Restriction fragment length polymorphism of the apolipoprotein A-I gene among inhabitants of Novosibirsk]

Restriction fragments' length polymorphism in the region of apolipoprotein A-I (apo A-I) gene was investigated in Novosibirsk (Siberia, USSR) population. Correlation between PstI apo A-I alleles (2,2 kb-P1; 3,3 kb-P2) and total cholesterol, triglycerides, high density polyproteins cholesterol, and apolipoprotein A-I level was analysed. A tendency to increase in cholesterol index of atherogenicity and to decrease in high density lipoproteins cholesterol as well as apolipoprotein A-I level was shown to occur for P1P2 genotype patients.     

Haemophilus influenzae antibiotic resistant strains isolated in Moscow in 2002-2004]

The results of 3-year observation on emergence of H. influenzae antibiotic resistant strains in Moscow are summarized. The study included 566 strains isolated from patients in 2002-2004. The susceptibility was determined by the 2-fold microdilution method on the Haemophilus test medium. The percentage of the resistant strains isolated in 2002, 2003 and 2004 was the following: ampicillin --4.9, 3.2 and 3.6%, tetracycline--3.3, 3.2 and 1.8% and co-trimoxazole--10.9, 20.9 and 20% respectively. The strains isolated in 2003 and 2004 were resistant to azithromycin in 0.6 and 1.8% of the isolates and to clarithromycin in 1.3 and 3.2% of the isolates respectively. Five isolates differed by the minimum resistance to ampicillin whereas producing no beta-lactamase (BLNAR strains). The drugs of choice for the treatment of respiratory tract infections mainly due to H. influenzae, i.e. acute otitis and sinusitis, chronic bronchitis exacerbation and sometimes pneumonia remain betalactam antibiotics. From the microbiological viewpoint the inhibitor-protected aminopenicillins, cefuroxime and cefotaxime have no significant advantages vs. amoxycillin. The use of cotrimoxazole and chloramphenicol should be considered inexpedient.     

The isolation of nucleases Sm1 and Sm2 in preparative amounts and a study of their antigenicity]

Two enzyme forms of endonuclease (Sm 1 and Sm 2) strain B10M1 in 60 and 100 mg respectively have been isolated from the culture fluid Serratia marcescens. The chromatographic and electrophoretic properties and N-terminal amino acid residues are different for both enzymes. The purification procedure consists of dialysis and ion-exchange chromatography on DEAE- and phosphocellulose. The yield of nucleases Sm1 and Sm2 are 14% and 28% respectively. The antigenic differences of nucleases Sm1 and Sm2 have been found by cross immunoenzyme analysis.     

Anonymous nuclear DNA markers in the American oyster and their implications for the heterozygote deficiency phenomenon in marine bivalves

A puzzling population-genetic phenomenon widely reported in allozyme surveys of marine bivalves is the occurrence of heterozygote deficits relative to Hardy-Weinberg expectations. Possible explanations for this pattern are categorized with respect to whether the effects should be confined to protein-level assays or are genomically pervasive and expected to be registered in both protein- and DNA-level assays. Anonymous nuclear DNA markers from the American oyster were employed to reexamine the phenomenon. In assays based on the polymerase chain reaction (PCR), two DNA-level processes were encountered that can lead to artifactual genotypic scorings: (a) differential amplification of alleles at a target locus and (b) amplification from multiple paralogous loci. We describe symptoms of these complications and prescribe methods that should generally help to ameliorate them. When artifactual scorings at two anonymous DNA loci in the American oyster were corrected, Hardy-Weinberg deviations registered in preliminary population assays decreased to nonsignificant values. Implications of these findings for the heterozygote-deficit phenomenon in marine bivalves, and for the general development and use of PCR-based assays, are discussed.      

Basolateral plasma membrane localization of ouabain-sensitive sodium transport sites in the secretory epithelium of the avian salt gland

The distribution of Na+ pump sites (Na+-K+-ATPase) in the secretory epithelium of the avian salt gland was demonstrated by freeze-dry autoradiographic analysis of [(3)H] ouabain binding sites. Kinetic studies indicated that near saturation of tissue binding sites occurred when slices of salt glands from salt-stressed ducks were exposed to 2.2 μM ouabain (containing 5 μCi/ml [(3)H]ouabain) for 90 min. Washing with label-free Ringer's solution for 90 min extracted only 10% of the inhibitor, an amount which corresponded to ouabain present in the tissue spaces labeled by [(14)C]insulin. Increasing the KCl concentration of the incubation medium reduced the rate of ouabain binding but not the maximal amount bound. In contrast to the low level of ouabain binding to salt glands of ducks maintained on a freshwater regimen, exposure to a salt water diet led to a more than threefold increase in binding within 9-11 days. This increase paralleled the similar increment in Na+-K+-ATPase activity described previously. [(3)H]ouabain binding sites were localized autoradiographically to the folded basolateral plasma membrane of the principal secretory cells. The luminal surfaces of these cells were unlabeled. Mitotically active peripheral cells were also unlabeled. The cell-specific pattern of [(3)H]ouabain binding to principal secretory cells and the membrane-specific localization of binding sites to the nonluminal surfaces of these cells were identical to the distribution of Na+-K+-ATPase as reflected by the cytochemical localization of ouabain-sensitive and K+-dependent nitrophenyl phosphatase activity. The relationship between the nonluminal localization of Na+-K+-ATPase and the possible role of the enzyme n NaCl secretion is considered in the light of physiological data on electrolyte transport in salt glands and other secretory epithelia.     

构建基于CRISPR/Cas9技术敲除ALOX5的质粒

旨在利用CRISPR/Cas9技术构建敲除花生四烯5-脂氧合酶基因(Arachidonate 5-lipoxygenase gene,ALOX5)的重组质粒。设计合成3对靶向敲除ALOX5第六外显子的sgRNA,将其分别插入到CRISPR/Cas9质粒骨架pX458载体中,转化感受态大肠杆菌DH5α后挑取克隆,通过测序评估重组质粒是否构建成功。将构建好的重组质粒转染293T细胞,在荧光显微镜下观察转染效果,挑取转染成功的细胞,用试剂盒提取转染细胞基因组DNA,PCR扩增含敲除位点的DNA片段,用测序技术获得核苷酸序列,用DNAStar软件分析转染细胞中ALOX5基因敲除情况。测序结果表明2对双链sgRNA寡核苷酸已插入质粒,且序列正确,靶向ALOX5基因的重组质粒pX458-sgRNAs-ALOX5构建成功。其在293T细胞中的转染效率约为50%,用一代测序法未检测到sgRNAs的切割效果。初步表明利用CRISPR/Cas9技术成功构建靶向ALOX5基因的重组质粒pX458-sgRNAs-ALOX5。     

Morphological aspects of blood digestion in a parasitic mite Bakericheyla chanayi

All life stages of B. chanayi (Acariformes: Cheyletidae) are characterized by occasional bloodsucking and a long period of digestion. No newly engorged mites were found during the period of their host birds' migration. The fine structure of the digestive tract of a blood-feeding acariform mite is described for the first time. The anterior midgut (AMG) is a place of blood digestion, while the posterior midgut (PMG) is involved in nitrogen metabolism forming guanine crystals as the main end-product. The AMG epithelium consists of digestive cells that probably arise from mitotically active basal cells with high synthesizing activity.As observed in ticks, blood digestion is accompanied by the formation of huge endosomes that serve as places of storage and sorting of ingested material. Digestive cells show different types of endocytotic activity as well as various late endosomes, which implies different subcellular pathways for different blood components. In both midgut regions, elimination of the excretory material occurs by apocrine secretion or by discharging of apical cell fragments (loaded with lysosomes) into the gut lumen. The formation of guanine granules occurs inside the lysosomes of PMG epithelial cells thus having much in common with intracellular digestion. Peculiarities of intracellular blood digestion were analyzed according to the modern hypothesis of endocytosis and compared to what is known in ticks.     

The early life history of two sympatric New Zealand octopuses: eggs and paralarvae of Octopus huttoni and Pinnoctopus cordiformis

This study combined morphological and morphometric information on egg clutches, egg capsules and paralarvae of two sympatric coastal octopuses from New Zealand waters, Octopus huttoni and Pinnoctopus cordiformis, to provide species-specific traits to identify their early life stages obtained from field surveys. Eggs of O. huttoni (2.5 mm length; 1 mm width) were entwined with one another forming strings that ranged from 11 to 25.8 mm in length. Eggs of P. cordiformis (6.4 mm length; 1.5 mm width) were significantly bigger than those of O. huttoni and were grouped in small clusters of about seven eggs. Paralarvae O. huttoni and P. cordiformis differed in hatching size (1.4 mm versus 3.1 mm mantle length), number of suckers per arm (four versus eight), number of lamellae per outer demibranch (five versus ten) and arrangements of chromatophores in the body surface (29 to 59 versus 91 to 179), respectively. The morphological traits described in hatchlings from the laboratory allowed comparisons with field-collected paralarvae, suggesting that such characters were reliable species-specific patterns to enable a consistent differentiation between the early life stages of these two sympatric species, even in the absence of the brooding female.