication gene.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
The mazEF toxin-antitoxin system as a novel antibacterial target in Acinetobacter baumannii

Ghafourian S, Good L, Sekawi Z, Hamat RA, Soheili S, Sadeghifard N, et al.

Mem Inst Oswaldo Cruz, 2014 Jul;109(4):502-5.
PMID: 25004148

Although analysis of toxin-antitoxin (TA) systems can be instructive, to date, there is no information on the prevalence and identity of TA systems based on a large panel of Acinetobacter baumannii clinical isolates. The aim of the current study was to screen for functional TA systems among clinical isolates of A. baumannii and to identify the systems' locations. For this purpose, we screened 85 A. baumannii isolates collected from different clinical sources for the presence of the mazEF, relBE and higBA TA genes. The results revealed that the genes coding for the mazEF TA system were commonly present in all clinical isolates of A. baumannii. Reverse transcriptase-polymerase chain reaction analysis showed that transcripts were produced in the clinical isolates. Our findings showed that TA genes are prevalent, harboured by chromosomes and transcribed within A. baumannii. Hence, activation of the toxin proteins in the mazEF TA system should be investigated further as an effective antibacterial strategy against this bacterium.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Fulltext Induction of apoptosis in MCF-7 cells by the hemagglutinin-neuraminidase glycoprotein of Newcastle disease virus Malaysian strain AF2240

Ghrici M, El Zowalaty M, Omar AR, Ideris A

Oncol Rep, 2013 Sep;30(3):1035-44.
PMID: 23807159 DOI: 10.3892/or.2013.2573

Newcastle disease virus (NDV) exerts its naturally occurring oncolysis possibly through the induction of apoptosis. We hypothesized that the binding of the virus to the cell via the hemagglutinin-neuraminidase (HN) glycoprotein may be sufficient to not only induce apoptosis but to induce a higher apoptosis level than the parental NDV AF2240 virus. NDV AF2240 induction of apoptosis in MCF-7 human breast cancer cells was analyzed and quantified. In addition, the complete HN gene of NDV strain AF2240 was amplified, sequenced and cloned into the pDisplay eukaryotic expression vector. HN gene expression was first detected at the cell surface membrane of the transfected MCF-7 cells. HN induction of apoptosis in transfected MCF-7 cells was analyzed and quantified. The expression of the HN gene alone was able to induce apoptosis in MCF-7 cells but it was a less potent apoptosis inducer compared to the parental NDV AF2240 strain. In conclusion, the NDV AF2240 strain is a more suitable antitumor candidate agent than its recombinant HN gene unless the latter is further improved by additional modifications.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Heterogeneous t(4;11) fusion transcripts in two infants with acute lymphoblastic leukemia

Gill HK, Ten SK, Dhaliwal JS, Moore S, Hassan R, Karim FA, et al.

Malays J Pathol, 2004 Dec;26(2):105-10.
PMID: 16329562

An RT-PCR assay detected the t(4;11) translocation in two infants with acute lymphoblastic leukemia (ALL). Case P76 was a 10-month-old, female infant, who presented with a WBC of 137.4 x 10(9)/l and a pre-pre-B ALL immunophenotype. Case P120 was a 6-month-old female infant, with a WBC > 615 x 10(9)/l and a pre-pre-B ALL immunophenotype. RT-PCR of cDNA from both these cases generated a 656 bp and a 542 bp respectively, which sequencing confirmed as t(4;11) fusion transcripts. The primers and conditions selected for this assay are compatible with a one-step multiplex PCR for the main translocations in childhood ALL.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
TEL-AML1 frequency in multi-ethnic Malaysian pediatric acute lymphoblastic leukemia

Gill HK, Keoh TS, Dhaliwal JS, Moore S, Kim TS, Hassan R, et al.

Cancer Genet. Cytogenet., 2005 Jan 15;156(2):129-33.
PMID: 15642392

Eighty-eight multi-ethnic Malaysian pediatric acute lymphoblastic leukemia (ALL) patients were screened for the TEL-AML1 rearrangement by reverse transcription-polymerase chain reaction (RT-PCR). Fluorescence in situ hybridization (FISH) was used as an independent screen for 30 cases and to confirm RT-PCR positive cases. Seventeen patients, or 19%, were found to be t(12;21) positive. Ethnically the group comprised 12 Malays, 4 Chinese, and 1 Indian. All patients, including 1 with an unusual blast cell morphology who suffered an early relapse and death, were characteristic TEL-AML1 cases in cell count, age, ALL subset classification, and fusion transcript expressed. This study shows that in Malaysia, TEL-AML1 is found in the same distinct ALL subset and at a similar frequency as in other diverse childhood ALL cohorts.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Fulltext Defining p47-phox deficient Chronic Granulomatous Disease in a Malay family

Gill HK, Kumar HC, Dhaliwal JS, Zabidi F, Sendut IH, Noah RM, et al.

Asian Pac J Allergy Immunol, 2012 Dec;30(4):313-20.
PMID: 23393912

BACKGROUND: The most common autosomal form of Chronic Granulomatous Disease, p47-phox deficient CGD, generally features a GT (deltaGT) deletion in the GTGT sequence at the start of exon 2 on the NCF-1 gene. This consistency is due to the coexistence of and the recombination between 2 homologous pseudogenes (psi s) and NCF-1. The GTGT: deltaGT ratio mirrors the NCF-I: NCF-1 psi ratio and is 2:4 in normal individuals.
OBJECTIVE: To determine the molecular basis of the Autosomal-CGD in a family with 2 children, a male and female, affected by the disease. The female patient suffered recurrent infection, retinitis pigmentosa and discoid lupus.
METHODS: Chemiluminescence (CL) was used to study the respiratory burst, while genetic analysis was done by RT-PCR, PCR, deltaGT and the 20bp gene scans.
RESULTS: The CL response of the patient was profoundly low. The patient's p47-phox band was absent in the RT-PCR for NADPH-oxidase component mRNAs. The deltaGT scan showed that the patient's GTGT: deltaGT ratio was 0:6, the parents' and the younger brother's was 1:5 and the younger sister's was 2:4. Examination of other NCF-1/ NCF-1 psi s differences showed that the father had a compound deltaGT allele ie. deltaGT-20bp, inherited by the patient, and that both parents had compound GTGT alleles with a single 30bp segment in intron 1.
CONCLUSIONS: The patient was a classic, homozygous deltaGT p47-phox deficient CGD with one allele harbouring a compound deltaGT-20bp gene. The deltaGT and 20bp gene scans offer a relatively simple and efficient means of defining a p47-phox deficient CGD patient.
Key words: Chronic Granulomatous Disease, Primary Immunodeficiency, NCF-1, p47-phox, NADPH-oxidas

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction
Micromanipulation of culture niche permits long-term expansion of dental pulp stem cells--an economic and commercial angle

Govindasamy V, Ronald VS, Totey S, Din SB, Mustafa WM, Totey S, et al.

In Vitro Cell Dev Biol Anim, 2010 Oct;46(9):764-73.
PMID: 20725801 DOI: 10.1007/s11626-010-9332-0

Stem cells isolated from dental pulp possess the capacity for self-renewal and the potential for multi-lineage differentiation. However, dental pulp stem cells have different characteristics in terms of their culture conditions. The success of stem cells culture is governed by its micro-environmental niche. Therefore, we studied the effects of culture niche on long-term expansion of dental pulp stem cells in terms of cell morphology, growth kinetics, senescence pattern, cell surface marker expression differentiation capacity, and seeding plating density of dental pulp stem cells in four different, widely used media composition Among the various basal media tested, α-minimum essential media and knock out-minimum essential media supplemented with 10% fetal bovine serum were found to be the most optimal media composition in preserving the phenotypic characteristics and differentiation potential for prolonged periods as compared with DMEM-F12 and DMEM-LG. Plating density has been shown to affect overall yield. As a conclusion, the adoption of an appropriate culture system significantly improved cell yield, thus enabling the attainment of sufficient yields for therapeutic applications economizing in terms of cost of production and minimizing seeding cell density for maximum yield.

Matched MeSH terms: Reverse Transcriptase Polymerase Chain Reaction

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