A sensitive method for determination of trace amounts of chromate (III) with terbium (III) sodium hexametaphosphate chelate as fluorescent probe

Based on the fluorescence quenching of Terbium (III)‐sodium hexametaphosphate (Tb/SHMP) chelates in the presence chromate (III), a sensitive fluorimetric method was developed for the determination of trace amounts of chromium (III) in aqueous solutions. Under the optimum conditions, the linear calibration graph was obtained (R = 0.996). The linear range and detection limit of Cr (III) were 7.69 × 10^?7 to 1.15 × 10^?4 mol L^?1 and 4.50 × 10^?7 mol L^?1, respectively. The proposed method had a wider linear range and was proved to be very sensitive, rapid and simple. The method was applied successfully to the determination of chromium (III) in the synthetic samples and real water samples. Moreover, the reaction mechanism was discussed through the fluorescence lifetime and proved to be dynamic quenching behavior. Copyright © 2010 John Wiley & Sons, Ltd.     

A non-radioactive method to detect RNA or DNA using an oligonucleotide probe with bromodeoxyuridine free ends, a monoclonal antibody against bromodeoxyuridine and immunogold silver staining

A non radioactive method for probing RNA or DNA on dot and Northern blots using a synthetic oligonucleotide with bromodeoxyuridine free ends is described. The present experiment was carried out with human testis and placental RNA's. The probe was the 21 base long sequence coding for the amino acids 18 to 24 of the insulin-like growth factor I (IGF-I) with two bromodeoxyuridine dinucleotides added at the 5' and 3' ends. The probe was detected with a monoclonal antibody against bromodeoxyuridine and immunogold silver staining (IGSS). Our method was compared to the peroxydase (HRP) revelation of the same probe. The results obtained show a lower background with IGSS than with HRP revelation. A sensitivity similar to that of 32P labelling was found with the advantages of an increase in the rapidity of the procedure (24 hours instead of 9 days exposure) and the absence of handling radioactive substances. Moreover, as the monoclonal antibody against BrdU detects single stranded DNA only, the use of BrdU free ends-labelled oligonucleotide allows the development of the revelation procedure without any previous denaturation of the hybrid. This particular point is an indisputable advantage for detecting hybridization in situ.     

Improvement of ciliate identification and quantification: a new protocol for fluorescence in situ hybridization (FISH) in combination with silver stain techniques

A new protocol for taxon specific probe based fluorescent in situ hybridization was developed for the identification and quantification of ciliates in microbial communities. Various fixatives and experimental parameters were evaluated and optimized with respect to cell permeability and morphological preservation. Optimum results were adaption by obatined of a modified fixation method using Bouin's solution. Furthermore, conventional staining procedures such as different Protargol stain techniques and a silver nitrate impregnation method were modified and can now be applied in combination with fluorescence in situ hybridization. The new protocol allows a rapid and reliable identification as well as quantification of ciliates based upon classical morphological aspects and rRNA based phylogenetic relationships performed in one experiment. Furthermore, a set of specific probes targeting different regions of the 18S rRNA was designed for Glaucoma scintillans Ehrenberg, 1830 and tested by applying this new approach of combining in situ cell hybridization with conventional staining techniques.     

A new species of Selaginella from Hainan Island, China, with a key to the Hainan species

Selaginella hainanensis X.C.Zhang & Noot. sp. nov. , a new species from Hainan Island, South China, is described and illustrated. Diagnostic notes and a key to the species distributed in Hainan are also provided.  © 2005 The Linnean Society of London, Botanical Journal of the Linnean Society , 2005, 148 , 323–327.     

A new approach to detect a particular DNA sequence by UV-immobilization of its hybridization complex with a highly specific probe resulting from ligation of a tandem of short oligonucleotides in solution

A new approach was proposed for detecting amplified DNA fragments by hybridization with a highly selective oligonucleotide probe obtained by ligation of a tandem of three short oligonucleotides (pN8 + pN4 + pN8' Bio) in solution, with subsequent UV-immobilization of the hybridization product on a nylon membrane and its colorimetric detection with the streptavidin-alkaline phosphatase technique. Owing to the high selectivity of ligation, the 20-mer ligation product was detected on a membrane only when it was completely complementary to a template fragment. The results showed that any single-nucleotide substitution in the tetramer-binding site can be localized and identified with the use of all 12 possible tetramers.     

A novel nucleic acid-binding protein in the cyanobacterium Synechococcus sp. PCC6301: a soluble 33-kDa polypeptide with high sequence similarity to ribosomal protein S1

Cyanobacteria are prokaryotes that carry out plant-type photosynthesis and contain several eukaryotic-type RNA-binding proteins. Using a single-stranded DNA column, a 33-kDa protein was isolated and characterized from Synechococcus sp. PCC6301. This protein of 293 amino acids is similar in overall structure to the ribosomal protein S1 found in the same species, and contains three repeated units that are highly similar to the S1 motif originally found in the ribosomal protein S1 of Escherichia coli. However, the 33-kDa protein was found not to be associated with ribosomes and its nucleic acid binding specificity is distinct from that of the ribosomal protein S1. As this protein has high affinity for both single- and double-stranded DNA, as well as for poly(G) and poly(A), we tentatively named it nucleic acid-binding protein 1 (Nbp1). Received: 8 October 1999 / Accepted: 24 January 2000     

A new plant protein interacts with eIF3 and 60S to enhance virus‐activated translation re‐initiation

The plant viral re‐initiation factor transactivator viroplasmin (TAV) activates translation of polycistronic mRNA by a re‐initiation mechanism involving translation initiation factor 3 (eIF3) and the 60S ribosomal subunit (60S). QJ;Here, we report a new plant factor—re‐initiation supporting protein (RISP)—that enhances TAV function in re‐initiation. RISP interacts physically with TAV in vitro and in vivo. Mutants defective in interaction are less active, or inactive, in transactivation and viral amplification. RISP alone can serve as a scaffold protein, which is able to interact with eIF3 subunits a/c and 60S, apparently through the C‐terminus of ribosomal protein L24. RISP pre‐bound to eIF3 binds 40S, suggesting that RISP enters the translational machinery at the 43S formation step. RISP, TAV and 60S co‐localize in epidermal cells of infected plants, and eIF3–TAV–RISP–L24 complex formation can be shown in vitro. These results suggest that RISP and TAV bridge interactions between eIF3‐bound 40S and L24 of 60S after translation termination to ensure 60S recruitment during repetitive initiation events on polycistronic mRNA; RISP can thus be considered as a new component of the cell translation machinery.     

Mycoplasma bovis isolates from dairy calves in Japan have less susceptibility than a reference strain to all approved macrolides associated with a point mutation (G748A) combined with multiple species‐specific nucleotide alterations in 23S rRNA

Erythromycin, tylosin and tilmicosin are approved for use in cattle in Japan, the latter two being used to treat Mycoplasma bovis infection. In this study, 58 M. bovis isolates obtained from Japanese dairy calves all exhibited reduced susceptibility to these macrolides, this widespread reduced susceptibility being attributable to a few dominant lineages. All 58 isolates contained the G748A variant in both the rrl3 and rrl4 alleles of 23S rRNA, whereas a reference strain (PG45) did not. G748 localizes in the central loop of domain II (from C744 to A753) of 23S rRNA, which participates in binding to mycinose, a sugar residue present in both tylosin and tilmicosin. A number of in vitro‐ selected mutants derived from M. bovis PG45 showed reduced susceptibility to tylosin and tilmicosin and contained a nucleotide insertion within the central loop of domain II of rrl3 (U747–G748Ins_CU/GU or A743–U744Ins_UA), suggesting that mutations around G748 confer this reduced susceptibility phenotype. However, other Mycoplasma species containing G748A were susceptible to tylosin and tilmicosin. Sequence comparison with Escherichia coli revealed that M. bovis PG45 and isolates harbored five nucleotide alterations (U744C, G745A, U746C, A752C and A753G) in the central loop of domain II of 23S rRNA, whereas other Mycoplasma species lacked at least two of these five nucleotide alterations. It was therefore concluded that G748 mutations in combination with species‐specific nucleotide alterations in the central loop of domain II of 23S rRNA are likely sufficient to reduce susceptibility of M. bovis to tylosin and tilmicosin.