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251.
Gwendoline Kint Kathleen AJ Sonck Geert Schoofs David De Coster Jos Vanderleyden Sigrid CJ De Keersmaecker 《BMC microbiology》2009,9(1):198
Background
Quorum sensing is a term describing a bacterial communication system mediated by the production and recognition of small signaling molecules. The LuxS enzyme, catalyzing the synthesis of AI-2, is conserved in a wide diversity of bacteria. AI-2 has therefore been suggested as an interspecies quorum sensing signal. To investigate the role of endogenous AI-2 in protein expression of the Gram-negative pathogen Salmonella enterica serovar Typhimurium (S. Typhimurium), we performed a 2D-DIGE proteomics experiment comparing total protein extract of wildtype S. Typhimurium with that of a luxS mutant, unable to produce AI-2. 相似文献252.
William J Holmes Richard AJ Darby Martin DB Wilks Rodney Smith Roslyn M Bill 《Microbial cell factories》2009,8(1):35-14
Background
The optimisation and scale-up of process conditions leading to high yields of recombinant proteins is an enduring bottleneck in the post-genomic sciences. Typical experiments rely on varying selected parameters through repeated rounds of trial-and-error optimisation. To rationalise this, several groups have recently adopted the 'design of experiments' (DoE) approach frequently used in industry. Studies have focused on parameters such as medium composition, nutrient feed rates and induction of expression in shake flasks or bioreactors, as well as oxygen transfer rates in micro-well plates. In this study we wanted to generate a predictive model that described small-scale screens and to test its scalability to bioreactors. 相似文献253.
254.
Use of Ichip for High-Throughput In Situ Cultivation of “Uncultivable” Microbial Species 总被引:1,自引:0,他引:1
D. Nichols N. Cahoon E. M. Trakhtenberg L. Pham A. Mehta A. Belanger T. Kanigan K. Lewis S. S. Epstein 《Applied and environmental microbiology》2010,76(8):2445-2450
One of the oldest unresolved microbiological phenomena is why only a small fraction of the diverse microbiological population grows on artificial media. The “uncultivable” microbial majority arguably represents our planet''s largest unexplored pool of biological and chemical novelty. Previously we showed that species from this pool could be grown inside diffusion chambers incubated in situ, likely because diffusion provides microorganisms with their naturally occurring growth factors. Here we utilize this approach and develop a novel high-throughput platform for parallel cultivation and isolation of previously uncultivated microbial species from a variety of environments. We have designed and tested an isolation chip (ichip) composed of several hundred miniature diffusion chambers, each inoculated with a single environmental cell. We show that microbial recovery in the ichip exceeds manyfold that afforded by standard cultivation, and the grown species are of significant phylogenetic novelty. The new method allows access to a large and diverse array of previously inaccessible microorganisms and is well suited for both fundamental and applied research.It has been known for over a century that the overwhelming majority of microbial species do not grow on synthetic media in vitro and remain unexplored (13, 32, 37, 39, 40, 43). The rRNA and metagenomics approaches demonstrated a spectacular diversity of these uncultivated species (11, 21, 25-27, 30, 36). Accessing this “missing” microbial diversity is of significant interest for both basic and applied sciences and has been recognized as one of the principal challenges for microbiology today (12, 29, 41). In recent years, technical advances in cultivation methodologies have recovered a diverse set of ecologically relevant species (1, 3, 5, 7, 15, 20, 24, 28, 33, 42). However, by and large the gap between microbial diversity in nature and that in culture collections remains unchanged, and most microbial phyla still have no cultivable representatives (25, 29). Earlier, we developed a novel method of in situ cultivation of environmental microorganisms inside diffusion chambers (15). The rationale for such an approach was that diffusion would provide cells inside the chamber with naturally occurring growth components and enable those species that grew in nature at the time of the experiment to also grow inside the diffusion chambers. Expectedly, this method yields a rate of microbial recovery many times larger than those of standard techniques. Even so, this method is laborious and does not allow an efficient, high-throughput isolation of microbial species en masse. This limits the method''s applicability, for example, in the drug discovery effort. Here we transform this methodology into a high-throughput technology platform for massively parallel cultivation of “uncultivable” species. Capitalizing on earlier microfluidics methods developed for microbial storage and screening (4, 16), we have designed and tested an isolation chip, or ichip for short, which consists of hundreds of miniature diffusion chambers. If each diffusion minichamber is loaded with a single cell, the resulting culture is monospecific. The ichip thus allows microbial growth and isolation into pure culture in one step. Here we demonstrate that cultivation of environmental microorganisms inside the ichip incubated in situ leads to a significantly increased colony count over that observed on synthetic media. Perhaps even more significantly, species grown in ichips are different from those registered in standard petri dishes and are highly novel. 相似文献
255.
256.
Phosphoglucose isomerase (PGI) allozymes were isolated from the wing-
polymorphic water strider, Limnoporus canaliculatus, and were characterized
biochemically with respect to temperature-dependent kinetic and
thermostability properties. At higher temperatures, the allozymes exhibited
significant differences in Michaelis constant (Km) values for substrates of
both the forward and reverse reaction directions. Results were consistent
with expectations of adaptive kinetic differentiation based on the
latitudinal variation of PGI allele frequencies. PGI genotypes also
differed with regard to maximal velocity (Vmax)/Km ratios at higher
temperatures. These differences were due primarily, if not exclusively, to
allozyme-dependent variation in Km values. The allozymes also exhibited
dramatic differences in thermostability. However, no thermostability
differences were observed when the substrate analogue 6-phosphogluconate
was present in the incubation medium. The data from this study, together
with data from Mytilus edulis and Metridium senile on temperature-dependent
kinetic variation among PGI allozymes, form a consistent picture of natural
selection influencing the clinal variation of alleles at this locus in
these three phylogenetically distant organisms. More definitive support of
this hypothesis, however, must await additional studies on the
physiological effects of the allozymic variation as well as direct
measurements of fitness differences among the enzyme genotypes.
相似文献
257.
Rjean Carmichael Alain Belanger Simon Caron Fernand Labrie M. R. Sairam 《Steroids》1982,40(6):641-649
In order to better understand the effects of LHRH administration on testicular function in adult rat, we compared the inhibitory effects of LH and the LHRH analogue [D-Ser-(TBU)6, des-Gly-NH2 10]LHRH ethylamide upon testicular steroidogenesis and LH, FSH and prolactin receptor contents. Administration of LH as well as LHRH analogue resulted in a marked decrease of LH receptor levels, accompanied by a blockage at the level of 17-hydroxylase activity. We have been able to demonstrate that multiple LH administration can achieve a testicular desensitization comparable to that observed after LHRH agonist treatment. 相似文献
258.
J W Bacus M G Belanger R K Aggarwal F E Trobaugh 《The journal of histochemistry and cytochemistry》1976,24(1):195-201
Digital image processing and pattern recognition techniques were applied to determine the feasibility of a natural n-space subgrouping of normal and abnormal peripheral blood erythrocytes into well separated categories. The data consisted of 325 digitized red cells from 11 different cell classes. The analysis resulted in five features: (a) size, (b) roundness, (c) spicularity, (d) eccentricity and (e) central gray level distribution. These features separated the data into six distinct condensed subgroups of red cells. Each subgroup consisted of morphologically similar cells: (a) macrocytes, (b) normocytes, (c) schistocytes, acanthocytes and burr cells, (d) microcytes and spherocytes, (e) elliptocytes, sickle cells and pencil forms and (f) target cells. The concept of a quantitative "red cell differential" was introduced, utilizing these subgroup definitions to establish subpopulations of red cells, with quantifiable indices for the diagnosis of anemia, at the specimen level. 相似文献
259.
260.
Evidence for the thiamine biosynthetic pathway in higher-plant plastids and its developmental regulation 总被引:1,自引:0,他引:1
Faith C. Belanger Thomas Leustek Boyang Chu Alan L. Kriz 《Plant molecular biology》1995,29(4):809-821
Thiamine or vitamin B-1, is an essential constituent of all cells since it is a cofactor for two enzyme complexes involved in the citric acid cycle, pyruvate dehydrogenase and -ketoglutarate dehydrogenase. Thiamine is synthesized by plants, but it is a dietary requirement for humans and other animals. The biosynthetic pathway for thiamine in plants has not been well characterized and none of the enzymes involved have been isolated. Here we report the cloning and characterization of two cDNAs representing members of the maize thi1 gene family encoding an enzyme of the thiamine biosynthetic pathway. This assignment was made based on sequence homology to a yeast thiamine biosynthetic gene and by functional complementation of a yeast strain in which the endogenous gene was inactivated. Using immunoblot analysis, the thi1 gene product was found to be located in a plastid membrane fraction. RNA gel blot analysis of various tissues and developmental stages indicated thi1 expression was differentially regulated in a manner consistent with what is known about thiamine synthesis in plants. This is the first report of cDNAs encoding proteins involved in thiamine biosynthesis for any plant species. 相似文献