Mutation of recA gene of Vibrio cholerae: Towards the development of attenuated vibrio cholerae vaccine strains

Despite over hundred years of study, the intestinal pathogen Vibrio cholerae still causes epidemic outbreaks in areas of the world. The endemic areas include India, Asia, Africa, the Mediterranean, and more recently, South and Central America, Mexico and the United States (Faruque et a/, 1998). This Gram-negative bacterium cplonizes the human intestine and causes potentially fatal diarrhoeal disease- the Cholera. A number of genes associated with the virulence of Vibrios had so far been identified by researchers of cholera. These genes are gaining popularity among researchers especially in the manipulation of them towards the production of an effective cholera vaccine. In this research, however, the recA gene was the center of interest. It was reported by Ghosh et al (1985) that the Vibrio cholerae have a RecA system that is analogous to the Escherichia coli RecA system. The RecA protein is the central enzyme for homologous recombination, DNA strand exchange, and DNA repair mechanism (Kuzminov, 1999). In the development of live attenuated, oral cholera vaccine, the mutation of recA gene was strongly recommended by some researchers to render the vaccine strains recombination deficient (Goldberg and Mekalanos, 1986; Taylor et al, 1994; Kenner et al, 1995; Kaper et al, 1994; Boyd and Waldor, 1999). Hence, the general approach of study is to mutate the recA gene in existing vaccine strain (VCUSM-2) to stabilize and prevent the strain from further acquiring the undesirable gene when released to the environment. The study began with a deletional mutation in the 1.2kb Vibrio cholerae recA gene previously cloned onto the vector, pTOP02.1. The EstEll was recognized as the unique restriction enzyme site which existed twice in the middle of Vibrio cholerae recA gene at the location of 71 Obp and 782bp. Upon restriction with EstEll, a small fragment of gene with the Isize of 72bp was deleted. The sticky overhang was polished and this was followed by a selfblunt end ligation of the linearized plasmid. The deletional mutation of recA gene produced a + 1 frameshift mutation starting from the ligation junction to the down stream of recA gene. As a result a stop codon was introduced at 748bp of the gene, and hence results in a truncated recA gene product. The successfully mutated Vibrio cholerae recA gene was then subcloned onto the pBluescriptiiSK before it was transferred onto the pCVD442, a conjugatable suicide plasmid for conjugation with the cholera vaccine strain, VCUSM-2

Maternal mRNAs are regulated by diverse P body–related mRNP granules during early Caenorhabditis elegans development

Processing bodies (P bodies) are conserved mRNA–protein (mRNP) granules that are thought to be cytoplasmic centers for mRNA repression and degradation. However, their specific functions in vivo remain poorly understood. We find that repressed maternal mRNAs and their regulators localize to P body–like mRNP granules in the Caenorhabditis elegans germ line. Surprisingly, several distinct types of regulated granules form during oocyte and embryo development. 3′ untranslated region elements direct mRNA targeting to one of these granule classes. The P body factor CAR-1/Rap55 promotes association of repressed mRNA with granules and contributes to repression of Notch/glp-1 mRNA. However, CAR-1 controls Notch/glp-1 only during late oogenesis, where it functions with the RNA-binding regulators PUF-5, PUF-6, and PUF-7. The P body protein CGH-1/Rck/Dhh1 differs from CAR-1 in control of granule morphology and promotes mRNP stability in arrested oocytes. Therefore, a system of diverse and regulated RNP granules elicits stage-specific functions that ensure proper mRNA control during early development

Screening for intermediate and severe forms of thalassaemia in discarded red blood cells: optimization and feasibility

Detection and quantification of Hb subtypes of human blood is integral to presumptive identification of thalassaemias. It has been used in neonatal screening of thalassaemia and Hb variants. The use of discarded red blood cells following processing of the cord blood for stem cells provides readily available diagnostic material for thalassaemia screening. In this study, we determined the range of Hb subtypes in 195 consecutive cord blood samples collected for cord blood banking. The 'cord blood samples' analysed were those of the remaining red blood cells after the cord blood was processed for stem cell storage. Quantification of Hb subtypes by high performance liquid chromatography (HPLC) was done on BioRad Variant II Hb testing system. Only 73 (36.5%) of the samples could be analyzed neat without dilution. With a 1:300 dilution with wash solution the acceptable area as recommended by the manufacturer for reading of a C-gram within the 1 to 3 million ranges were achieved in all. Eighteen (9%) 12 showed classical Hb Barts (γ4) prerun peaks were confirmed by Sebia Hydrasys automated Hb gel electrophoresis and quantified by Sebia Capillarys 2 capillary electrophoresis. Only 1 (0.5%) was presumptively identified with HbH disease. Due to the limited number of samples no beta-thalassaemia major, Hb E beta-thalassaemia and Hb Barts hydrops fetalis were found. The HPLC assay was possible at a cost US$ 5 per sample and a turnover time of 10 samples per hour without technical difficulties. This study reports an effective and valuable protocol for thalassaemia screening in red blood cells which would otherwise be discarded during cord blood processing. Cord blood with severe and intermediate forms of thalassaemia can be preselected and not stored

Coordination complexes of thiazyl rings: Synthesis, structure and DFT computational analysis of CpCr(CO)x (x = 2,3) complexes of fluorinated and non-fluorinated 1lamda^3-1,2,4,6-thiatriazinyls, with differing Cr—S bond orders.

Sherpa Romeo green journal. Permission to archive accepted author manuscript.Reaction of [3,5-Ph2-C2N3S]2 with [CpCr(CO)3]2 in toluene at RT forms an adduct via a Cr—S bond, formulated as CpCr(CO)3SN3C2Ph2, which has fitting NMR, IR and combustion analysis data. The structure was determined by a single-crystal X-ray structure diffraction study (P21/n, a = 8.4611(17)Å; b = 20.509(4) Å, c = 11.757(2) Å, = 104.453(7)°). The Cr—S bond-length = 2.4908(11) Å corresponds to a bond-order of 1.0 from > 90 values for CpCr(CO)x or Cp*Cr(CO)x moieties (x = 2,3) bonded to S which are used to establish a Pauling-type bond order scale specific to this class of compounds. Similar reactions of fluorinated thiatriazinyls derived from [3-Ph-5-CF3-C2N3S]2 or [4-MeOC6H4-5-CF3-C2N3S]2 are accompanied by the loss of CO to produce CpCr(CO)2SN3C2PhCF3 (P, a = 8.0929(8) Å; b = 10.3160(10) Å, c = 11.2405(11), = 70.032(2)°, = 72.076(2)°, = 82.375(2)° ) and CpCr(CO)2SN3(CCF3)(C6H4OCH3) (P21/c, a = 8.1311(7)Å; b = 24.284(2) Å, c = 9.1025(8) Å, = 97.218(2)°), also fully characterized by spectroscopy and crystallography. Their measured Cr—S bond-lengths, 2.2987(14) Å and 2.2965(11) Å, correspond to bond orders of 1.5. (U/R)B3PW91/6-311+G(2df,2p)//B3PW91/6-31G(2d,p) hybrid DFT calculations show that the tricarbonyl complex has an unusual bond. However, the dicarbonyl complexes of the fluorinated thiatriazinyls are -bonded.Ye

Multiple mechanisms collectively regulate clathrin-mediated endocytosis of the epidermal growth factor receptor

Four independent mechanisms for uptake of activated EGFR are identified by a combination of receptor mutagenesis and RNA interference approaches

Differential cross section measurements for the production of a W boson in association with jets in proton–proton collisions at √s = 7 TeV

Measurements are reported of differential cross sections for the production of a W boson, which decays into a muon and a neutrino, in association with jets, as a function of several variables, including the transverse momenta (pT) and pseudorapidities of the four leading jets, the scalar sum of jet transverse momenta (HT), and the difference in azimuthal angle between the directions of each jet and the muon. The data sample of pp collisions at a centre-of-mass energy of 7 TeV was collected with the CMS detector at the LHC and corresponds to an integrated luminosity of 5.0 fb[superscript −1]. The measured cross sections are compared to predictions from Monte Carlo generators, MadGraph + pythia and sherpa, and to next-to-leading-order calculations from BlackHat + sherpa. The differential cross sections are found to be in agreement with the predictions, apart from the pT distributions of the leading jets at high pT values, the distributions of the HT at high-HT and low jet multiplicity, and the distribution of the difference in azimuthal angle between the leading jet and the muon at low values.United States. Dept. of EnergyNational Science Foundation (U.S.)Alfred P. Sloan Foundatio

Penilaian Kinerja Keuangan Koperasi di Kabupaten Pelalawan

This paper describe development and financial performance of cooperative in District Pelalawan among 2007 - 2008. Studies on primary and secondary cooperative in 12 sub-districts. Method in this stady use performance measuring of productivity, efficiency, growth, liquidity, and solvability of cooperative. Productivity of cooperative in Pelalawan was highly but efficiency still low. Profit and income were highly, even liquidity of cooperative very high, and solvability was good

Juxtaposing BTE and ATE – on the role of the European insurance industry in funding civil litigation

One of the ways in which legal services are financed, and indeed shaped, is through private insurance arrangement. Two contrasting types of legal expenses insurance contracts (LEI) seem to dominate in Europe: before the event (BTE) and after the event (ATE) legal expenses insurance. Notwithstanding institutional differences between different legal systems, BTE and ATE insurance arrangements may be instrumental if government policy is geared towards strengthening a market-oriented system of financing access to justice for individuals and business. At the same time, emphasizing the role of a private industry as a keeper of the gates to justice raises issues of accountability and transparency, not readily reconcilable with demands of competition. Moreover, multiple actors (clients, lawyers, courts, insurers) are involved, causing behavioural dynamics which are not easily predicted or influenced. Against this background, this paper looks into BTE and ATE arrangements by analysing the particularities of BTE and ATE arrangements currently available in some European jurisdictions and by painting a picture of their respective markets and legal contexts. This allows for some reflection on the performance of BTE and ATE providers as both financiers and keepers. Two issues emerge from the analysis that are worthy of some further reflection. Firstly, there is the problematic long-term sustainability of some ATE products. Secondly, the challenges faced by policymakers that would like to nudge consumers into voluntarily taking out BTE LEI

Search for stop and higgsino production using diphoton Higgs boson decays

Results are presented of a search for a "natural" supersymmetry scenario with gauge mediated symmetry breaking. It is assumed that only the supersymmetric partners of the top-quark (stop) and the Higgs boson (higgsino) are accessible. Events are examined in which there are two photons forming a Higgs boson candidate, and at least two b-quark jets. In 19.7 inverse femtobarns of proton-proton collision data at sqrt(s) = 8 TeV, recorded in the CMS experiment, no evidence of a signal is found and lower limits at the 95% confidence level are set, excluding the stop mass below 360 to 410 GeV, depending on the higgsino mass

Impacts of the Tropical Pacific/Indian Oceans on the Seasonal Cycle of the West African Monsoon

The current consensus is that drought has developed in the Sahel during the second half of the twentieth century as a result of remote effects of oceanic anomalies amplified by local land–atmosphere interactions. This paper focuses on the impacts of oceanic anomalies upon West African climate and specifically aims to identify those from SST anomalies in the Pacific/Indian Oceans during spring and summer seasons, when they were significant. Idealized sensitivity experiments are performed with four atmospheric general circulation models (AGCMs). The prescribed SST patterns used in the AGCMs are based on the leading mode of covariability between SST anomalies over the Pacific/Indian Oceans and summer rainfall over West Africa. The results show that such oceanic anomalies in the Pacific/Indian Ocean lead to a northward shift of an anomalous dry belt from the Gulf of Guinea to the Sahel as the season advances. In the Sahel, the magnitude of rainfall anomalies is comparable to that obtained by other authors using SST anomalies confined to the proximity of the Atlantic Ocean. The mechanism connecting the Pacific/Indian SST anomalies with West African rainfall has a strong seasonal cycle. In spring (May and June), anomalous subsidence develops over both the Maritime Continent and the equatorial Atlantic in response to the enhanced equatorial heating. Precipitation increases over continental West Africa in association with stronger zonal convergence of moisture. In addition, precipitation decreases over the Gulf of Guinea. During the monsoon peak (July and August), the SST anomalies move westward over the equatorial Pacific and the two regions where subsidence occurred earlier in the seasons merge over West Africa. The monsoon weakens and rainfall decreases over the Sahel, especially in August.Peer reviewe