[Packaging of Subchromosomal-Size DNA Molecules in Chromatin Bodies in the Ciliate Macronucleus].

A fundamental difference between somatic nuclei (macronuclei) of ciliates and cell nuclei of higher eukaryotes is that the macronuclear genome is a huge number (up to tens or hundreds of thousands) of gene-sized (0.5-25 kb) or subchromosomal (up to 2000 kb) minichromosomes. Electron microscopy shows that macronuclear chromatin usually looks like chromatin bodies or fibrils 200-300 nm thick in the interphase. However, the question of how many DNA molecules are contained in an individual chromatin body remains open. The organization of chromatin in macronuclei was studied in the ciliates Didinium nasutum and three Paramecium sp, which differ in pulsed-field gel electrophoresis (PFGE) karyotype, and compared with the model of topologically associated domains (TADs) of higher eukaryotic nuclei. PFGE showed that the sizes of macronuclear DNAs ranged from 50 to 1700 kb, while the majority of the molecules were less than 500 kb in length. A comparative quantitative analysis of the PFGE and electron microscopic data showed that each chromatin body contained one minichromosome in P. multimicronucleatum in the logarithmic growth phase, while bodies in the D. nasutum macronucleus contained two or more DNA molecules each. Chromatin bodies aggregated during starvation, when activity of the macronuclei decreased, leading to an increase of chromatin body size or the formation of 200- to 300-nm fibrils of several chromatin bodies. A model was proposed to explain the formation of such structures. In terms of topological characteristics, macronuclear chromatin bodies with subchromosomal DNA molecules were found to correspond to higher eukaryotic TADs.

[Comparative description of macronuclear electrophoretic karyotypes of Paramecium primaurelia and Paramecium novaurelia sibling species].

The macronuclear genomes of two sibling species belonging to the Paramecium aurelia complex--P. primaurelia and P. novaurelia--were analyzed by pulsed-field gel electrophoresis (PFGE). Their electrophoretic karyotypes showed a continuous spectrum of different-sized DNA molecules with a species-specific pattern of banding, which was reproducible and did not change with strain senescence. Thus, P. aurelia macronuclear PFGE profiles could be described by an approach analogous to that used for the description of metaphase chromosome banding patterns. At first, well-identifiable regions (size fractions) of a PFGE profile, which can be seen at any resolution, are determined. Then, the bands of the second order of resolution (subfractions) can be defined in some of these regions. The blocks of the first and second orders can be utilized as inner markers of the PFGE profile allowing precise comparison of different PFGE profiles. Such comparative analysis has demonstrated some marking differences in organization of the macronuclear genomes of P. primaurelia and P. novaurelia, and low level of intraspecific polymorphism. Worth noting is that the P. novaurelia strain isolated in USA was different from all other analyzed P. novaurelia strains originating from Europe. The nature of banding of P. aurelia PFGE profiles is discussed. The revealed high order and stability of the macronuclear genome organization makes possible searching for new approaches to study the mechanisms of this non-trivial genome formation and maintenance. Further PFGE analysis of the macronuclear genomes of the other Paramecium species in relation with the Paramecium phylogeny may provide insights into the directions of the evolution of the macronuclear genome in Ciliata.

[Identification of Chlorella viruses in Paramecium bursaria clones by pulse-field electrophoresis].

The ciliates Paramecium bursaria contain endosymbiotic green algae Chlorella spp. in their cytoplasm. The algae isolated from P. bursaria are sensitive to large DNA-containing viruses of the family Phycodnaviridae. The type virus of this family is PBCV-1 (Paramecium bursaria Chlorella virus). Investigation of the total DNA of P. bursaria clones by pulse-field electrophoresis (PEP) revealed a pronounced band on PEP profiles of some P. bursaria clones; the band was formed by DNA molecules of approx. 300 kb. This band probably contained the DNA of chlorella virus. Two approaches were used in the present work to confirm this hypothesis. Microbiological tests were used to scan a collection of P. bursaria clones for specific types of viruses; the 300-kb band was revealed only in the PEP profiles of virus-containing clones. Blot hybridization of P. bursaria total DNA separated by pulse-field electrophoresis with the virus-specific probe revealed that the band under study contained the DNA of a chlorella virus. Paramecium clones were shown to contain approx. 10(5) copies of nonintegrated viral DNA.

[Determination of the minimum size of Gallus gallus domesticus chicken microchromosome by a pulse electrophoresis method]. / Opredelenie minimal'nogo razmera mikrokhromosom kuritsy Gallus gallus domesticus metodom pul's-élektroforeza.

The karyotype of the chicken Gallus gallus domesticus was studied by means of pulsed-field electrophoresis. An electrokaryogram was obtained for the microchromosomal (MI) portion of the chicken genome. Chicken MIs were separated into two fractions. A fraction with a higher mobility included MIs sized 3.4-4.8 Mb; the lower size limit of a less mobile fraction corresponded to MIs of approximately 5 Mb. The smallest MI in the chicken karyotype was estimated at 3.4 +/- 0.25 Mb.