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91.
Combined inactivation and expression strategy to study gene function under physiological conditions: application to identification of new Escherichia coli adhesins 下载免费PDF全文
In bacteria, whereas disruption methods have been improved recently, the use of plasmid complementation strategies are still subject to limitations, such as cloning difficulties, nonphysiological levels of gene expression, or a requirement for antibiotics as plasmid selection pressure. Moreover, because of the pleiotropic modifications of cell physiology often induced by plasmid-based complementation, these strategies may introduce biases when biological process such as adhesion or biofilm formation are studied. We developed a plasmid-free approach that combines the lambda-red linear DNA recombination method with site-directed insertion of a repression and expression (RExBAD) cassette which places a functional pBAD promoter upstream of a target gene. We showed that this method permits both inactivation and modulation of most Escherichia coli gene expression, including expression of toxin and essential genes. We used this strategy to study adhesion and bacterial biofilms by placing the RExBAD cassette in front of the tra operon, encoding the DNA transfer and pilus genes on the F conjugative plasmid, and in front of flu, the antigen 43 (Ag43) autotransporter adhesin-encoding gene. In silico analysis revealed the existence of 10 genes with homology to the Ag43 gene that were good candidates for genes that encode putative new adhesins in E. coli. We used the RExBAD strategy to study these genes and demonstrated that induction of expression of four of them is associated with adhesion of E. coli to abiotic surfaces. The potential use of the RExBAD approach to study the function of cryptic or uncharacterized genes in large-scale postgenomic functional analyses is discussed. 相似文献
92.
Ga?l Chambonnier Lorène Roux David Redelberger Firas Fadel Alain Filloux Melissa Sivaneson Sophie de Bentzmann Christophe Bordi 《PLoS genetics》2016,12(5)
In response to environmental changes, Pseudomonas aeruginosa is able to switch from a planktonic (free swimming) to a sessile (biofilm) lifestyle. The two-component system (TCS) GacS/GacA activates the production of two small non-coding RNAs, RsmY and RsmZ, but four histidine kinases (HKs), RetS, GacS, LadS and PA1611, are instrumental in this process. RetS hybrid HK blocks GacS unorthodox HK autophosphorylation through the formation of a heterodimer. PA1611 hybrid HK, which is structurally related to GacS, interacts with RetS in P. aeruginosa in a very similar manner to GacS. LadS hybrid HK phenotypically antagonizes the function of RetS by a mechanism that has never been investigated. The four sensors are found in most Pseudomonas species but their characteristics and mode of signaling may differ from one species to another. Here, we demonstrated in P. aeruginosa that LadS controls both rsmY and rsmZ gene expression and that this regulation occurs through the GacS/GacA TCS. We additionally evidenced that in contrast to RetS, LadS signals through GacS/GacA without forming heterodimers, either with GacS or with RetS. Instead, we demonstrated that LadS is involved in a genuine phosphorelay, which requires both transmitter and receiver LadS domains. LadS signaling ultimately requires the alternative histidine-phosphotransfer domain of GacS, which is here used as an Hpt relay by the hybrid kinase. LadS HK thus forms, with the GacS/GacA TCS, a multicomponent signal transduction system with an original phosphorelay cascade, i.e. H1LadS→D1LadS→H2GacS→D2GacA. This highlights an original strategy in which a unique output, i.e. the modulation of sRNA levels, is controlled by a complex multi-sensing network to fine-tune an adapted biofilm and virulence response. 相似文献
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A 3beta-hydroxysteroid dehydrogenase (3betaHSD) was demonstrated in term human fetal membranes (chorion and amnion) with both dehydroepiandrosterone (3beta-hydroxy-5-androsten-17-one) and pregnenolone (3beta-hydroxy-5-pregnen-20-one as substrates, and the subcellular distribution substrate and nucleotide specificity of the enzyme was studied. In both membranes the microsomal fraction (particles which sedimented at 105,000 g after 90 min) had the highest specific activity. The chorion was more active than the amnion but the enzyme in both tissues had similar substrate and nucleotide specificity. NAD was the preferred cofactor, and pregnenolone was a better substrate than dehydroepiandrosterone in the presence of NAD. However, with NADP as cofactor both steroids were equally good substrates. When the 3beta-hydroxysteroid dehydrogenase activity of chorion microsomes was compared with that of placental microsomes, the specific activities were found to be of the same order of magnitude, and the substrate, nucleotide specificity and steroid binding properties were almost identical. 相似文献
95.
Stefan Haffke Barbara Kusterer F. Joachim Fromme Steffen Roux Bernd Hackauf Thomas Miedaner 《Bioenergy Research》2014,7(1):424-429
Winter rye (Secale cereale L.) is becoming increasingly important as substrate for biogas production in Central Europe. Dry matter yield has evolved as a breeding goal comparably important to the traditional grain yield. We analyzed the covariation between both traits and tested other agronomic traits for their correlation to dry matter yield that could be used for prediction of biomass yield. A set of 258 experimental hybrids were tested for dry matter yield harvested at late milk stage and grain yield harvested at full ripening at three to four locations in Germany in 2011 and 2012. We observed a wide range of dry matter yield (10–24 Mg ha?1) and grain yield (6–15 Mg ha?1) among testcross progenies. Genetic variances were significantly (P?<?0.01) different from zero for all traits. High entry-mean heritabilities (0.92–0.94) were found for plant height measurements and moderate heritabilities for grain and dry matter yield (0.52 and 0.49, respectively). Relative efficiencies for selection of dry matter yield estimated by second (EC 51–55) and third (EC 73) measurements of plant height were 1.24 and 0.98 respectively, compared to 0.52 for grain yield. Indirect selection for high dry matter yield using late plant height measurements should be successful. Using grain yield for indirect selection was less effective. The observed broad genetic variation for biomass yield in elite hybrid rye gives good prospects for the use as a resource of renewable energy. Plant height is a good predictor of dry matter yield but should be selected together with improved lodging resistance. 相似文献
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97.
Mammal Research - Foraging animals must contend with fluctuating environmental variables that affect foraging success, including conditions like wind noise, which could diminish the usefulness of... 相似文献
98.
Genetic analysis shows low levels of hybridization between African wildcats (Felis silvestris lybica) and domestic cats (F. s. catus) in South Africa 下载免费PDF全文
Johannes J. Le Roux Llewellyn C. Foxcroft Marna Herbst Sandra MacFadyen 《Ecology and evolution》2015,5(2):288-299
Hybridization between domestic and wild animals is a major concern for biodiversity conservation, and as habitats become increasingly fragmented, conserving biodiversity at all levels, including genetic, becomes increasingly important. Except for tropical forests and true deserts, African wildcats occur across the African continent; however, almost no work has been carried out to assess its genetic status and extent of hybridization with domestic cats. For example, in South Africa it has been argued that the long‐term viability of maintaining pure wildcat populations lies in large protected areas only, isolated from human populations. Two of the largest protected areas in Africa, the Kgalagadi Transfrontier and Kruger National Parks, as well as the size of South Africa and range of landscape uses, provide a model situation to assess how habitat fragmentation and heterogeneity influences the genetic purity of African wildcats. Using population genetic and home range data, we examined the genetic purity of African wildcats and their suspected hybrids across South Africa, including areas within and outside of protected areas. Overall, we found African wildcat populations to be genetically relatively pure, but instances of hybridization and a significant relationship between the genetic distinctiveness (purity) of wildcats and human population pressure were evident. The genetically purest African wildcats were found in the Kgalagadi Transfrontier Park, while samples from around Kruger National Park showed cause for concern, especially combined with the substantial human population density along the park's boundary. While African wildcat populations in South Africa generally appear to be genetically pure, with low levels of hybridization, our genetic data do suggest that protected areas may play an important role in maintaining genetic purity by reducing the likelihood of contact with domestic cats. We suggest that approaches such as corridors between protected areas are unlikely to remain effective for wildcat conservation, as the proximity to human settlements around these areas is projected to increase the wild/domestic animal interface. Thus, large, isolated protected areas will become increasingly important for wildcat conservation and efforts need to be made to prevent introduction of domestic cats into these areas. 相似文献
99.
Gustison ML le Roux A Bergman TJ 《Philosophical transactions of the Royal Society of London. Series B, Biological sciences》2012,367(1597):1847-1859
Primates are intensely social and exhibit extreme variation in social structure, making them particularly well suited for uncovering evolutionary connections between sociality and vocal complexity. Although comparative studies find a correlation between social and vocal complexity, the function of large vocal repertoires in more complex societies remains unclear. We compared the vocal complexity found in primates to both mammals in general and human language in particular and found that non-human primates are not unusual in the complexity of their vocal repertoires. To better understand the function of vocal complexity within primates, we compared two closely related primates (chacma baboons and geladas) that differ in their ecology and social structures. A key difference is that gelada males form long-term bonds with the 2-12 females in their harem-like reproductive unit, while chacma males primarily form temporary consortships with females. We identified homologous and non-homologous calls and related the use of the derived non-homologous calls to specific social situations. We found that the socially complex (but ecologically simple) geladas have larger vocal repertoires. Derived vocalizations of geladas were primarily used by leader males in affiliative interactions with 'their' females. The derived calls were frequently used following fights within the unit suggesting that maintaining cross-sex bonds within a reproductive unit contributed to this instance of evolved vocal complexity. Thus, our comparison highlights the utility of using closely related species to better understand the function of vocal complexity. 相似文献
100.
Latka MH Fielding K Gray GE Bekker LG Nchabeleng M Mlisana K Nielson T Roux S Mkhize B Mathebula M Naicker N de Bruyn G Kublin J Churchyard GJ;HVTN Phambili study team 《PloS one》2012,7(4):e31387