MyMedR

Displaying publications 81 - 100 of 224 in total

Abstract:

Sort:

Fulltext Diagnosis and potential treatments for acute hepatopancreatic necrosis disease (AHPND): a review

Santos HM, Tsai CY, Maquiling KRA, Tayo LL, Mariatulqabtiah AR, Lee CW, et al.

Aquac Int, 2020;28(1):169-185.
PMID: 32834683 DOI: 10.1007/s10499-019-00451-w

Acute hepatopancreatic necrosis disease (AHPND) or formerly known as early mortality syndrome (EMS) is an emerging disease that has caused significant economic losses to the aquaculture industry. The primary causative agent of AHPND is Vibrio parahaemolyticus, a Gram-negative rod-shaped bacterium that has gained plasmids encoding the fatal binary toxins Pir A/Pir B that cause rapid death of the infected shrimp. In this review, the current research studies and information about AHPND in shrimps have been presented. Molecular diagnostic tools and potential treatments regarding AHPND were also included. This review also includes relevant findings which may serve as guidelines that can help for further investigation and studies on AHPND or other shrimp diseases.

Matched MeSH terms: Plasmids
Fulltext Differentiation of chromoplasts and other plastids in plants

Sadali NM, Sowden RG, Ling Q, Jarvis RP

Plant Cell Rep, 2019 Jul;38(7):803-818.
PMID: 31079194 DOI: 10.1007/s00299-019-02420-2

Plant cells are characterized by a unique group of interconvertible organelles called plastids, which are descended from prokaryotic endosymbionts. The most studied plastid type is the chloroplast, which carries out the ancestral plastid function of photosynthesis. During the course of evolution, plastid activities were increasingly integrated with cellular metabolism and functions, and plant developmental processes, and this led to the creation of new types of non-photosynthetic plastids. These include the chromoplast, a carotenoid-rich organelle typically found in flowers and fruits. Here, we provide an introduction to non-photosynthetic plastids, and then review the structures and functions of chromoplasts in detail. The role of chromoplast differentiation in fruit ripening in particular is explored, and the factors that govern plastid development are examined, including hormonal regulation, gene expression, and plastid protein import. In the latter process, nucleus-encoded preproteins must pass through two successive protein translocons in the outer and inner envelope membranes of the plastid; these are known as TOC and TIC (translocon at the outer/inner chloroplast envelope), respectively. The discovery of SP1 (suppressor of ppi1 locus1), which encodes a RING-type ubiquitin E3 ligase localized in the plastid outer envelope membrane, revealed that plastid protein import is regulated through the selective targeting of TOC complexes for degradation by the ubiquitin-proteasome system. This suggests the possibility of engineering plastid protein import in novel crop improvement strategies.

Matched MeSH terms: Plasmids/genetics
Fulltext Disruption of adeB gene has a greater effect on resistance to meropenems than adeA gene in Acinetobacter spp. isolated from University Malaya Medical Centre

Wong EW, Yusof MY, Mansor MB, Anbazhagan D, Ong SY, Sekaran SD

Singapore Med J, 2009 Aug;50(8):822-6.
PMID: 19710984

The AdeABC pump of Acinetobacter spp. confers resistance to various antibiotic classes. This pump is composed of the AdeA, AdeB, and AdeC proteins where AdeB is a member of the resistance-nodulation-division efflux pump superfamily. The adeA, adeB, and adeC genes are contiguous and adjacent to adeS and adeR, which are transcribed in the opposite direction and which specify proteins homologous to sensors and regulators of two-component systems, respectively. In this study, an attempt is made to elucidate the role of the AdeABC efflux pump in carbapenem resistance in Acinetobacter spp.

Matched MeSH terms: Plasmids/metabolism
Diverse Plasmids Harboring blaCTX-M-15 in Klebsiella pneumoniae ST11 Isolates from Several Asian Countries

Kim SY, Ko KS

Microb Drug Resist, 2019 Mar;25(2):227-232.
PMID: 30212274 DOI: 10.1089/mdr.2018.0020

To reveal whether an increase of CTX-M-15-producing Klebsiella pneumoniae ST11 isolates is due to clonal dissemination across the countries, plasmids (pHK02-026, pM16-13, pIN03-01, and pTH02-34) were extracted from four K. pneumoniae isolates collected in Hong Kong, Malaysia, Thailand, and Indonesia, respectively. Complete sequencing of blaCTX-M-15-carrying plasmids was performed. In addition to the four plasmids, a previously sequenced plasmid (pKP12226) of a K. pneumoniae ST11 isolate from Korea was included in the analysis. While pIN03-01 and pTH02-34, which belonged to the incompatibility group IncX3, showed nearly the same structure, the others of IncF1A or IncFII exhibited very different structures. The number and kinds of antibiotic genes found in the plasmids were also different from each other. Cryptic prophage genes were identified in all five blaCTX-M-15-harboring plasmids from the ST11 isolates; P1-like region in pKP12226, CPZ-55 prophage region in pHK02-026, phage shock operon pspFABCD in pM16-13, and SPBc2 prophage yokD in pIN03-01 and pTH02-34. The plasmids with blaCTX-M-15 in the prevailing K. pneumoniae ST11 isolates in Asian countries might emerge from diverse origins by recombination. The prevalence of CTX-M-15-producing K. pneumoniae ST11 clone in Asian countries is not mainly due to the dissemination of a single strain.

Matched MeSH terms: Plasmids
Fulltext Doubly Spliced RNA of Hepatitis B Virus Suppresses Viral Transcription via TATA-Binding Protein and Induces Stress Granule Assembly

Tsai KN, Chong CL, Chou YC, Huang CC, Wang YL, Wang SW, et al.

J Virol, 2015 Nov;89(22):11406-19.
PMID: 26339052 DOI: 10.1128/JVI.00949-15

The risk of liver cancer in patients infected with the hepatitis B virus (HBV) and their clinical response to interferon alpha therapy vary based on the HBV genotype. The mechanisms underlying these differences in HBV pathogenesis remain unclear. In HepG2 cells transfected with a mutant HBV(G2335A) expression plasmid that does not transcribe the 2.2-kb doubly spliced RNA (2.2DS-RNA) expressed by wild-type HBV genotype A, the level of HBV pregenomic RNA (pgRNA) was higher than that in cells transfected with an HBV genotype A expression plasmid. By using cotransfection with HBV genotype D and 2.2DS-RNA expression plasmids, we found that a reduction of pgRNA was observed in the cells even in the presence of small amounts of the 2.2DS-RNA plasmid. Moreover, ectopic expression of 2.2DS-RNA in the HBV-producing cell line 1.3ES2 reduced the expression of pgRNA. Further analysis showed that exogenously transcribed 2.2DS-RNA inhibited a reconstituted transcription in vitro. In Huh7 cells ectopically expressing 2.2DS-RNA, RNA immunoprecipitation revealed that 2.2DS-RNA interacted with the TATA-binding protein (TBP) and that nucleotides 432 to 832 of 2.2DS-RNA were required for efficient TBP binding. Immunofluorescence experiments showed that 2.2DS-RNA colocalized with cytoplasmic TBP and the stress granule components, G3BP and poly(A)-binding protein 1 (PABP1), in Huh7 cells. In conclusion, our study reveals that 2.2DS-RNA acts as a repressor of HBV transcription through an interaction with TBP that induces stress granule formation. The expression of 2.2DS-RNA may be one of the viral factors involved in viral replication, which may underlie differences in clinical outcomes of liver disease and responses to interferon alpha therapy between patients infected with different HBV genotypes.

Matched MeSH terms: Plasmids/genetics
Fulltext Draft Genome Sequence of a Vibrio parahaemolyticus Strain, KS17.S5-1, with Multiple Antibiotic Resistance Genes, Which Causes Acute Hepatopancreatic Necrosis Disease in Penaeus monodon in the West Coast of Peninsular Malaysia

Devadas S, Bhassu S, Christie Soo TC, Mohamed Iqbal SN, Yusoff FM, Shariff M

Microbiol Resour Announc, 2018 Jul;7(2).
PMID: 30533806 DOI: 10.1128/MRA.00829-18

We report the first draft genome sequence of a Vibrio parahaemolyticus strain (VpAHPND), which causes acute hepatopancreatic necrosis disease (AHPND) in Penaeus monodon. The strain has a pVA1-like plasmid carrying pirAvp and pirBvp genes. Whole-genome comparisons revealed >98% similarity to VpAHPND isolates from Thailand, Mexico, and Vietnam.

Matched MeSH terms: Plasmids
Fulltext Draft Genome Sequence of the Shrimp Pathogen Vibrio parahaemolyticus ST17.P5-S1, Isolated in Peninsular Malaysia

Devadas S, Bhassu S, Christie Soo TC, Yusoff FM, Shariff M

Microbiol Resour Announc, 2018 Sep;7(11).
PMID: 30533648 DOI: 10.1128/MRA.01053-18

We sequenced the genome of Vibrio parahaemolyticus strain ST17.P5-S1, isolated from Penaeus vannamei cultured in the east coast of Peninsular Malaysia. The strain contains several antibiotic resistance genes and a plasmid encoding the Photorhabdus insect-related (Pir) toxin-like genes, pirAvp and pirBvp, associated with acute hepatopancreatic necrosis disease (AHPND).

Matched MeSH terms: Plasmids
Dynamics of PEGylated-dextran-spermine nanoparticles for gene delivery to leukemic cells

Amini R, Jalilian FA, Abdullah S, Veerakumarasivam A, Hosseinkhani H, Abdulamir AS, et al.

Appl Biochem Biotechnol, 2013 Jun;170(4):841-53.
PMID: 23615733 DOI: 10.1007/s12010-013-0224-0

Leukemic cells are hard-to-transfect cell lines. Many transfection reagents which can provide high gene transfer efficiency in common adherent cell lines are not effective to transfect established blood cell lines or primary leukemic cells. This study aims to examine a new class of cationic polymer non-viral vector, PEGylated-dextran-spermine (PEG-D-SPM), to determine its ability to transfect the leukemic cells. Here, the optimal conditions of the complex preparation (PEG-D-SPM/plasmid DNA (pDNA)) were examined. Different weight-mixing (w/w) ratios of PEG-D-SPM/pDNA complex were prepared to obtain an ideal mixing ratio to protect encapsulated pDNA from DNase degradation and to determine the optimal transfection efficiency of the complex. Strong complexation between polymer and pDNA in agarose gel electrophoresis and protection of pDNA from DNase were detected at ratios from 25 to 15. Highest gene expression was detected at w/w ratio of 18 in HL60 and K562 cells. However, gene expression from both leukemic cell lines was lower than the control MCF-7 cells. The cytotoxicity of PEG-D-SPM/pDNA complex at the most optimal mixing ratios was tested in HL60 and K562 cells using MTS assay and the results showed that the PEG-D-SPM/pDNA complex had no cytotoxic effect on these cell lines. Spherical shape and nano-nature of PEG-D-SPM/pDNA complex at ratio 18 was observed using transmission electron microscopy. As PEG-D-SPM showed modest transfection efficiency in the leukemic cell lines, we conclude that further work is needed to improve the delivery efficiency of the PEG-D-SPM.

Matched MeSH terms: Plasmids/genetics; Plasmids/chemistry
Effect of induced expression of an antimicrobial peptide melittin on Chlamydia trachomatis and Mycoplasma hominis infections in vivo

Lazarev VN, Shkarupeta MM, Titova GA, Kostrjukova ES, Akopian TA, Govorun VM

Biochem Biophys Res Commun, 2005 Dec 16;338(2):946-50.
PMID: 16246304

A plasmid construct was designed in which the gene of antimicrobial peptide melittin is controlled by the tetracycline-responsive promoter of human cytomegalovirus, aided by a constitutively expressed trans-activator protein gene. Its vaginal administration and induction of melittin gene transcription with doxycycline markedly suppressed subsequent genital tract infection of mice by Mycoplasma hominis and Chlamydia trachomatis. At least half of the melittin-protected animals proved free of either pathogen within 3-4 weeks. Recombinant plasmids expressing genes of antimicrobial peptides hold much promise as agents for prevention and control of urogenital latent infections.

Matched MeSH terms: Plasmids/administration & dosage
Effect of signal peptides on the secretion of β-cyclodextrin glucanotransferase in Lactococcus lactis NZ9000

Subramaniam M, Baradaran A, Rosli MI, Rosfarizan M, Khatijah Y, Raha AR

J. Mol. Microbiol. Biotechnol., 2012;22(6):361-72.
PMID: 23295307 DOI: 10.1159/000343921

Cyclodextrin glucanotransferase (CGTase) is an extracellular enzyme which catalyzes the formation of cyclodextrin from starch. The production of CGTase using lactic acid bacterium is an attractive alternative and safer strategy to produce CGTase. In this study, we report the construction of genetically modified Lactococcus lactis strains harboring plasmids that secrete the Bacillus sp. G1 β-CGTase, with the aid of the signal peptides (SPs) SPK1, USP45 and native SP (NSP). Three constructed vectors, pNZ:NSP:CGT, pNZ:USP:CGT and pNZ:SPK1:CGT, were developed in this study. Each vector harbored a different SP fused to the CGTase. The formation of halo zones on starch plates indicated the production and secretion of β-CGTase by the recombinants. The expression of this enzyme is shown by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and zymogram analysis. A band size of ∼75 kDa corresponding to β-CGTase is identified in the intracellular and the extracellular environments of the host after medium modification. The replacement of glucose by starch in the medium was shown to induce β-CGTase production in L. lactis. Although β-CGTase production is comparatively low in NZ:SPK1:CGT, the SP SPK1 was shown to have higher secretion efficiency compared to the other SPs used in this study.

Matched MeSH terms: Plasmids
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
Fulltext Electrosprayed Alginate Nanoparticles as CRISPR Plasmid DNA Delivery Carrier: Preparation, Optimization, and Characterization

Alallam B, Altahhan S, Taher M, Mohd Nasir MH, Doolaanea AA

Pharmaceuticals (Basel), 2020 Jul 22;13(8).
PMID: 32707857 DOI: 10.3390/ph13080158

Therapeutic gene editing is becoming more feasible with the emergence of the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR-associated protein (Cas) system. However, the successful implementation of CRISPR/Cas9-based therapeutics requires a safe and efficient in vivo delivery of the CRISPR components, which remains challenging. This study presents successful preparation, optimization, and characterization of alginate nanoparticles (ALG NPs), loaded with two CRISPR plasmids, using electrospray technique. The aim of this delivery system is to edit a target gene in another plasmid (green fluorescent protein (GFP)). The effect of formulation and process variables were evaluated. CRISPR ALG NPs showed mean size and zeta potential of 228 nm and -4.42 mV, respectively. Over 99.0% encapsulation efficiency was achieved while preserving payload integrity. The presence of CRISPR plasmids in the ALG NPs was confirmed by Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy. The tests revealed that the nanoparticles were cytocompatible and successfully introduced the Cas9 transgene in HepG2 cells. Nanoparticles-transfected HepG2 was able to edit its target plasmid by introducing double-strand break (DSB) in GFP gene, indicating the bioactivity of CRISPR plasmids encapsulated in alginate nanoparticles. This suggests that this method is suitable for biomedical application in vitro or ex vivo. Future investigation of theses nanoparticles might result in nanocarrier suitable for in vivo delivery of CRISPR/Cas9 system.

Matched MeSH terms: Plasmids
Electrotransformation of thermophilic bacterium Caldimonas manganoxidans

Arai T, Aikawa S, Sudesh K, Kondo T, Kosugi A

J Microbiol Methods, 2022 01;192:106375.
PMID: 34793853 DOI: 10.1016/j.mimet.2021.106375

Caldimonas manganoxidans is a Gram-negative, thermophilic, bioplastic-producing bacterium that is a promising strain to overcome the drawbacks of existing bioplastic manufacturing methods. However, genetic manipulation of this species has not previously been studied. Here, we developed an optimized electrotransformation protocol for C. manganoxidans by screening conditions, including the bacterial growth phase, electroporation buffer, pulse strength, and recovery time. The optimized transformation protocol obtained (3.1 ± 0.78) × 108 colony-forming units/μg DNA of plasmid pBBR1MCS-2. High transformation efficiency was observed when using plasmid DNA isolated from C. manganoxidans. The DNA methylases of Escherichia coli did not affect the transformation efficiency of C. manganoxidans. The electrotransformation technique proposed here will be beneficial for the genetic manipulation of thermophilic Caldimonas species.

Matched MeSH terms: Plasmids/genetics
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
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
Fulltext Engineering the lactococcal mevalonate pathway for increased sesquiterpene production

Song AA, Abdullah JO, Abdullah MP, Shafee N, Othman R, Noor NM, et al.

FEMS Microbiol Lett, 2014 Jun;355(2):177-84.
PMID: 24828482 DOI: 10.1111/1574-6968.12469

Isoprenoids are a large, diverse group of secondary metabolites which has recently raised a renewed research interest due to genetic engineering advances, allowing specific isoprenoids to be produced and characterized in heterologous hosts. Many researches on metabolic engineering of heterologous hosts for increased isoprenoid production are focussed on Escherichia coli and yeasts. E. coli, as most prokaryotes, use the 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway for isoprenoid production. Yeasts on the other hand, use the mevalonate pathway which is commonly found in eukaryotes. However, Lactococcus lactis is an attractive alternative host for heterologous isoprenoid production. Apart from being food-grade, this Gram-positive prokaryote uses the mevalonate pathway for isoprenoid production instead of the MEP pathway. Previous studies have shown that L. lactis is able to produce sesquiterpenes through heterologous expression of plant sesquiterpene synthases. In this work, we analysed the gene expression of the lactococcal mevalonate pathway through RT-qPCR to successfully engineer L. lactis as an efficient host for isoprenoid production. We then overexpressed the mvk gene singly or co-expressed with the mvaA gene as an attempt to increase β-sesquiphellandrene production in L. lactis. It was observed that co-expression of mvk with mvaA doubled the amount of β-sesquiphellandrene produced.

Matched MeSH terms: Plasmids/genetics
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
Enhanced secretory production of hemolysin-mediated cyclodextrin glucanotransferase in Escherichia coli by random mutagenesis of the ABC transporter system

Low KO, Mahadi NM, Abdul Rahim R, Rabu A, Abu Bakar FD, Abdul Murad AM, et al.

J Biotechnol, 2010 Dec;150(4):453-9.
PMID: 20959127 DOI: 10.1016/j.jbiotec.2010.10.001

The hemolysin transport system was found to mediate the release of cyclodextrin glucanotransferase (CGTase) into the extracellular medium when it was fused to the C-terminal 61 amino acids of HlyA (HlyAs(61)). To produce an improved-secretion variant, the hly components (hlyAs, hlyB and hlyD) were engineered by directed evolution using error-prone PCR. Hly mutants were screened on solid LB-starch plate for halo zone larger than the parent strain. Through screening of about 1 × 10(4) Escherichia coli BL21(DE3) transformants, we succeeded in isolating five mutants that showed a 35-217% increase in the secretion level of CGTase-HlyAs(61) relative to the wild-type strain. The mutation sites of each mutant were located at HlyB, primarily along the transmembrane domain, implying that the corresponding region was important for the improved secretion of the target protein. In this study we describe the finding of novel site(s) of HlyB responsible for enhancing secretion of CGTase in E. coli.

Matched MeSH terms: Plasmids/genetics
Fulltext Enhancers of Agrobacterium-mediated transformation of Tibouchina semidecandra selected on the basis of GFP expression

Thau, Wilson Lym Yon, Henry, Erle Stanley, Janna Ong Abdullah

Trop Life Sci Res, 2010;21(2):-.
MyJurnal

Genetic engineering is a powerful tool for the improvement of plant traits. Despite reported successes in the plant kingdom, this technology has barely scratched the surface of the Melastomataceae family. Limited studies have led to some optimisation of parameters known to affect the transformation efficiency of these plants. The major finding of this study was to optimise the presence of selected enhancers [e.g., monosaccharides (D-glucose, D-galactose and D-fructose), tyrosine, aluminium chloride (AICI3) and ascorbic
acid] to improve the transformation efficiency of Tibouchina semidecandra. Agrobacterium tumefaciens strain LBA4404 harbouring the disarmed plasmid pCAMBIA1304 was used to transform shoots and nodes of T. semidecandra. Different concentrations of the transformation enhancers were tested by using green fluorescent protein (GFP) as a reporter. The results obtained were based on the percentage of GFP expression, which was observed 14 days post-transformation. A combination of 120 µM galactose and 100
µM tyrosine supplemented with 600 µM AICI3 in the presence of 15 mg/l ascorbic acid gave the highest percentage of positive transformants for T. semidecandra shoots. Whereas 60 µM galactose and 50 µM tyrosine with 200 µM AICI3 in the presence of 15 mg/l ascorbic acid was optimum for T. semidecandra nodes. The presence of the hygromycin phosphotransferase II (hptII) transgene in the genomic DNA of putative
T. semidecandra transformants was verified by PCR amplification with specific primers.

Matched MeSH terms: Plasmids
Enterobacteriaceae resistant to third-generation cephalosporins and quinolones in fresh culinary herbs imported from Southeast Asia

Veldman K, Kant A, Dierikx C, van Essen-Zandbergen A, Wit B, Mevius D

Int J Food Microbiol, 2014 May 2;177:72-7.
PMID: 24607424 DOI: 10.1016/j.ijfoodmicro.2014.02.014

Since multidrug resistant bacteria are frequently reported from Southeast Asia, our study focused on the occurrence of ESBL-producing Enterobacteriaceae in fresh imported herbs from Thailand, Vietnam and Malaysia. Samples were collected from fresh culinary herbs imported from Southeast Asia in which ESBL-suspected isolates were obtained by selective culturing. Analysis included identification by MALDI-TOF mass spectrometry, susceptibility testing, XbaI-PFGE, microarray, PCR and sequencing of specific ESBL genes, PCR based replicon typing (PBRT) of plasmids and Southern blot hybridization. In addition, the quinolone resistance genotype was characterized by screening for plasmid mediated quinolone resistance (PMQR) genes and mutations in the quinolone resistance determining region (QRDR) of gyrA and parC. The study encompassed fifty samples of ten batches of culinary herbs (5 samples per batch) comprising nine different herb variants. The herbs originated from Thailand (Water morning glory, Acacia and Betel leaf), Vietnam (Parsley, Asian pennywort, Houttuynia leaf and Mint) and Malaysia (Holy basil and Parsley). By selective culturing 21 cefotaxime resistant Enterobacteriaceae were retrieved. Array analysis revealed 18 isolates with ESBL genes and one isolate with solely non-ESBL beta-lactamase genes. Mutations in the ampC promoter region were determined in two isolates with PCR and sequencing. The isolates were identified as Klebsiella pneumoniae (n=9), Escherichia coli (n=6), Enterobacter cloacae complex (n=5) and Enterobacter spp. (n=1). All isolates tested were multidrug resistant. Variants of CTX-M enzymes were predominantly found followed by SHV enzymes. PMQR genes (including aac(6')-1b-cr, qnrB and qnrS) were also frequently detected. In almost all cases ESBL and quinolone resistance genes were located on the same plasmid. Imported fresh culinary herbs from Southeast Asia are a potential source for contamination of food with multidrug resistant bacteria. Because these herbs are consumed without appropriate heating, transfer to human bacteria cannot be excluded.

Matched MeSH terms: Plasmids/genetics

Filters

Please provide feedback to Administrator (afdal@afpm.org.my)

External Links