Molecular genetic organization and origin of plasmid pBS52 with a broad range of bacterial hosts

The data are presented on the localization of genetic determinants of resistance to streptomycin, ampicillin and sulfanilamides on the physical map of conjugative R plasmid pBS52 of 38,000 bp which has a broad bacterial host range and belongs to a new incompatibility group. The plasmid has a natural "polylinker" site (less than 200 bp) containing (in the order of arrangement) the recognition sites for restriction enzymes: BamHI-EcoRI-PstI-EcoRV-BglII (PvuII). The comparative analysis shows that pBS52 contains a segment homologous to DNA of plasmid RSE1010 (IncP-4). The evolutionary origin of plasmid pBS52 is discussed. The recA-independent formation of the mini-derivatives of pBS373 and pBS374 types during the transformation of Escherichia coli with pBS52 plasmid DNA has been shown. Plasmids pBS373 and yBS374 are capable of autonomous replication in Pseudomonas putida and P. aeruginosa cells, which is provided by the rep system of IncP-4 replicon.     

钙离子在蟾蜍卵母细胞成熟分裂过程中的作用

缺钙处理的中华大蟾蜍卵母细胞,在孕酮作用下,仍能显示与恢复成熟分裂有关的早期启动变化,即卵内cAMP含量下降。但在缺钙条件下,孕酮不能促使卵母细胞进一步产生具有生物活性的促成熟因子,这可能与缺钙条件下卵母细胞内蛋白质磷酸化反应普遍低下有关。在外环境中有足量钙离子的条件下,即使无孕酮刺激,二价阳离子载体A_(23187)亦能诱发卵母细胞GVBD。这些结果无疑证明外源钙离子内流,以及由此可能导致的卵内游离钙离子增加,与卵母细胞恢复和完成成熟分裂有密切关系。     

Mutations of plasmid pBS286 blocking the initial stages of naphthalene oxidation induced by Tn5

A collection of Tn5 transposon Nah- mutants of the plasmid pBS286 was obtained. The insertion sites were localized and orientation of Tn5 determined. The mutants obtained were biochemically analyzed, the nah-region map of the plasmid being elaborated. Structural genes of the nah operon were shown to be organized similarly to those of the nah1 operon of the NAH7 plasmid discussed in the literature. The data obtained are in favour of the previously published information on the presence of elements operating the pBS286 plasmid. The results are given indicating a possibility of regulating the expression of catechol splitting meta-pathway genes with participation of products on early stages of naphthalene oxidation.     

Molecular genetic studies of the DNA promotor regions in Bacillus thuringiensis subsp. galleriae 69-6. I. Structural and functional analysis

Three recombinant plasmids pPBT9, pPBT10 and pPBT74 carrying promoter-containing regions of DNA of Bacillus thuringiensis which are responsible for the expression of the promoterless tet gene, were studied. In the in vitro experiments, it had been shown that these promoter-active HindIII fragments of bacillar DNA contained RNA polymerase binding sites. The AluI subfragments that specifically bind to Escherichia coli RNA polymerase promote the tet gene expression, similar to the whole HindIII fragments. Sequence analysis revealed that the approximately 220 base pair AluI subfragment of the bacillar insertion of the pPBT10 plasmid contained sites typical for "-10" and "-35" homology regions of promoters specific for sigma 55-RNA polymerase from Bac. subtilis. The 1.45 kb HindII bacillar fragment of the plasmid pPBT9 had three AluI subfragments that bind to E. coli RNA polymerase. Only approximately 400 base pair AluI subfragment among these restored the tet gene expression in vivo. Bireplicon pBP plasmids were constructed that promoted the expression of the enterobacterial antibiotic resistance gene under the control of Bac. thuringiensis promoters in Bac. subtilis cells.     

环境温度影响蟾蜍卵母细胞成熟能力的机制之实验分析

长年饲养在高温(28—30℃)环境中雌性中华大蟾蜍,它们的卵母细胞可以长足,但经激素处理时,生发泡不破裂,仅显示成熟过程早期阶段的变化。值得注意的是,在孕酮刺激后的高温卵卵质中,出观了一种能诱发低温卵恢复减数分裂的物质,称作为“依赖冬眠因子的促成熟物质”(HF-MPS)。HF-MPS 与MPF 有不少相似之处,如孕酮处理后,它们在卵质中出现的时间相仿,它们的形成均不依赖于转录水平,而是依赖于翻译水平的蛋白质合成活动;但亦存在不同之处,如MPF 诱发低温卵GVBD 时程不受温度影响,而HF-MPS 在10℃环境中,诱发低温卵GVBD 的时间明显延缓;MPF 不仅能诱发低温卵GVBD,而且同样能诱发高温卵GVBD,然而,HF-MPS 只能诱发低温卵GVBD。由此表明,MPF 和HF-MPS 似乎是截然不同的两类活性蛋白质。高温卵缺少低温诱发产生的“冬眠因子”,所以不能恢复cdc 2基因的转录活动,不能实现MPF 自身催化扩增作用,不能保证孕酮处理后的卵母细胞完成正常成熟的全过程变化。足见,低温是中华大蟾蜍卵母细胞恢复减数分裂过程中的必要条件,是导致中华大蟾蜍现有区域分布的内在原因之一。     

Synthesis of human insulin gene. VIII. Construction of expression vectors for fused proinsulin production in Escherichia coli

Analysis of Tn1725 insertions in the Pif+ plasmid pRS2496 showed the maximum limits of the F pif region to be between 43.7 and 47.15 on the 100-kb map of the F plasmid. The effect of these insertions on the expression of pif polypeptides indicated that two of the pif genes, pifA and pifC, lie within a polycistronic operon.     

Cloning and expression of Clostridium thermocellum F7 cellulase genes in Escherichia coli and Bacillus subtilis cells

The Clostridium thermocellum total DNA Sau 3A fragments' library was constructed on the basis of shuttle vector pMK4 for the Escherichia coli - Bacillus subtilis. 14 clones with endoglucanase activity and one with beta-glucosidase activity were selected in E. coli cells. Recombinant plasmids pCE were characterized by structural instability of various degree in B. subtilis cells. The results of the physical mapping, analysis of gene products in E. coli mini-cells as well as the DNA-DNA blot hybridization have led to conclusion on cloning of 7 individual genes for endoglucanases. Up to 3 polypeptides of various molecular weight corresponding to the products of cel gene were revealed in E. coli mini-cells containing the recombinant plasmids. The hybridization analysis demonstrated considerable homology of the majority of cel genes.     

不同发育时期中华绒螯蟹血淋巴渗透压的分析

中华绒螯蟹血淋巴渗透压随不同的生长阶段而异;不论是在淡水中,还是在不同盐浓度的海水中,都具有保持血淋巴高渗调节控制的能力。青春期前,6—9月份,血淋巴渗透压自390mOsm/Kg H_2O升至560mOsm/Kg H_2O;青春期后,9—12月份,自555mOsm/Kg H_2O升至656mOsm/Kg H_2O,但12月到翌年3月份却明显下降。雌蟹从淡水移入海水后的血淋巴渗透压调升速度,青春期后是青春期前的三倍左右。