Chemically-induced affinity star restriction specificity: a novel TspGWI/sinefungin endonuclease with theoretical 3-bp cleavage frequency

The type IIS/IIC restriction endonuclease TspGWI recognizes the sequence 5'-ACGGA-3', cleaving DNA 11/9 nucleotides downstream. Here we show that sinefungin, a cofactor analog of S-adenosyl methionine, induces a unique type of relaxation in DNA recognition specificity. In the presence of sinefungin, TspGWI recognizes and cleaves at least 12 degenerate variants of the original recognition sequence that vary by single base pair changes from the original 5-bp restriction site with only a single degeneracy per variant appearing to be allowed. In addition, sinefungin was found to have a stimulatory effect on cleavage at these nondegenerate TspGWI recognition sites, irrespective of their number or the DNA topology. Interestingly, no fixed "core" could be identified among the new recognition sequences. Theoretically, TspGWI cleaves DNA every 1024 bp, while sinefungin-induced activity cleaves every 78.8 bp, corresponding to a putative 3-bp long recognition site. Thus, the combination of sinefungin and TspGWI represents a novel frequent cutter, next only to CviJI/CviJI*, that should prove useful in DNA cloning methodologies.     

Bioplastics from microorganisms

The term 'biomaterials' includes chemically unrelated products that are synthesised by microorganisms (or part of them) under different environmental conditions. One important family of biomaterials is bioplastics. These are polyesters that are widely distributed in nature and accumulate intracellularly in microorganisms in the form of storage granules, with physico-chemical properties resembling petrochemical plastics. These polymers are usually built from hydroxy-acyl-CoA derivatives via different metabolic pathways. Depending on their microbial origin, bioplastics differ in their monomer composition, macromolecular structure and physical properties. Most of them are biodegradable and biocompatible, which makes them extremely interesting from the biotechnological point of view.     

Producing drugs from marine sponges

Marine sponges are potential sources of many unique metabolites, including cytotoxic and anticancer compounds. Natural sponge populations are insufficient or inaccessible for producing commercial quantities of metabolites of interest. This review focuses on methods of producing sponge biomass to overcome supply limitations. Production techniques discussed include aquaculture in the sea, the controlled environments of aquariums, and culture of sponge cells and primmorphs. Cultivation in the sea and aquariums are currently the only practicable and relatively inexpensive methods of producing significant quantities of sponge biomass. In the future, metabolite production from cultured sponge cells and primmorphs may become feasible. Obtaining a consistent biomass yield in aquariums requires attention to many factors that are discussed in this work.     

Hemisynthesis and preliminary evaluation of novel endocannabinoid analogues

Three new endocannabinoid analogues in which amide moiety was replaced either by oxomethylene group or ester moiety with simultaneous substitution of both alpha-hydrogens with methyl groups were synthesized and their abilities to interact with CB1-receptor and FAAH were investigated.     

In vitro reconstruction of the biosynthetic pathway of peptidoglycan cytoplasmic precursor in Pseudomonas aeruginosa

Bacterial peptidoglycan is the cell wall component responsible for maintaining cell integrity against osmotic pressure. Biosynthesis of the cytoplasmic precursor UDP-N-acetylmuramyl pentapeptide is catalyzed by the Mur enzymes. Genomic analysis of the three regions encoding Mur proteins was achieved. We have cloned and over-expressed the murA, -B, -D, -E and -F genes of Pseudomonas aeruginosa in pET expression system by adding a His-Tag to the C-termini of the proteins. Mur proteins were purified to homogeneity by a single chromatographic step on affinity nickel columns. Protein identities were verified through N-terminal sequencing. Enzyme activity was proved by the identification of the pathway's final product.     

Diatoms of Dindifelou fall (upper Basin of the Gambia River,Senegal): floristic inventory

The Dindifelou fall, located in south‐eastern Senegal, receives its water from discharging springs located in Dande plate‐land. These waters form, at the foot of the cliff, a pond that drains into the Thiokoye, a tributary of the Gambia River. The water quality of the pond and the stream is very good based on physicochemical analyses. Around the pond region, a microclimate characterized by a higher humidity and a lower temperature compared with the surrounding area is present. Diatoms study is carried out in different habitat types associated with this fall. The samples were taken by scraping the surface of rocks and rinsing fern leaves with sparkling water. Dande plate‐land is made of Upper Proterozoic sandstones whose gravity accumulations litter the bottom of the pond and the stream. This study allowed us to inventory 61 species and varieties of diatoms belonging to 28 genera. The most represented genera are Achnanthidium (ten species), Eunotia and Pinnularia (eight species each). Twelve species and varieties are reported for the first time in Senegambia including five of the genera Achnanthidium. The microflora is dominated by Achnantidium minutissimum and Achnanthidium catenatum, followed by Ulnaria ulna and various species of the genera Eunotia and Brachysira. Aerial habitats are more diversified here than the epilithic samples.     

Interspecific differences in root architecture among maize and triticale genotypes grown under drought,waterlogging and soil compaction

Environmental stresses (soil compaction, drought, waterlogging) cause changes in plants’ root system structure, also affecting the growth of above-ground parts. The aim of this study was to estimate phenotypic variation among maize and triticale genotypes in root penetration ability through petrolatum-wax-layer (RPA). Also, the effect of shortage or excess of soil water on dry matter of shoots and roots and morphological changes in root system structure in sensitive and resistant maize and triticale genotypes grown in low or high soil compaction level was evaluated. To estimate RPA index, the petrolatum-wax-layer method (PWL) was used. The strength of three petrolatum-wax concentrations 60, 50 and 40 % was 0.52, 1.07 and 1.58 MPa, respectively. High coefficients of variation (CV) were observed in 0.52 and 1.07 MPa and for maize were 19.2 and 21.7 %, and for triticale, 12.5 and 18.3 %, respectively. The data indicate that the use of PWL technique is an effective screening method, and makes it possible to divide the genotypes into resistant and sensitive groups. The second part of this study investigated a multistress effect of soil compaction combined with drought or waterlogging on root and shoot growth and morphological changes in root system structure of maize and triticale genotypes differing in susceptibility to environmental stresses. Seedlings were grown for 4 weeks in root-boxes under conditions of low (LSC 1.1 g cm^?3) or severe (SSC 1.6 g cm^?3) soil compaction. Drought or waterlogging stresses were applied for 2 weeks from 14th to 28th day. In comparison to LSC treatment, in SSC treatment the decrease in dry matter of shoots and roots was greater for sensitive genotypes of maize and triticale (Ancora, CHD-147). Soil drought or waterlogging caused greater decrease of dry matter of shoots and roots in seedlings grown in SSC in comparison to LSC. The root penetration index (RPI) was estimated as a ratio of root dry matter in 15–40 cm root-box layer to total root dry matter. On the basis of RPI it was possible to group the genotypes according to their ability to distribute roots in soil profile. In comparison to LSC, SSC exerted a strong influence on the length of seminal and seminal adventitious roots, as well as the number and length of L- and S-type lateral roots developed on seminal and nodal roots. In both species the restriction effect of soil compaction on number and length of roots was more severe in sensitive (Ankora, CHD-147) than in resistant (Tina, CHD-247) genotypes. The restriction in roots propagation was greater in triticale than in maize. Exposure to drought or waterlogging in the case of genotypes grown in LSC and SSC treatments caused a decrease in number and length of particular components of root system structure. In both species the decrease of root number and length in plants grown under waterlogging was greater than under drought. The observed changes in root system were greater in sensitive (Ankora, CHD147) than in resistant (Tina, CHD-247) genotypes. Statistically significant correlations were found between RPA and RPI and also between these indexes and soil compaction, drought and waterlogging susceptibility indexes. This indicates that genotypes resistant to soil compaction were resistant to drought or waterlogging and also that genotypes resistant to drought were resistant to waterlogging.     

The participation of plant cell organelles in compatible and incompatible potato virus Y-tobacco and -potato plant interaction

Potyviruses replicate and express their genomes in the cytoplasm in closely related membranous structures such as the endoplasmic reticulum or in the vicinity of the ER. The present research demonstrates the participation of plant cell organelles based on ultrastructural examination of compatible and incompatible interactions in tobacco- and potato-potato virus Y (PVY) necrotic strains. In two interaction types, PVY^N Wi and PVY^NTN particles were documented inside cell nuclei. Virus cytoplasmic inclusions and particles were associated with nuclear envelope pore complexes. Moreover, the PVY capsid protein was immunolocalised in the cell nucleus and nucleolus. Our results for the first time show PVY particles and capsid proteins inside the mitochondrion in compatible interactions, whereas in hypersensitive responses these interactions were identified inside chloroplasts. The PVY particles attached to mitochondria caused association groups of these organelles. The ultrastructural analysis clearly demonstrated both the dynamics of the endoplasmatic reticulum in two types of PVY interactions and connections between PVY cytoplasmic inclusions and particles with its membranous structures. Moreover, we demonstrated strongly localised immunodetection of the PVY capsid protein on the surface and in the vicinity of ER in cases of hypersensitive response as well as in compatible interaction.     

Genetic Diversity among Thermophilic Bacteria Isolated from Geothermal Sites by Using Two PCR Typing Methods

Microbial communities thriving at two hot springs, Hammam Pharaon (Pharaoh's Bath) and Oyoun Mossa (Moses springs), in Egypt was studied by cultural and molecular methods. Thirteen morphologically distinct strains of facultative anaerobic thermophilic bacterial isolates have been characterized and identified using phenotypic and genotypic characters including RAPD-PCR, ERIC-PCR typing, plasmid analysis and 16S rRNA sequencing. All isolates produced plasmid DNA with various sizes ranging from 0.7 kb to a larger plasmid 7.2 kb. The bacterial strains could tolerate a temperature range between 45 to 85°C and a pH between 4–11. Also, sulphate-reducing bacteria (SRB) in the thermal springs were investigated with combined biochemical and molecular approaches. A sulphate-reducing bacteria medium containing lactate was used for enrichment and isolation, which yielded Gram negative, rod shaped, anaerobic, non-spore-forming and motile bacteria capable of reducing sulphate to sulphide. These grew at temperatures ranging from 30 to 50°C and could use pyruvate, lactate and ethanol as electron donors. The dissimilatory sulphite reductase (DSR) gene sequences of eleven representative isolates revealed that the strains belonged to the sulphur reducing bacterial species Desulfovibrio vulgaris. 16S rRNA gene partial sequence results indicated the presence of novel or existing species of Bacillus (one species), Anoxybacillus (four species) and Geobacillus (eight species). In this study phenotypic and genotypic diversity were applied for the first time to differentiate thermophilic bacteria of such geothermal sites in Sinai, Egypt.