Oligonucleotides have been ligated efficiently on solid-phase using CuAAC and SPAAC chemistry to produce up to 186-mer triazole linked DNA products. Multiple sequential ligation reactions can be carried out by using a masked azide approach. This work suggests a novel modular approach to the synthesis of large complex oligonucleotide analogues.
A new class of modified oligonucleotides (combination probes) has been designed and synthesised for use in genetic analysis and RNA detection. Their chemical structure combines an intercalating anchor with a reporter fluorophore on the same thymine nucleobase. The intercalator (thiazole orange or benzothiazole orange) provides an anchor, which upon hybridisation of the probe to its target becomes fluorescent and simultaneously stabilizes the duplex. The anchor is able to communicate via FRET to a proximal reporter dye (e.g. ROX, HEX, ATTO647N, FAM) whose fluorescence signal can be monitored on a range of analytical devices. Direct excitation of the reporter dye provides an alternative signalling mechanism. In both signalling modes, fluorescence in the unhybridised probe is switched off by collisional quenching between adjacent intercalator and reporter dyes. Single nucleotide polymorphisms in DNA and RNA targets are identified by differences in the duplex melting temperature, and the use of short hybridization probes, made possible by the stabilisation provided by the intercalator, enhances mismatch discrimination. Unlike other fluorogenic probe systems, placing the fluorophore and quencher on the same nucleobase facilitates the design of short probes containing multiple modifications. The ability to detect both DNA and RNA sequences suggests applications in cellular imaging and diagnostics.
Acetylation is an important, reversible posttranslational modification to a protein. In a previous study, we found that there were a large number of acetylated sites in various nutrient storage proteins of the silkworm haemolymph. In this study, we confirmed that acetylation can affect the stability of nutrient storage protein Bombyx mori apolipophorin‐III (BmApoLp‐III). First, the expression of BmApoLp‐III could be upregulated when BmN cells were treated with the deacetylase inhibitor panobinostat (LBH589); similarly, the expression was downregulated when the cells were treated with the acetylase inhibitor C646. Furthermore, the increase in acetylation by LBH589 could inhibit the degradation and improve the accumulation of BmApoLp‐III in BmN cells treated with cycloheximide and MG132 respectively. Moreover, we found that an increase in acetylation could decrease the ubiquitination of BmApoLp‐III and vice versa; therefore, we predicted that acetylation could improve the stability of BmApoLp‐III by competing for ubiquitination and inhibiting the protein degradation pathway mediated by ubiquitin. Additionally, BmApoLp‐III had an antiapoptosis function that increased after LBH589 treatment, which might have been due to the improved protein stability after acetylation. These results have laid the foundation for further study on the mechanism of acetylation in regulating the storage and utilization of silkworm nutrition.
The production of low-cost biologically activated carbons (BACs) is urgent need of environmental protection and ecological sustainability. Hence, walnut shells were treated by traditional pyrolysis, direct KOH impregnation and combined activation composed of hydrothermal carbonization and two-step H3PO4- and pyrolysis-activation process to obtain porous carbon with high adsorption capacity. It was found that the best adsorption capacity for iodine and organic dye methylene blue (MB) can be obtained using the KOH impregnation at impregnation ratio of 1:1 or combined activation comprising of 2 h H3PO4 activation and 1 h pyrolysis activation at 1000°C. The produced KOH, H3PO4/pyrolysis activated BACs at the optimum conditions are superior to that of commercial ACs, 9.4 and 1.3 times for MB removal, 4 and 4.5 times for iodine number respectively. Characterization results demonstrated their porous structure with very good textural properties such as high BET surface area (1689.1 m2/g, 1545.3 m2/g) and high total pore volume (0.94 cm3/g, 0.96 cm3/g). The N2 adsorption-desorption isotherm of H3PO4/pyrolysis activated hydrochar suggested the co-existence of micro and meso-pores. Moreover, they are more effective for the removal of Fe(III) and Cr(VI) from aqueous solution than the commercial AC, suggesting a promising application in the field of water treatment.
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