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  1. Fulltext A Comprehensive Review on Phytochemistry and Pharmacological Activities of Clinacanthus nutans (Burm.f.) Lindau
    Khoo LW, Audrey Kow S, Lee MT, Tan CP, Shaari K, Tham CL, et al.
    Evid Based Complement Alternat Med, 2018;2018:9276260.
    PMID: 30105077 DOI: 10.1155/2018/9276260
    Clinacanthus nutans (Burm.f.) Lindau (Acanthaceae), commonly known as Sabah snake grass, is a vegetable and a well-known herb that is considered an alternative medicine for insect bites, skin rashes, herpes infection, inflammation, and cancer and for health benefits. Current review aims to provide a well-tabulated repository of the phytochemical screening, identification and quantification, and the pharmacological information of C. nutans according to the experimental design and the plant preparation methods which make it outstanding compared to existing reviews. This review has documented valuable data obtained from all accessible library databases and electronic searches. For the first time we analyzed the presence of flavonoids, triterpenoids, steroids, phytosterols, and glycosides in C. nutans based on the results from phytochemical screening which are then further confirmed by conventional phytochemical isolation methods and advanced spectroscopic techniques. Phytochemical quantification further illustrated that C. nutans is a good source of phenolics and flavonoids. Pharmacological studies on C. nutans revealed that its polar extract could be a promising anti-inflammation, antiviral, anticancer, immune and neuromodulating, and plasmid DNA protective agent; that its semipolar extract could be a promising antiviral, anticancer, and wound healing agent; and that its nonpolar extract could be an excellent anticancer agent.
    Matched MeSH terms: Plasmids
  2. Efficacy of an inactivated recombinant vaccine encoding a fimbrial protein of Pasteurella multocida B:2 against hemorrhagic septicemia in goats
    Mohd Yasin IS, Mohd Yusoff S, Mohd ZS, Abd Wahid Mohd E
    Trop Anim Health Prod, 2011 Jan;43(1):179-87.
    PMID: 20697957 DOI: 10.1007/s11250-010-9672-5
    This study was carried out to determine the antibody responses and protective capacity of an inactivated recombinant vaccine expressing the fimbrial protein of Pasteurella multocida B:2 following intranasal vaccination against hemorrhagic septicemia in goats. Goats were vaccinated intranasal with 10(6) CFU/mL of the recombinant vaccine (vaccinated group) and 10(6) CFU/mL of pET32/LIC vector without fimbrial protein (control group). All three groups were kept separated before all goats in the three groups were challenged with 10(9) CFU/mL of live pathogenic P. multocida B:2. During the course of study, both serum and lung lavage fluid were collected to evaluate the antibody levels via enzyme-linked immunosorbent assay. It was found that goats immunized with the inactivated recombinant vaccine developed a strong and significantly (p 
    Matched MeSH terms: Plasmids/genetics
  3. Synthesis of isatin thiosemicarbazones derivatives: in vitro anti-cancer, DNA binding and cleavage activities
    Ali AQ, Teoh SG, Salhin A, Eltayeb NE, Khadeer Ahamed MB, Abdul Majid AM
    Spectrochim Acta A Mol Biomol Spectrosc, 2014 May 5;125:440-8.
    PMID: 24607427 DOI: 10.1016/j.saa.2014.01.086
    New derivatives of thiosemicarbazone Schiff base with isatin moiety were synthesized L1-L6. The structures of these compounds were characterized based on the spectroscopic techniques. Compound L6 was further characterized by XRD single crystal. The interaction of these compounds with calf thymus (CT-DNA) exhibited high intrinsic binding constant (k(b)=5.03-33.00×10(5) M(-1)) for L1-L3 and L5 and (6.14-9.47×10(4) M(-1)) for L4 and L6 which reflect intercalative activity of these compounds toward CT-DNA. This result was also confirmed by the viscosity data. The electrophoresis studies reveal the higher cleavage activity of L1-L3 than L4-L6. The in vitro anti-proliferative activity of these compounds against human colon cancer cell line (HCT 116) revealed that the synthesized compounds (L3, L6 and L2) exhibited good anticancer potency.
    Matched MeSH terms: Plasmids/metabolism
  4. The application of naked DNA plasmid (DrZP3) and recombinant adenovirus (Ad-rZP3) in rat animal model to determine comparative efficacy of ZP3-Immunocontraceptive vaccines
    Mohd-Lila MA, Yee LK, Cen LS, Bala JA, Balakrishnan KN, Allaudin ZN, et al.
    Microb Pathog, 2019 Sep;134:103572.
    PMID: 31163251 DOI: 10.1016/j.micpath.2019.103572
    The common physical and chemical methods for controlling rat pest are less than satisfactory and inhumane. Immunocontraception approach has been considered more humane and it can be accomplished by inducing the relevant host immune response that block further development of reproductive gametes. ZP3 proteins are known to play very important role during sperm-ovum fertilization. It is a self-antigen and only localized in female ovaries. Therefore, an immunization with ZP3 protein elsewhere will induce a generalize host immune response against ZP3 protein. This study employed rat ZP3 (rZP3) gene prepared from its cDNA of Rattus rattus diardii. It was delivered and expressed in vivo by naked plamid DNA (DrZP3) or recombinant ZP3-Adenovirus (Ad-rZP3). Expression studies in vitro with DrZP3 or Ad-ZP3 showed rZP3 proteins were successfully expressed in Vero cells. Hyperimmune serum against rZP3 that were prepared by immunizing several rats with purified rZP3-pichia yeast fusion protein showed it blocked sperms from binding DrZP3-transfected Vero cells. Female Sprague Dawley rats immunized with DrZP3 demonstrated a long-term effect for significant reduction of fertility up to 92.6%. Ovaries from rats immunized with DrZP3 were severely atrophied with disappearance of primordial follicles from ovarian cortex with an increased in the amount of oocyte-free cell clusters. Female rats immunized with Ad-rZP3 demonstrated 27% reduction of fertility. The infertility induced by Ad-rZP3 is comparatively low and ineffective. This could be due to a strong host immune response that suppresses the recombinant virus itself resulted in minimum rZP3 protein presentation to the host immune system. As a result, low antibody titers produced against rZP3 is insufficient to block oocytes from maturity and fertilization. Therefore, immunization with DrZP3 for immunocontraception is more effective than Ad-rZP3 recombinant adenovirus. It is proposed to explore further on the use of adenovirus or other alternative viruses to deliver ZP3 protein and for the development of enhanced expression of rZP3 in target host.
    Matched MeSH terms: Plasmids
  5. Fulltext Gene delivery potential of biofunctional carbonate apatite nanoparticles in lungs
    Alhaji SY, Chowdhury EH, Rosli R, Hassan F, Abdullah S
    Biomed Res Int, 2014;2014:646787.
    PMID: 25143941 DOI: 10.1155/2014/646787
    Existing nonviral gene delivery systems to lungs are inefficient and associated with dose limiting toxicity in mammalian cells. Therefore, carbonate apatite (CO3Ap) nanoparticles were examined as an alternative strategy for effective gene delivery to the lungs. This study aimed to (1) assess the gene delivery efficiency of CO3Ap in vitro and in mouse lungs, (2) evaluate the cytotoxicity effect of CO3Ap/pDNA in vitro, and (3) characterize the CO3Ap/pDNA complex formulations. A significantly high level of reporter gene expression was detected from the lung cell line transfected with CO3Ap/pDNA complex prepared in both serum and serum-free medium. Cytotoxicity analysis revealed that the percentage of the viable cells treated with CO3Ap to be almost similar to the untreated cells. Characterization analyses showed that the CO3Ap/pDNA complexes are in a nanometer range with aggregated spherical structures and tended to be more negatively charged. In the lung of mice, highest level of transgene expression was observed when CO3Ap (8 μL) was complexed with 40 μg of pDNA at day 1 after administration. Although massive reduction of gene expression was seen beyond day 1 post administration, the level of expression remained significant throughout the study period.
    Matched MeSH terms: Plasmids/metabolism
  6. Fulltext Limited expression of non-integrating CpG-free plasmid is associated with increased nucleosome enrichment
    Habib O, Mohd Sakri R, Ghazalli N, Chau DM, Ling KH, Abdullah S
    PLoS One, 2020;15(12):e0244386.
    PMID: 33347482 DOI: 10.1371/journal.pone.0244386
    CpG-free pDNA was reported to facilitate sustained transgene expression with minimal inflammation in vivo as compared to CpG-containing pDNA. However, the expression potential and impact of CpG-free pDNA in in vitro model have never been described. Hence, in this study, we analyzed the transgene expression profiles of CpG-free pDNA in vitro to determine the influence of CpG depletion from the transgene. We found that in contrast to the published in vivo studies, CpG-free pDNA expressed a significantly lower level of luciferase than CpG-rich pDNA in several human cell lines. By comparing novel CpG-free pDNA carrying CpG-free GFP (pZGFP: 0 CpG) to CpG-rich GFP (pRGFP: 60 CpGs), we further showed that the discrepancy was not influenced by external factors such as gene transfer agent, cell species, cell type, and cytotoxicity. Moreover, pZGFP exhibited reduced expression despite having equal gene dosage as pRGFP. Analysis of mRNA distribution revealed that the mRNA export of pZGFP and pRGFP was similar; however, the steady state mRNA level of pZGFP was significantly lower. Upon further investigation, we found that the CpG-free transgene in non-integrating CpG-free pDNA backbone acquired increased nucleosome enrichment as compared with CpG-rich transgene, which may explain the observed reduced level of steady state mRNA. Our findings suggest that nucleosome enrichment could regulate non-integrating CpG-free pDNA expression and has implications on pDNA design.
    Matched MeSH terms: Plasmids/genetics*
  7. Fulltext Advancements in reprogramming strategies for the generation of induced pluripotent stem cells
    Lai MI, Wendy-Yeo WY, Ramasamy R, Nordin N, Rosli R, Veerakumarasivam A, et al.
    J Assist Reprod Genet, 2011 Apr;28(4):291-301.
    PMID: 21384252 DOI: 10.1007/s10815-011-9552-6
    Direct reprogramming of somatic cells into induced pluripotent stem (iPS) cells has emerged as an invaluable method for generating patient-specific stem cells of any lineage without the use of embryonic materials. Following the first reported generation of iPS cells from murine fibroblasts using retroviral transduction of a defined set of transcription factors, various new strategies have been developed to improve and refine the reprogramming technology. Recent developments provide optimism that the generation of safe iPS cells without any genomic modification could be derived in the near future for the use in clinical settings. This review summarizes current and evolving strategies in the generation of iPS cells, including types of somatic cells for reprogramming, variations of reprogramming genes, reprogramming methods, and how the advancement iPS cells technology can lead to the future success of reproductive medicine.
    Matched MeSH terms: Plasmids
  8. Crystal structure of fuculose aldolase from the Antarctic psychrophilic yeast Glaciozyma antarctica PI12
    Jaafar NR, Littler D, Beddoe T, Rossjohn J, Illias RM, Mahadi NM, et al.
    Acta Crystallogr F Struct Biol Commun, 2016 11 01;72(Pt 11):831-839.
    PMID: 27827354
    Fuculose-1-phosphate aldolase (FucA) catalyses the reversible cleavage of L-fuculose 1-phosphate to dihydroxyacetone phosphate (DHAP) and L-lactaldehyde. This enzyme from mesophiles and thermophiles has been extensively studied; however, there is no report on this enzyme from a psychrophile. In this study, the gene encoding FucA from Glaciozyma antarctica PI12 (GaFucA) was cloned and the enzyme was overexpressed in Escherichia coli, purified and crystallized. The tetrameric structure of GaFucA was determined to 1.34 Å resolution. The overall architecture of GaFucA and its catalytically essential histidine triad are highly conserved among other fuculose aldolases. Comparisons of structural features between GaFucA and its mesophilic and thermophilic homologues revealed that the enzyme has typical psychrophilic attributes, indicated by the presence of a high number of nonpolar residues at the surface and a lower number of arginine residues.
    Matched MeSH terms: Plasmids/metabolism; Plasmids/chemistry
  9. Fulltext Prevalence of plasmid-bearing and plasmid-free Chlamydia trachomatis infection among women who visited obstetrics and gynecology clinics in Malaysia
    Yeow TC, Wong WF, Sabet NS, Sulaiman S, Shahhosseini F, Tan GM, et al.
    BMC Microbiol, 2016 Mar 18;16:45.
    PMID: 26987367 DOI: 10.1186/s12866-016-0671-1
    BACKGROUND: The 7.5 kb cryptic plasmid of Chlamydia trachomatis has been shown to be a virulence factor in animal models, but its significance in humans still remains unknown. The aim of this study was to investigate the prevalence and potential involvement of the C. trachomatis cryptic plasmid in causing various clinical manifestations; including infertility, reproductive tract disintegrity, menstrual disorder, and polycystic ovarian syndrome (PCOS) among genital C. trachomatis-infected patients.
    RESULTS:A tot l of 180 female patients of child bearing age (mean 30.9 years old, IQR:27-35) with gynecological complications and subfertility issues, who visited Obstetrics and Gynecology clinics in Kuala Lumpur, Malaysia were recruited for the study. Prevalence of genital chlamydial infection among these patients was alarmingly high at 51.1% (92/180). Of the 92 chlamydia-infected patients, 93.5% (86/92) were infected with plasmid-bearing (+) C. trachomatis while the remaining 6.5% (6/92) were caused by the plasmid-free (-) variant. Our data showed that genital C. trachomatis infection was associated with infertility issues, inflammation in the reproductive tract (mucopurulent cervicitis or endometriosis), irregular menstrual cycles and polycystic ovarian syndrome (PCOS). However, no statistical significance was detected among patients with plasmid (+) versus plasmid (-) C. trachomatis infection. Interestingly, plasmid (+) C. trachomatis was detected in all patients with PCOS, and the plasmid copy numbers were significantly higher among PCOS patients, relative to non-PCOS patients.
    CONCLUSION: Our findings show a high incidence of C. trachomatis infection among women with infertility or gynecological problems in Malaysia. However, due to the low number of plasmid (-) C. trachomatis cases, a significant role of the plasmid in causing virulence in human requires further investigation of a larger cohort.
    KEYWORDS: Chlamydia trachomatis; Infertility; Plasmid; Reproductive system disorders
    Study site: Obstetrics and Gynecology clinic, University Malaya Medical Centre (UMMC), Kuala Lumpur, Malaysia
    Matched MeSH terms: Plasmids/genetics*; Plasmids/metabolism
  10. Inadvertent expression of dengue 2 virus NS3-EGFP fusion protein in Escherlchla coll using the pEGFP-N 1 mammalian expression vector
    Norazizah S, AbuBakar S
    JUMMEC, 1999;4:41-46.
    Dengue 2 New Guinea C (NGC) virus NS3 protein, a potentially important virulence factor was cloned to the N-terminus of the Aeqirorea victoria enhanced green fluorescent protein (EGFP) using the pEGFP-N1 mammalian expression vector. During amplification of the recombinant plasmid in E. coli, transformants expressing the EGFP were detected in vivo when viewed using fluorescence microscopy. This inadvertent expression of the recombinant fusion protein was confirmed further by detection of the T7.Tag peptide cloned to the aluino terminal of the fusion protein using T 7.Tag specific monoclonal antibody. These findings represent perhaps the first reported expression of the T7.Tag-NS3-EGFP fusion protein using the pEGFP-N1 mammalian expression vector in E. coli. KEYWORDS: Dengue, NS3, pEGFP-N1, fusion protein.
    Matched MeSH terms: Plasmids
  11. Characterisation of simple sequence repeats in the Asian seabass, Lates Calcarifer by random sequencing
    Wan KL, Chong PP, Adura Mohd. Adnan
    Sains Malaysiana, 2011;40.
    In recent years, there has been considerable interest in simple sequence repeats (SSRs) particularly as molecular markers with applications in many different fields. We have carried out an effort to identify and analyse SSRs in the genome of the Asian seabass, Lates calcarifer by random sequencing. Genomic DNA was isolated from the muscle tissue of L. calcarifer, sheared by nebulisation and ligated into plasmid vector. Recombinant clones were selected randomly from the genomic libraries constructed. Subsequently, plasmid DNA was extracted and subjected to one-pass sequencing. A total of 4175 random sequences, also known as genome survey sequences (GSSs), with a total length of 1.7 Mb was generated. Screening of the whole L. calcarifer GSS data set allowed for the identification of a total of 151 perfect (100% similarity) SSRs. These SSR consensus patterns spread over a wide range of size (1 to 226 bp). The most frequent consensus pattern is dinucleotide, which represents 60% of all SSRs identified. The dinucleotides (AC)n, (AT)n and (AG)n were also found to occur frequently in the L. calcarifer genome. Sequence comparison between L. calcarifer and other fish species showed variation in repeat content, indicating the different ways in which repeats may evolve in the genome of these species. Data generated from this random sequencing of the L. calcarifer genome should serve as a valuable resource for further studies of this organism.
    Matched MeSH terms: Plasmids
  12. Fulltext Antibiotic susceptibility profile of Haemophilus influenzae and transfer of co-trimoxazole resistance determinants
    Mohd-Zain Z, Kamsani NH, Ismail IS, Ahmad N
    Trop Biomed, 2012 Sep;29(3):372-80.
    PMID: 23018500 MyJurnal
    Prior to the implementation of Haemophilus influenzae type b vaccination worldwide, H. influenzae has been one of the main causative agents of community acquired pneumonia and meningitis in children. Due to the lack of information on the characteristics of the H. influenzae isolates that have previously been collected in Malaysia, the H. influenzae were assessed of their microbial susceptibility to commonly used antibiotics. Emphasis was made on strains that were resistance to co-trimoxazole (SXT) and their mode of transfer of the antibiotic resistance determinants were examined. A collection of 34 H. influenzae isolates was serotyped and antimicrobial susceptibility tests were performed to 11 antibiotics. To the isolates that were found to be resistant to co-trimoxazole, minimum inhibition concentration (MIC) to SXT was performed using Etest while agar dilution method was used to measure the individual MICs of trimethoprim (TMP) and sulfamethoxazole (SUL). These isolates were also examined for presence of plasmid by PCR and isolation method. Conjugal transfers of SXT-resistant genes to SXT-susceptible hosts were performed to determine their rate of transfer. Result showed that 20.6% of the total number of isolates was serotype B while the remaining was non-typeable. Antimicrobial susceptibility profile of all the isolates revealed that 58.8% was resistant to at least one antibiotic. Majority of these isolates were equally resistant to ampicillin and tetracycline (29.4% each), followed by resistance to SXT (26.5%). From nine isolates that were found to be SXT-resistant, five contained plasmid/s. Conjugal transfer experiment showed that these five isolates with plasmid transferred SXT-resistance determinants at a higher frequency than those without. From these observations, it is postulated that plasmid is not involved in the transfer of SXT-resistance genes but presence of plasmid facilitates their transfer. The information obtained from this study provides some basic knowledge on the antimicrobial susceptibility pattern of the H. influenzae isolates and their mode of transfer of SXT-resistance genes.
    Matched MeSH terms: Plasmids/genetics
  13. Culture-independent-based analyses identified site-specific microbial in two oil-wells from Malaysia
    Cheng-Yee Fish-Low, Chee HY, Ainon Hamzah
    Sains Malaysiana, 2015;44:1625-1633.
    Microbial communities of two oil reservoirs from Malaysia, denoted as Platform Bo and Platform Pe were studied using
    culture-independent approach. Environmental DNA was extracted and the universal amplified ribosomal region (UARR)
    was target amplified for both prokaryotes and eukaryotes. The amplified products were purified and cloned into pTZ57R/T
    vector to construct the 16S/18S rDNA library. Restriction endocucleases HhaI and MspI were used to screen the library.
    From that, 125 and 253 recombinant plasmid representative clones from Platform Bo and Platform Pe, respectively, were
    sent for DNA sequencing. Twenty-six operational taxonomic units (OTUs) consist of 20 genera detected at Platform Bo
    and 17 OTUs consist of 13 genera detected at Platform Pe. Marinobacter and Acinetobacter species co-occurred in both
    platforms whereas the rest are site-specific. Gammaproteobacteria accounted for 86.0% of the microbial community in
    Platform Bo, where OTUs affiliated to Marinobacter, Pseudomonas and Marinobacterium that were the most abundant. The
    major OTUs in the Platform Pe were with affinities to Achromobacter, followed by Stenotrophomonas and Serratia. The
    only archaeal isolates were detected in Platform Pe, which affiliated to Thermocladium. The singletons and doubletons
    accounted for about 50.0% of the OTU abundance in both platforms, which considerably significant despite their rare
    occurrence.
    Matched MeSH terms: Plasmids
  14. Fulltext Increased recovery and improved purity of PHA from recombinant Cupriavidus necator
    Anis SN, Iqbal NM, Kumar S, Al-Ashraf A
    Bioengineered, 2013 Mar-Apr;4(2):115-8.
    PMID: 23018620 DOI: 10.4161/bioe.22350
    A simple procedure for recovering biodegradable polymer from bacterial cells has been developed using economical and environmentally friendly solvent or chemicals. Recombinant bacterium, Cupriavidus necator harboring pBBR1MCS-C2 plasmid polyhydroxyalkanoate (PHA) synthase gene was used for the production of copolymer P(3HB-co-3HHx) from crude palm kernel oil (CPKO). NaOH was chosen in this study as it could give high purity and recovery yield. Increase of NaOH concentration had resulted in an increase of the PHA purity, but the recovery yield had decreased. The greater improvement of PHA purity and recovery were achieved by incubating the freeze-dried cells (10-30 g/L) in NaOH (0.1 M) for 1-3 h at 30°C and polishing using 20% (v/v) of ethanol. The treatment caused negligible degradation of the molecular weight of PHA recovered from the bacterial cells. The present review also highlights other extraction methods to provide greater insights into economical and sustainable recovery of PHA from bacterial cells.
    Matched MeSH terms: Plasmids/genetics
  15. Overcoming the challenge of transduction of human T-cells with chimeric antigen receptor (CAR) specific for ERBB2 antigen
    Munisvaradass R, Ding SSL, Ee AHK, Syahril Abdullah, Mok PL, Kumar S, et al.
    Sains Malaysiana, 2017;46:1831-1838.
    Breast cancer is one of the most common malignancies among woman. Decades of scientific study have linked the
    overexpression of ERBB2 antigen to aggressive tumors. To target aggressive breast cancer, chimeric antigen receptor
    (CAR) technology can be utilized. For this, human T-cells are transduced with a gene sequence encoding a CAR that is
    specific for tumor-associated antigens (TAAs). These genetically-engineered CAR transduced T-cells (CAR-T cells) are
    able to target the tumor antigen without the need for major histocompatibility complex (MHC) recognition, rendering
    it a potentially universal immunotherapeutic option. However, efficient transduction of therapeutic gene into human
    T-cells and further cell expansion are challenging. In this study, we reported a successful optimization of a transduction
    protocol using spinoculation on CD3+ T-cells with different concentrations of lentiviral plasmid encoding the CAR gene.
    CD3+T-cells were isolated from the peripheral blood mononuclear cells (PBMCs). The constructed CAR gene was inserted
    into a lentiviral plasmid containing the green fluorescent protein (GFP) tag and lentiviral particles were produced. These
    lentiviral particles were used to transduce activated T-cells by spinoculation. T-cells were activated using Dynabeadconjugated
    CD3/CD28 human T-cell activator and interleukin-2 (IL-2) before transduction. CD3+ T-cells were selected
    and GFP expression, which indicated transduction, was observed. Future studies will focus on in vitro and in vivo models
    to determine the efficiency of CAR-T cells in specifically targeting ERBB2-expressing cells.
    Matched MeSH terms: Plasmids
  16. Fulltext Engineering and Validation of a Vector for Concomitant Expression of Rare Transfer RNA (tRNA) and HIV-1 nef Genes in Escherichia coli
    Mualif SA, Teow SY, Omar TC, Chew YW, Yusoff NM, Ali SA
    PLoS One, 2015;10(7):e0130446.
    PMID: 26147991 DOI: 10.1371/journal.pone.0130446
    Relative ease in handling and manipulation of Escherichia coli strains make them primary candidate to express proteins heterologously. Overexpression of heterologous genes that contain codons infrequently used by E. coli is related with difficulties such as mRNA instability, early termination of transcription and/or translation, deletions and/or misincorporation, and cell growth inhibition. These codon bias -associated problems are addressed by co-expressing ColE1-compatible, rare tRNA expressing helper plasmids. However, this approach has inadequacies, which we have addressed by engineering an expression vector that concomitantly expresses the heterologous protein of interest, and rare tRNA genes in E. coli. The expression vector contains three (argU, ileY, leuW) rare tRNA genes and a useful multiple cloning site for easy in-frame cloning. To maintain the overall size of the parental plasmid vector, the rare tRNA genes replaced the non-essential DNA segments in the vector. The cloned gene is expressed under the control of T7 promoter and resulting recombinant protein has a C-terminal 6His tag for IMAC-mediated purification. We have evaluated the usefulness of this expression vector by expressing three HIV-1 genes namely HIV-1 p27 (nef), HIV-1 p24 (ca), and HIV-1 vif in NiCo21(DE3) E.coli and demonstrated the advantages of using expression vector that concomitantly expresses rare tRNA and heterologous genes.
    Matched MeSH terms: Plasmids/genetics
  17. Fulltext Production and purification of polymerization-competent HIV-1 capsid protein p24 (CA) in NiCo21(DE3) Escherichia coli
    Teow SY, Mualif SA, Omar TC, Wei CY, Yusoff NM, Ali SA
    BMC Biotechnol, 2013;13:107.
    PMID: 24304876 DOI: 10.1186/1472-6750-13-107
    HIV genome is packaged and organized in a conical capsid, which is made up of ~1,500 copies of the viral capsid protein p24 (CA). Being a primary structural component and due to its critical roles in both late and early stages of the HIV replication cycle, CA has attracted increased interest as a drug discovery target in recent years. Drug discovery studies require large amounts of highly pure and biologically active protein. It is therefore desirable to establish a simple and reproducible process for efficient production of HIV-1 CA.
    Matched MeSH terms: Plasmids/genetics
  18. Fulltext Engineering an L-cell line that expresses insulin under the control of the glucagon-like peptide-1 promoter for diabetes treatment
    Rasouli M, Ahmad Z, Omar AR, Allaudin ZN
    BMC Biotechnol, 2011 Nov 03;11:99.
    PMID: 22047106 DOI: 10.1186/1472-6750-11-99
    BACKGROUND: Diabetes mellitus is a complicated disease with a pathophysiology that includes hyperinsulinemia, hyperglycemia and other metabolic impairments leading to many clinical complications. It is necessary to develop appropriate treatments to manage the disease and reduce possible acute and chronic side effects. The advent of gene therapy has generated excitement in the medical world for the possible application of gene therapy in the treatment of diabetes. The glucagon-like peptide-1 (GLP-1) promoter, which is recognised by gut L-cells, is an appealing candidate for gene therapy purposes. The specific properties of L-cells suggest that L-cells and the GLP-1 promoter would be useful for diabetes therapy approaches.

    RESULTS: In this study, L-cells were isolated from a primary intestinal cell line to create suitable target cells for insulin expression studies. The isolated cells displayed L-cell properties and were therefore used as an L-cell surrogate. Next, the isolated L-cells were transfected with the recombinant plasmid consisting of an insulin gene located downstream of the GLP-1 promoter. The secretion tests revealed that an increase in glucose concentration from 5 mM to 25 mM induced insulin gene expression in the L-cells by 2.7-fold. Furthermore, L-cells quickly responded to the glucose stimulation; the amount of insulin protein increased 2-fold in the first 30 minutes and then reached a plateau after 90 minutes.

    CONCLUSION: Our data showed that L-cells efficiently produced the mature insulin protein. In addition, the insulin protein secretion was positively regulated with glucose induction. In conclusion, GLP-1 promoter and L-cell could be potential candidates for diabetes gene therapy agents.

    Matched MeSH terms: Plasmids/genetics*; Plasmids/chemistry
  19. Enhanced recovery and purification of P(3HB-co-3HHx) from recombinant Cupriavidus necator using alkaline digestion method
    Anis SN, Nurhezreen MI, Sudesh K, Amirul AA
    Appl Biochem Biotechnol, 2012 Jun;167(3):524-35.
    PMID: 22569781 DOI: 10.1007/s12010-012-9677-9
    A simple, efficient and economical method for the recovery of P(3HB-co-3HHx) was developed using various chemicals and parameters. The initial content of P(3HB-co-3HHx) in bacterial cells was 50-60 wt%, whereas the monomer composition of 3HHx used in this experiments was 3-5 mol%. It was found that sodium hydroxide (NaOH) was the most effective chemical for the recovery of biodegradable polymer. High polyhydroxyalkanoate purity and recovery yield both in the range of 80-90 wt% were obtained when 10-30 mg/ml of cells were incubated in NaOH at the concentration of 0.1 M for 60-180 min at 30 °C and polished using 20 % (v/v) of ethanol.
    Matched MeSH terms: Plasmids/genetics
  20. Epidemiology and molecular characterization of nosocomially transmitted multidrug-resistant Klebsiella pneumoniae
    Parasakthi N, Vadivelu J, Ariffin H, Iyer L, Palasubramaniam S, Arasu A
    Int J Infect Dis, 2000;4(3):123-8.
    PMID: 11179914
    OBJECTIVES: To describe the epidemiology, antimicrobial susceptibility, genomic profiles, and control of a nosocomial outbreak of multidrug-resistant Klebsiella pneumoniae (MRKP) that occurred in the pediatric oncology unit of the University of Malaya Medical Centre in Kuala Lumpur.

    MATERIALS AND METHODS: A prospective epidemiologic and microbiologic study was conducted of MRKP isolated from the blood and wound of a boy with necrotizing fasciitis after a 7-day course of ceftazidime and amikacin. In the following 2 weeks, phenotypically similar MRKP were isolated from the blood cultures of four other patients and rectal swabs of another three patients and two liquid soap samples located in the same ward.

    RESULTS: Antimicrobial profiles demonstrated that all the isolates were resistant to ceftazidime, sensitive to imipenem and ciprofloxacin, and confirmed to be extended-spectrum beta-lactamase producers. Plasmids of varying molecular weights were present in all isolates. In eight of these isolates, which included four from blood, there were common large molecular weight plasmids ranging from 80 kb to 100 kb. Pulsed-field gel electrophoresis analysis using XbaI demonstrated six different DNA profiles, A to F. Profile A was shared by two blood culture isolates and were related by 91%. Profile B was found in one rectal swab isolate and one isolate from liquid soap and were related by 94%. Profile C was shared by one blood isolate and one liquid soap isolate and showed 100% relatedness. Profiles D, E, and F each were demonstrated by one blood isolate and two rectal swab isolates, respectively. These showed only 65% relatedness.

    CONCLUSIONS: The MRKP strains in this outbreak were not clonal in origin. The decline of the outbreak after 4 weeks was attributed to the reemphasis of standard infection control procedures and the implementation of a program that addressed sites of environmental contamination.

    Matched MeSH terms: Plasmids/genetics
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