Significance of codon usage and irregularities of rare codon distribution in genes for expression of BspLU11III methyltransferases.

Genes of adenine-specific DNA-methyltransferase M.BspLU11IIIa and cytosine-specific DNA-methyltransferase M.BspLU11IIIb of the type IIG BspLU11III restriction-modification system from the thermophilic strain Bacillus sp. LU11 were expressed in E. coli. They contain a large number of codons that are rare in E. coli and are characterized by equal values of codon adaptation index (CAI) and expression level measure (E(g)). Rare codons are either diffused (M.BspLU11IIIa) or located in clusters (M.BspLU11IIIb). The expression level of the cytosine-specific DNA-methyltransferase was increased by a factor of 7.3 and that of adenine-specific DNA only by a factor of 1.25 after introduction of the plasmid pRARE supplying tRNA genes for six rare codons in E. coli. It can be assumed that the plasmid supplying minor tRNAs can strongly increase the expression level of only genes with cluster distribution of rare codons. Using heparin-Sepharose and phosphocellulose chromatography and gel filtration on Sephadex G-75 both DNA-methyltransferases were isolated as electrophoretically homogeneous proteins (according to the results of SDS-PAGE).

[Effect of homologous peptides of differentiation factors HLDF and PEDF on preimplantation development of mice in vitro]. / Vliianie gomologichnykh peptidov factorov differentsirovki HLDF i PEDF na doimplantatsionnoe razvitie myshei in vitro.

We studied the effect of synthetic peptides PEDF-6 and HLDF-6 on preimplantation development of mouse embryos in vitro. PEDF-6 peptide corresponds to fragment 351-356 and of pigment epithelium-derived differentiation factor (PEDF), while HLDF-6 peptide corresponds to fragment 84-89 of differentiation factor HLDF isolated from HL-60 cell line. Despite high homology, these peptides had different effects on the early development. PEDF-6 had no effect on the cleavage of 2-4-cell embryos but decelerated blastocyst formation from such embryos and disturbed their structure. In the presence of HLDF-6 the blastomeres divided more actively as compared to the control and a higher number of embryos developed to the blastocyst stage. The effects of both peptides were stage-specific: the affect the embryos at early cleavage stages and, apparently, determine their further development at that moment although do not directly affect formation of the blastocysts.

[The simulation of the cooperative effect of development in a culture of early mouse embryos after irradiation with electromagnetic waves in the millimeter range]. / Imitatsiia kooperativnogo éffekta razvitiia v kul'ture rannikh zarodyshei myshi posle oblucheniia élektromagnitnymi volnami millimetrovogo diapazona.

We have found that two-cell mouse embryos cultured in vitro can be stimulated by electromagnetic irradiation in the millimeter range. After 30 min of exposure, they acquire the ability to develop in culture on their own and can reach the stage of blastocyst in a relatively large volume of Whitten cultural medium (150 microliters) without serum or growth factors. It is proposed that millimeter range electromagnetic waves activate metabolic processes and specifically the synthesis of factors controlling early embryonic development in culture.

[Effect of millimeter waves on the early development of the mouse and sea urchin embryo]. / Deistvie millimetrovykh voln na rannee razvitie zarodyshei myshei i morskikh ezhei.

The action of nonthermal electromagnetic radiation (EMR) of the millimeter range on the early development of murine and sea urchin embryos was investigated. An MRTA-01E-03 generator with a frequency of 54-78 GHz and radiation intensity of 0.06 mWt/cm2 was used. The embryos were irradiated during 30 min at the stage of two blastomeres. The number of murine embryos that reached the blastocyst stage increased (up to 97.3% in comparison with 87.5% in control). The total time of cultivation up to the blastocyst stage was also shorter (72 h) than in control (96 h). The irradiation had effect on the development of sea urchin embryos only if embryos with a weakened viability were tested. The results indicate that millimeter electromagnetic radiation has a stimulating effect on the early development of embryos, increasing the resistance of embryos to unfavorable environmental conditions.

[Non-Mendelian inheritance of the Un(s) mutant gene in mice]. / Nemendelevskoe nasledovanie mutantnogo gena Un(s) u myshei.

The phenotypic expression of the mutation Uns (gene Pax-1) were studied in the progeny of heterozygous mice. In all crosses, the sex ratio close to 1:1 was observed in the progeny. However, among sons and daughters the segregation for the mutant character depended on the variant of the cross variant. When the Uns/+ mice were intercrossed, the ratio of mutant to normal females was 3:1, while the corresponding ratio among males was 0.9:1. Studying preimplantation development in vitro and postimplantation development in vivo revealed no significant mortality in embryos obtained by crossing heterozygous mice. Ultrastructural analysis revealed no distortions in synapsis and desynapsis of axial elements of the autosomal chromosomes in Uns/+ mice. The segregation found seems to result either from different fertilizing ability of normal sperm and sperm carrying the Uns mutation or, which is more probable, from the genetic imprinting of the gene Uns.

[The positive effect of pretreatment with serotonin and gangliosides on the development of early mouse embryos after cryopreservation]. / Polozhitel'noe vliianie predobrabotki serotoninom i gangliozidami na razvitie rannikh zarodyshei myshei posle kriokonservatsii.

We have demonstrated that viability of preimplantation mouse embryos F1 (C57B1/6 x CBA) after cryoconservation at the stage of four blastomeres improves after pretreatment with serotonin (5 HT, 5 microM) or total gangliosides of bovine brain gangliosides (TG, 3 microM) added to the cultivation medium. After thawing, 64% of embryos preincubated with total brain gangliosides and 49% of embryos preincubated with serotonin developed to the stage of blastocyst during the cultivation in vitro; in the control, no more than 25% of embryos reaches this stage, and all these embryos were abnormal. Possible mechanisms of protective action of these compounds is discussed. We conclude that mouse embryos subjected to freezing-thawing procedure can be used to examine the role of serotonin and gangliosides in the regulatory processes of mammalian preimplantation development.

[Two site-specific endonucleases from the thermophilic OV Bacillus species strain]. / Dve sait-spetsificheskie éndonukleazy iz termofil'nogo shtamma Bacillus species OV.

Two site-specific endonucleases, BspOVI and BspOVII, were isolated from a thermophilic strain Bacillus species OV. The activities of both enzymes are maximal at 48 degrees C and do not depend on ATP and S-adenosyl-L-methionine. BspOVI recognizes the sequence [sequence: see text] and cleaves it as indicated by arrows. Thus, BspOVI is a IIN-subclass endonuclease isoshizomer of Eam1105I. BspOVI is very stable during storage. The enzyme can be used for direct T/A cloning of PCR products. BspOVII recognizes and cleaves the sequence [sequence: see text]; thus, BspOVII is an isoshizomer of CIaI. The cleavage by BspOVII is blocked by dam methylation of adenine inside the recognition site.

[The effect of the mutant genes Ta, Ra and Sd on early embryogenesis in mice]. / Vliianie mutantnykh genov Ta, Ra, Sd na rannii émbriogenez myshei.

We studied early embryonic mortality of mice from mutant stocks Tabby (Ta, X-chromosome) and RaSd (RaSd/++, chromosome 2) maintained in the heterozygous state in F1 CBA x C57B1/6 hybrid. Tabby and RaSd mice were reciprocally crossed with F1 mice and examined for the morphological status of embryos washed from the oviduct on the third day of pregnancy, when the stage of eight blastomeres is normally attained. Mortality was evaluated from the number of embryos which did not reach the expected stage by this time. The results have shown that 2-4 cell embryos, which have received gene Ta with the X-chromosome of the female parent, differed from embryos with F1 genotype at the same stage of development by their increased mortality rate, whereas among embryos obtained from RaSd, the mortality was mainly observed before cleavage. Death of embryos receiving the mutant gene from hemizygous Ta males or heterozygous RaSd/++ males was not significantly different from the mortality of embryos without these mutations.