Random amplified polymorphic DNA fingerprinting (RAPD fingerprinting) enables the determination of inter- and intraspecific genetic diversity in Euplotes ciliates on a molecular genetic level. Even ...single ciliates can be genetically characterized by this method. Three Euplotes species and several genetically recombined strains of E. octocarinatus (Carter) and E. aediculatus (Pierson) were compared and identified by two oligonucleotide probes. The similarity of fingerprints, measured by a band sharing index (D), was 0.38-0.46 when different species of Euplotes were compared. D was 0.60-0.78 when strains of the same species were compared. RAPD fingerprinting was then used to investigate the genetic structure in a natural population of E. daidaleos (Diller & Kounaris). The results revealed a clonal population structure. Altogether, 130 individuals of E. daidaleos were isolated from a small pond in Münster (Germany). Their genetic diversity was very low. The population consisted of only two different genotypes, with a D-value of 0.97 and a numerical proportion of 94:6. This indicates that sexual reproduction seldom occurs in this population. The fingerprint pattern of a strain of E. daidaleos (strain 13), which originated from a different habitat, was not found in the ciliates of the population from Münster (D = 0.60-0.72). This finding may allow the experimental analysis of the adaptive significance of predator-induced defense in natural habitats of Euplotes. After a release of ciliates of strain 13 and any defenseless mutant of it, into the characterized natural population of E. daidaleos, they could be identified and predation dependent changes of genotype frequencies measured.
The polypeptide release factor gene, eRF1, of Blepharisma japonicum (Bj-eRF1) was cloned and sequenced. Its coding region was 1314 base pairs and encodes a protein of 437 amino acids. The cloned gene ...was expressed in Escherichia coli and the recombinant Bj-eRF1 polypeptide was purified by Ni2+-nitrilotriacetic acid agarose and Superose12 chromatography. Pull-down analysis showed that the recombinant Bj-eRF1 interacts with the heterologously-expressed release factor, eRF3C, of Euplotes octocarinatus.
The gamma-tubulin (gamma-Tub)-encoding gene (gamma-tub) of Euplotes octocarinatus was amplified from macronuclear DNA with the help of the polymerase chain reaction (PCR) and sequenced. The ...polypeptide deduced from the gene consists of 462 amino acids (aa). It shares 61% aa identity with the Aspergillus nidulans gamma-Tub. The gene contains an in-frame TGA codon and two small pre-mRNA introns (36 and 26 bp). We suggest that the TGA, like TGA codons in the pheromone-encoding genes of E. octocarinatus, codes for a cysteine. This suggestion is supported by the finding that in the gamma-Tub of other organisms, a cysteine is located at this position. Sequencing the mRNA revealed that the introns are absent from the gamma-tub transcripts. The second intron constitutes the shortest one reported so far. We have also sequenced the 5'- and 3'-flanking regions of the gene up to the telomeres and report here the entire sequence of the macronuclear DNA molecule carrying gamma-tub.
Three macronuclear genes encoding putative nuclear protein kinases of the ciliate Euplotes octocarinatus syngen 1 were isolated and sequenced. All three deduced gene products share significant ...properties with a group of recently identified nuclear serine/threonine protein kinases named Ndr. The three predicted proteins contain the twelve conserved catalytic subdomains of protein kinases and 22 near universally-conserved amino acids residues that are characteristic of serine/threonine protein kinases. In addition, there is an ∼30 amino acid-peptide insertion between subdomains VII and VIII that contains a potential nuclear localization signal. Sequence analysis suggests that expression of the Eondr2 gene requires a +1 programmed translational frameshift for its translation. Comparison of the deduced EoNdr2 with other known Ndr protein kinases implies that a +1 ribosomal frameshift occurs at the motif AAATAA.
New primer molecules have been synthesized to increase the adhesion strength between a copper leadframe and an epoxy molding compound in microelectronical devices. The coupling agents were ...preliminarily chemisorbed at the surface of copper plates via special binding groups like thiol, disulfide, ethylene diamine and phthalocyanine. Binding to the epoxy resin was performed via an hydroxyl group. Linear hydrocarbon spacers with various chain lengths connected the copper- and epoxy-binding groups. The self-assembled layers of the organic coupling agents at the metal surface were characterized by X-ray photoelectron spectroscopy. Thermogravimetric analysis was used to study the coating with respect to its corrosion oxidation inhibition. Shear tests clearly indicated that the coupling agents increase adhesion strength and are stable even in extreme humidity and thermal conditions in analogy to IPC-Level-1 pretreatment. Thus, delamination of the microelectronical packages was prevented.
We investigate the shear viscosity of a pion gas in relativistic kinetic theory, using the Nambu-Jona-Lasinio model to construct the pion mass and the pi-pi interaction at finite temperature. Whereas ...at low temperatures the scattering properties and, hence, the viscosity are in agreement with lowest-order chiral perturbation theory, we find strong medium modifications in the crossover region. Here the system is strongly coupled and the scattering lengths diverge, similarly as for ultra-cold Fermi gases at a Feshbach resonance. As a consequence, the ratio eta/s is found to be strongly reduced as compared to calculations without medium-modified masses and scattering amplitudes. However, the quantitative results are very sensitive to the details of the applied approximations.
The predatory ciliate
Lembadion bullinum releases a chemical compound (L-factor) which induces morphological changes in another ciliate,
Euplotes octocarinatus, and some of its relatives. The changes ...render
Euplotes so extended that
Lembadion has difficulties engulfing this prey. We have previously shown that the L-factor is a protein with a mass of 31.5 kDa. Here we report the primary structure of the L-factor deduced from cDNA, the heterologous expression of the
Lembadion gene encoding the L-factor, the production of an antibody directed against this factor and the labeling of the cell surface of Lembadion with this antibody. From this and from peculiarities in the amino acid composition of the L-factor we conclude that the L-factor is a surface protein of
Lembadion or at least a major part of it. To our knowledge the L-factor is the first kairomone whose origin and structure has been identified.
Deviations from the universal genetic code have evolved independently several times in ciliated protozoa. Thus, in some species UAA and UAG are no longer used as termination codons, but are read as ...glutamine, whereas in the genus Euplotes, UGA is translated as cysteine. We have investigated the nature of the tRNACys isoacceptor responsible for decoding UGA in Euplotes cells. Southern hybridization analyses indicated that a single DNA molecule of 630 bp encoding tRNACys exists in the macronucleus of Euplotes octocarinatus. Cloning and sequencing of this fragment revealed that it contains only one copy of a tRNACys gene, which codes for a normal tRNACys with GCA anticodon. This is the first report of the characterization of a tRNA gene in any hypotrichous ciliate. It contains putative signals for initiation and termination of transcription by RNA polymerase III and can be transcribed efficiently in vitro in HeLa cell nuclear extract. Intensive studies on the DNA and tRNA level involving PCR analyses have not disclosed the existence of any tRNACys isoacceptor with UCA or ICA anticodons. Translation of the UGA codon by tRNACysGCA necessitates a G:A mispairing in the first anticodon position. We discuss a number of aspects which might contribute to the finding that a near-cognate tRNA isoacceptor efficiently translates the UGA stop codon.
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