Regeneration of Picea omorika plants via organogenesis

Picea omorika plants were regenerated from embryo and seedling shoot tip cultures. Adventitious and axillary shoots were produced on 1/2 MS medium containing benzyladenine and kinetin. Benzyladenine was more effective in bud induction, whereas kinetin hastened shoot development. Excised shoots were elongated on 1/3 MS medium without growth regulators, multiplied with kinetin and rooted with or without indole-3-butyric acid.Abbreviations BA N⁶-benzyladenine - 2IP N ⁶-(²-isopenteny) adenine - NAA -naphthaleneacetic acid - IAA indole-3-acetic acid - IBA indole-3-butyric acid     

Genetic transformation of alfalfa somatic embryos and their clonal propagation through repetitive somatic embryogenesis

Genetically transformed alfalfa (Medicago sativa L., cv. Zajearska 83) plantlets were obtained by inoculating somatic embryos with Agrobacterium tumefaciens strains A281/pGA472 and LBA4404/pBI121. Single somatic embryos, 5–7 mm long, were released from a repetitively embryogenic culture, wounded, and cocultivated with the bacteria. The agar-solidified culture medium contained mineral salts, vitamins, 40 g l^–1 sucrose, 1 g l^–1 yeast extract and 0.05 mg l^–1 BA. Five clones, transformed with A281/pGA472, and 4 clones transformed with LBA4404/pBI121, were selected for proliferation by repetitive somatic embryogenesis, on media containing 100 mg l^–1 of kanamycin. The transformation of kanamycin-resistant clones was confirmed by assaying the activity of neomycin phosphotransferase II and/or -glucuronidase enzymes, and by the Southern blot analysis. It is suggested that the transformation/regeneration system based on somatic embryogenesis may be suitable for establishing transgenic alfalfa lines. The relatively low frequency of embryo transformation is compensated for by abundant proliferation in secondary somatic embryogenesis.Abbreviations BA 6-benzyladenine - GUS -glucuronidase - Km kanamycin - NPTII neomycin phosphotransferase II - X-gluc 5-bromo-4-chloro-3-indolyl--glucuronic acid - BM basal medium     

Alterations in chemical shifts and exchange broadening upon peptide boronic acid inhibitor binding to α-lytic protease

-Lytic protease, a bacterial serine protease of 198 aminoacids (19800 Da), has been used as a model system for studies of catalyticmechanism, structure–function relationships, and more recently forstudies of pro region-assisted protein folding. We have assigned thebackbones of the enzyme alone, and of its complex with the tetrahedraltransition state mimic N-tert-butyloxycarbonyl-Ala-Pro-boroVal, usingdouble- and triple-resonance 3D NMR spectroscopy on uniformly¹⁵N- and ¹³C/¹⁵N-labeled protein.Changes in backbone chemical shifts between the uncomplexed and inhibitedform of the protein are correlated with distance from the inhibitor, thedisplacement of backbone nitrogens, and change in hydrogen bond strengthupon inhibitor binding (derived from previously solved crystal structures).A comparison of the solution secondary structure of the uninhibited enzymewith that of the X-ray structure reveals no significant differences.Significant line broadening, indicating intermediate chemical exchange, wasobserved in many of the active site amides (including three broadened toinvisibility), and in a majority of cases the broadening was reversed uponaddition of the inhibitor. Implications and possible mechanisms of this linebroadening are discussed.     

Oestradiol, a new hapten for detecting nucleic acid sequences by FISH

 Oestradiol has been conjugated to allylamine-dUTP with an 11-atom spacer to allow enzymatic incorporation of the label into DNA sequences. In a comparative DNA and mRNA FISH study we have used DNA probes that were either labelled with digoxigenin, biotin or oestradiol. Results show that oestradiol-labelled probes can detect DNA and RNA sequences in FISH equally well as digoxigenin- and biotin-labelled probes. Further, no crossreactivity between the various hapten-specific antibodies and the three haptens were observed. Binding of the rabbit anti-oestradiol antibody to endogenous oestrogen in various tissues was not observed under the conditions tested. In view of the increasing demands for multi-colour DNA and mRNA FISH applications, oestradiol is a welcome addition to the collection of haptens employed in FISH. Accepted 20 June 1997     

Mouse {beta}-galactoside {alpha}2,3-sialyltransferases: comparison of in vitro substrate specificities and tissue specific expression

Four types of ß-galactoside     

Nitrogen,phosphorus and Daphnia grazing in controlling phytoplankton biomass and composition – an experimental study

The role of nitrogen as a factor controllingphytoplankton biomass was studied in nutrientenrichment incubations in the laboratory using waterfrom pelagic region of two mesotrophic lakes ineastern Finland, Lake Kallavesi (in year 1994) andLake Juurusvesi (in year 1995). We used differentcombinations of phosphorus and nitrogen additions ina total of eight experiments. Furthermore, we includedDaphnia grazing treatment to the experimentaldesign in Lake Juurusvesi experiments. The nitrogentreatments did not increase chlorophyll aconcentration in any of the experiments compared withthe controls. Chlorophyll a content was highestin those nutrient treatments where phosphorus wasadded with or without nitrogen. Daphnia grazingdecreased chlorophyll a concentration comparedwith non-grazed treatments. In some cases grazing alsocaused higher ammonium concentrations. Theseexperiments, as well as the nutrient ratio of the lakewater used, suggest that phosphorus is likely tocontrol the amount of phytoplankton biomass. This revised version was published online in July 2006 with corrections to the Cover Date.     

In vitro microsome-mediated aflatoxin B1-DNA binding and its inhibition by cytosol of various organs of the hamster and quail

We studied thein vitro activation of aflatoxin B1 (B1) by microsomes and its inactivation by the cytosol of various quail and hamster organs, using B1-DNA binding as an index. The microsomal activity of the liver to bind B1 to DNA was not largely different between the two species and was higher than that of the other organs examined in either species. The microsomal activity of the kidney and lung was very low in the quail compared with the hamster, indicating the very small contribution of the lung and kidney microsomes to the activation of B1 in birds. Only the hamster liver cytosol showed strong inhibition of microsome-mediated B1-DNA binding.     

Binding of divalent copper and calcium ions to poly I

The effect ot Cu²⁺ and Ca²⁺ ions, on the ultraviolet differential (UVD) spectra of single-stranded poly I was studied and the coordination (Δε_b) and conformation (Δε_c) conponents of the spectra calculated The comparison of Δε_b and the UVD spectrum of protonated IMP leads to the conclusion that N(7) ot inosine-5'-monophosphate (IMP) is a coordinating site tor Ca²⁺ and Cu²⁺ ions on the polymer bases. The binding ot Ca²⁺ and Cu²⁺ ions causes differently directed displacements of the four absorption bands of poly I, which are observed in the wavenumber range (50-34) × 10³ cm⁻¹ The calculation of concentration dependencies tor the association constants (K“) ot Ca²⁺ and Cu²⁺ ions binding to poly I bases shows that the binding is cooperative The K“ values for the poly I + Ca²⁺ complex are two orders of magnitude lower than those for the poly 1 + Cu²⁺ complex At low ion concentrations, binding to the poly I phosphates predominates and increases the degree of the polynucleotide helicity. At higher concentrations the spectra are mainly affected by the ion binding to bases, which results in melting of the helical parts of poly I At Ca²⁺ concentrations exceeding 10⁻³ M light-scattering aggregates are formed. The degree of monomer order in them is close to that observed in multistranded helices of poly I     

The coding region of the human c-mos pseudogene contains Alu repeat insertions

We have determined the nucleotide sequence of an 841-bp fragment derived from a segment of the human genome previously cloned by Chumakov et al. [Gene 17 (1982) 19–26] and Zabarovsky et al. [Gene 23 (1983) 379–384] and containing regions homologous to the viral mos gene probe. This sequence displays homology with part of the coding region of the human and murine c-mos genes, contains several termination codons, and is interrupted by two Alu-family elements flanked by short direct repeats. Probably, the progenitor of the human c-mos gene was duplicated approximately at the time of mammalian divergence, was converted to a pseudogene, and acquired insertions of two Alu elements.