Analysis of Schizosaccharomyces pombe mitochondrial DNA replication by two dimensional gel electrophoresis

The entire mitochondrial genome of Schizosaccharomyces pombe ura4-294h ^-was analyzed by the 2D pulsed field gel electrophoresis technique developed by Brewer and Fangman. The genome consists of multimers with an average size of 100 kb and analysis of the overlapping restriction fragments of the complete mitochondrial DNA (mtDNA) genome resulted in simply Y 2D gel patterns. Large single-stranded DNA molecules or double-stranded DNA molecules containing large or numerous single-stranded regions were found in the S. pombe mtDNA preparation. The replication of mtDNA monomers was found to occur in either direction. On the basis of these results, a replication mechanism for S. pombe mtDNA that is most consistent with a rolling circle model is suggested.     

Effects of quality and developmental stage on the survival of IVF-derived bovine blastocysts cultured in vitro after freezing and thawing

Factors affecting viability of IVF-derived bovine blastocysts after freezing and thawing were investigated. A total of 1,101 ova matured and fertilized in vitro were cultured under 2 different conditions, 1) in TCM-199 on granulosa cell monolayers at 5% CO(2) in air and 2) in synthetic oviduct fluid (SOF) medium without somatic cell support at 5% CO(2), 5% O(2), 90% N(2). All blastocysts that developed from the 2 different culture systems were individually classified into 4 grades of embryo quality and were then frozen by conventional slow freezing. Developmental rates of the IVF-derived ova to blastocysts and the survival rates of the frozen-thawed blastocysts were not different between the SOF medium (16 and 49%) and the co-culture system (13 and 61%, respectively). Survival of frozen-thawed blastocysts was affected by embryo quality in both the SOF and co-culture systems (P<0.001). Blastocysts produced in vitro were also individually classified into 3 developmental stages and were then cultured for 3 d in the co-culture system with granulosa cells after freezing and thawing. There was a difference in the survival rate of frozen-thawed embryos between blastocyst developmental stages (early vs mid, P<0.05; mid vs expanded, P<0.01; early vs expanded, P<0.001). The post-thawing survival rate of blastocysts frozen at Day 7 (62%) of culture was higher compared with that of Day 8 (45%), but there was no difference in survival rate between Day 7 and 8 of culture. The results indicate that the quality and developmental stage of blastocysts are important factors influencing their survival after freezing and thawing.     

Expression and secretion of human lipocortin-1 by promoter and signal sequence of STA1 from Saccharomyces diastaticus

Summary For the secretion of human lipocortin-1 (LC-1) in yeast, a expression and secretion vector was constructed by using the promoter and signal sequence of glucoamylase gene (STA1) of Saccharomyces diastaticus. After the cDNA of human LC-1 was ligated with the secretion vector, the resulting hybrid plasmid was transformed into S. diastaticus. When the recombinant S. diastaticus was cultivated in YPD medium, LC-1 was expressed and secreted into the extracellular medium, yielding LC-1 protein at a concentration of 2.5 g/mL.     

Rapid screening of an SH2-containing gene using PCR amplification method

Summary To isolate a novel gene that contains an SH2 domain, we devised a rapid and nonradioactive cDNA library screening method using polymerase chain reaction (PCR). For PCR amplification, we designed degenerate oligonucleotide primers from the multialigned DNA sequences of SH2 domains. This method offers an inexpensive and efficient approach for the isolation of clones of interest from cDNA libraries.     

Changes and Relationship of PAF and TNF in Rats with Myocardial Ischaemia and Reperfusion Injury

IN THIS STUDY IT IS REPORTED THAT: (1) the levels of blood platelet-activating factor and serum tumour necrosis factor significantly increased after coronary ligation and reperfusion, compared with sham-ligated controls, in an anaesthetized rat model; (2) compared with vehicle controls, pretreatment with the PAF antagonist BN 50739 (10 mg/kg, i.v.) produced significant decreases in infarct size (from 29.6 +/- 4.0% to 22.4 +/- 2.1%, p < 0.05 after 3 h ligation, and from 28.5 +/- 9.5% to 10.5 +/- 4.5%, p < 0.01 after 4 h reperfusion) and the level of serum TNF (from 10.4 +/- 7.7 U/ml to 3.9 +/- 4.8 U/ml, p < 0.05); and (3) a significan positive correlation was found between the level of blood PAF or serum TNF and infarct size. The present results indicate that PAF and TNF may be important mediators involved in myocardial ischaemia and reperfusion injury, and that PAF antagonists may exert a protective effect on ischaemic or reperfused myocardium by inhibiting the interaction of PAF and TNF.     

软紫草愈伤组织的初步培养

外植体来源不同的软紫草愈伤组织产生紫草色素的能力各异。６０Ｃｏ－ｒ射线辐照处理后的Ｈ２无性系愈伤组织生长量和色素产生均属上乘。改良ＭＳ基本培养基添加１毫克／升ＫＴ,０．５毫克／升ＩＡＡ,５％（Ｗ／Ｖ）蔗糖对愈伤组织培养较为适宜。马铃薯提取液对愈伤组织生长有明显的促进作用。     

三褶脉紫菀中的新二萜甙

三褶脉紫菀（ＡｓｔｅｒａｇｅｒａｔｏｉｄｅｓＴｕｒｃｚ．）系菊科多年生草本植物,遍布全国,是民间常用的中药,有清热解毒、祛痰镇咳的功效［１,２］。化学工作者们曾从其同属植物紫菀（ＡｓｔｅｒｔａｒａｒｉｃｕｓＬ．ｆ．）中分离到紫菀酮（ｓｈｉｏｎｏｎｅ）、槲皮素（ｑｕｅｒｃｅｔｉｎ）、无羁萜（ｆｒｉｅｄｅｌｉｎ）、表无羁萜（ｅｐｉｆｒｉｅｄｅｌｉｎｅｌ）、毛叶醇（ｌａｃｈｎｏｐｈｙｌｌｏｌ）、乙酸毛叶酯（ｌａｃｈｎｏｐｈｙｌｌｏｌａｃｅｔａｔｅ）、茴香醚（ａｎｅｔｈｏｌｅ）以及紫菀三萜皂甙［２—４］…     

解离增强镧系元素荧光免疫分析灵敏度的改进

为提高解离增强镧系荧光免疫分析(DELFIA)的灵敏度或信／噪比,进行了一些重要的方法学研究．观察到了对于不同的铕量,荧光响应和信／噪比都随着增强液体积而明显地变化．对于确定的铕量,存在一个最佳体积,且铕量越小,其最佳体积也越小．实验中选择最佳体积是重要的．研究了增强液制备技术,发展了微滴定板条有效的清洗和干燥方法,使本底荧光明显降低．     

磁水对蕃茄酯酶同工酶的影响

蕃茄（ＬｙｃｏｐｅｒｓｉｃｏｍｅｓｃｕｌｅｎｔｕｍＭｉｌｌ）在各生长发育期酯酶同工酶谱带无明显变化,但经磁水处理后各生长发育阶段功能叶中酯酶同工酶酶谱及磷素含量与对照组相比存在显著差异,且具有酶谱负极端带数增加,正极端带数减少的趋势。总磷含量测定结果表明处理组均比对照组显著提高。这些变化在磁水处理著前后代中不存在,故认为磁水只是致使非编码顺序局部复制引起ＤＮＡ分子高级结构改变或是通过活化调节基因来调控酯酶同工酶,活化磷代谢,达到促进生长和增产的效果,而并非导致可遗传变异的结果。     

Different Localization of Two Ca2+ in Spinach Oxygen-Evolving Photosystem II Membranes. Evidence for Involvement of Only One Ca2+ in Oxygen Evolution

Localization of the two Ca²⁺ bound to oxygen-evolving photosystemII (PSII) membranes from spinach was investigated by fractionatingthe membranes into the PSII reaction center core complexes andperipheral antenna Chl a/b-proteins after solubilization withn-heptylthioglucoside. The core complex fraction contained oneCa²⁺ per PSII, while another Ca²⁺ was found in the solubilizedmajor light-harvesting Chl a/b-proteins (LHCII). LHCII isolatedwith Triton X-100 or dodecylmaltoside also contained Ca²⁺ inan amount corresponding to one per PSII. The Ca²⁺ bound to LHCIIcould not be removed by treatment with Chelex 100, which effectivelysequestered extraneous Ca²⁺ bound to LHCII, or by preparationof LHCII in the presence of 40 mM citrate. Localization of thetwo Ca²⁺ in different functional domain of PSII membranes conclusivelyindicates that the number of the bound Ca²⁺ that can functionin oxygen evolution is one per PSII. The results also suggestthat one Ca²⁺ has a structural role in the peripheral antennaassembly. (Received July 21, 1992; Accepted March 9, 1993)