Speciation in the Artemia genus: Mitochondrial DNA analysis of bisexual and parthenogenetic brine shrimps

From the cloned mitochondrial DNAs (mtDNAs) isolated from two bisexual species, one Mediterranean, Artemia salina, and one American, Artemia franciscana, and two parthenogenetic (diploid and tetraploid) strains of Artemia parthenogenetica collected in Spain, physical maps have been constructed and compared. They are extremely different among themselves, much more than the differences between Drosophila melanogaster and D. yakuba and in the same range of different mammalian species such as mouse/rat or man/cow. The nucleotide sequences of two regions of mtDNA encoding parts of the cytochrome c oxidase subunit I (COI) and cytochrome b (Cytb) genes have been determined in the two bisexual species and the two parthenogenetic strains. Comparisons of these sequences have revealed a high degree of divergence at the nucleotide level, averaging more than 15%, in agreement with the differences found in the physical maps. The majority of the nucleotide changes are silent and there is a strong bias toward transitions, with the CT substitutions being highly predominant. The evolutionary distance between the two Artemia parthenogenetica is high and there is no clear relationship with any of the bisexual species, including the one present nowadays in Spain. Using a combination of molecular (mtDNA) and morphological markers it is possible to conclude that all of these Artemia isolates should be actually considered as belonging to different species, even the two Artemia parthenogenetica diploidica and tetraploidica.On sabbatical leave from Departamento de Bioquímica, Facultad de Veterinaria, Universidad Complutense de Madridearly Italian artemiologists to designate the Medi-Beatriz Batuecas died in an accident during the Christmas holy days of 1988 after she had initiated this workCorrespondence to: R. Garesse     

Evidence for a life span-prolonging effect of a linear plasmid in a longevity mutant of Podospora anserina

The linear mitochondrial plasmid pAL2-1 of the long-lived mutant AL2 of Podospora anserina was demonstrated to be able to integrate into the high molecular weight mitochondrial DNA (mtDNA). Hybridization analysis and densitometric evaluation of the mitochondrial genome isolated from cultures of different ages revealed that the mtDNA is highly stable during the whole life span of the mutant. In addition, and in sharp contrast to the situation in certain senescence-prone Neurospora strains, the mutated P. anserina mtDNA molecules containing integrated plasmid copies are not suppressive to wild-type genomes. As demonstrated by hybridization and polymerase chain reaction (PCR) analysis, the proportion of mtDNA molecules affected by the integration of pAL2-1 fluctuates between 10% and 50%. Comparative sequence analysis of free and integrated plasmid copies revealed four differences within the terminal inverted repeats (TIRs). These point mutations are not caused by the integration event since they occur subsequent to integration and at various ages. Interestingly, both repeats contain identical sequences indicating that the mechanism involved in the maintenance of perfect TIRs is active on both free and integrated plasmid copies. Finally, in reciprocal crosses between AL2 and the wild-type strain A, some abnormal progeny were obtained. One group of strains did not contain detectable amounts of plasmid pAL2-1, although the mtDNA was clearly of the type found in the long-lived mutant AL2. These strains exhibited a short-lived phenotype. In contrast, one strain was selected that was found to contain wild-type A-specific mitochondrial genomes and traces of pAL2-1. This strain was characterized by an increased life span. Altogether these data suggest that the linear plasmid pAL2-1 is involved in the expression of longevity in mutant AL2.     

Molecular identification of the ten subunits of cytochrome-c reductase from potato mitochondria

Cytochrome-c reductase (EC 1.10.2.2.) from Solanum tuberosum L. comprises ten subunits with apparent molecular sizes of 55, 53, 51, 35, 33, 25, 14, 12, 11 and 10 kDa on 14% SDS-PAGE. The identity of the subunits was analysed by direct amino-acid sequencing via cyclic Edman degradation. A large-scale purification procedure for the enzyme complex based on affinity chromatography and gelfiltraton is described. All subunits were enzymatically fragmented and the generated peptides were separated by reverse-phase HPLC. Complete or partial sequence determination of 33 peptides comprising a total of nearly 500 amino acids showed, that cytochrome-c reductase from potato contains three respiratory proteins (cytochrome b, cytochrome c ₁ and the Rieske iron-sulfur protein), four small proteins with molecular sizes below 15 kDa (so-called Q-binding, hinge, cytochrome-c ₁-linked and core-linked proteins) and three proteins in the 50-kDa range which show similarity to members of the core/PEP/MPP protein family (core/processing enhancing protein/mitochondrial processing peptidase). In fact these subunits show highest sequence identity either to MPP or PEP, which is in line with earlier findings, that isolated cytochrome-c reductase from potato exhibits processing activity towards mitochondrial precursor proteins.Abbreviations MPP mitochondrial processing peptidase - PEP processing enhancing protein This research was supported by the Deutsche Forschungsgemeinschaft.     

杂交水稻及其“三系”线粒体DNA的AP—PCR指纹图谱

为了研究水稻（Ｏｒｙｚａ ｓａｔｉｖａ Ｌ．）细胞质雄性不育（ＣＭＳ）与线粒体基因组的关系,应用ＡＰ－ＰＣＲ 分析,用７ 个任意单引物对６ 种水稻品系线粒体ＤＮＡ 进行了扩增。水稻线粒体ＤＮＡ 的ＡＰ－ＰＣＲ 产物可分为三种类型：（１）所有供试品系均能扩增的片段,它们代表了线粒体ＤＮＡ 在进化上的保守性序列。有４ 个引物检测到这类片段。（２）２ 个以上水稻品系共同出现而在全部供试材料间存在差异的扩增片段,这类片段是检测水稻线粒体ＤＮＡ多态性的主要来源。（３）一种细胞质类型所特有的扩增片段,从引物Ｒ２ 和Ｖ５ 的扩增产物中发现了这类片段,它们可能与ＣＭＳ有关联。另外,ＷＡ型不育系珍汕９７Ａ 与其杂种之间在６ 个引物的扩增图谱上均存在不同程度的差异,说明两者的线粒体ＤＮＡ序列结构可能存在某种差别     

细胞电穿孔动态过程的荧光测量

利用改进后的Ｔｈ／ＤＰＡ荧光方法及探针ＥＢ对人血影及大鼠骨髓细胞电穿孔的动态过程及其与电脉冲参数的关系进行了系统的研究．测量结果表明,在临界点以上电场作用下,血影电穿孔在电击后０．２—０．３ｓ时达最大,在约０．８ｓ时愈合;而大鼠骨髓细胞电穿孔在电击后０．４—０．９ｓ达到最大,３－５ｓ左右愈合;电穿孔大小及扩大、愈合速率与电脉冲参数有关。１０－４０ｍｍｏｌ／Ｌ乙醇和５－２０ｍｍｏｌ／Ｌ成二醛抑制血影对Ｔｂ３＋离子的电通透,相同浓度的成二醛作用强于乙醇。这些结果将为电穿孔技术的合理应用提供参考。     

Further characterization of the respiratory deficient dum-1 mutation of Chlamydomonas reinhardtii and its use as a recipient for mitochondrial transformation

Summary The respiratory deficient dum-1 mutant of Chlamydomonas reinhardtii fails to grow in the dark because of a terminal 1.5 kb deletion in the linear 15.8 kb mitochondrial genome, which affects the apocytochrome b (CYB) gene. In contrast to the wild type where only mitochondrial genomes of monomer length are observed, the dum-1 genomes are present as a mixture of monomer and dimer length molecules. The mutant dimers appear to result from head-to-head fusions of two deleted molecules. Furthermore, mitochondrial genomes of dum-1 were also found to be unstable, with the extent of the deletion varying among single cell clones from the original mutant population. The dum-1 mutant also segregates, at a frequency of ca. 4% per generation, lethal minute colonies in which the original deletion now extends at least into the adjacent gene encoding subunit four of NAD dehydrogenase (ND4). We have used the dum-1 mutant as a recipient to demonstrate stable mitochondrial transformation in C. reinhardtii employing the biolistic method. After 4 to 8 weeks dark incubation, a total of 22 respiratory competent colonies were isolated from plates of dum-1 cells bombarded with C. reinhardtii mitochondrial DNA (frequency 7.3 × 10^–7) and a single colony was isolated from plates bombarded with C. smithii mitochondrial DNA (frequency 0.8 × 10^–7). No colonies were seen on control plates (frequency < 0.96 × 10^–9). All transformants grew normally in the dark on acetate media; 22 transformants were homoplasmic for the wild-type mitochondrial genome typical of the C. reinhardtii donor. The single transformant obtained from the C. smithii donor had a recombinant mitochondrial genome containing the donor CYB gene and the diagnostic HpaI and XbaI restriction sites in the gene encoding subunit I of cytochrome oxidase (COI) from the C. reinhardtii recipient. The characteristic deletion fragments of the dum-1 recipient were not detected in any of the transformants.     

Functional mitochondria in snake Bothrops alternatus erythrocytes and modulation of HbO2 affinity by mitochondrial ATP

Digitonin was applied to permeabilize the plasma membrane of Bothrops alternatus erythrocytes to study respiration, oxidative phosphorylation and Ca²⁺ transport by mitochondria in situ. These mitochondria oxidized added NAD-linked substrates, succinate and N,N,N, N-tetramethyl-p-phenylenediamine. Respiration was sensitive to rotenone and cyanide but not to antimycin A. This indicates that Bothrops mitochondria possess the respiratory complexes NADH-ubiquinone, succinate-ubiquinone, and ferrocytochrome c-oxygen oxidoreductases, although the lack of sensitivity to antimycin A raises doubt about the composition of the ubiquinol cytochrome c-reductase complex. An ability to build up and sustain a membrane potential was documented by their capacity to accumulate tetraphenylphosphonium and Ca²⁺ through an uncoupler-sensitive mechanism. Addition of ADP caused a transient decrease in the membrane potential, indicating that this is the predominant driving force for ATP synthesis as in most types of mitochondria. Uncoupling of phosphorylation from the oxidative process increased hemoglobin O₂ affinity, which suggests that ATP production by mitochondria may participate in modulation of O₂ transport by hemoglobin.Abbreviations membrane potential - BAE Bothrops alternatus erythrocytes - DNP 2,4-dinitrophenol - DPG 2,3-diphosphoglycerate - EGTA ethyleneglycol tetra-acetic acid - FCCP carbonylcyanide p-trifloromethoxyphenylhydrazone - TMPD N,N,N,N-tetramethyl-p-phenylenediamine - TPP⁺ tetraphenylphosphonium - TRIS tris-(hydroxymethyl)aminomethane     

一类单种群动态模型的稳定性

一、引言及引理考虑一类单种群动态模型对方程（1）总假设满足其中N（t）为种群在时刻t的数量或密度．由于方程（1）是描述种群增长的生态模型,因此我们考虑（1）的正解且满足初始条件：易证（1）和（2）有唯一正常数平衡点本文研究（1）关于N*的稳定性．（1）的特别情形是其中（1a）描述在食物资源严重不足种群数量充分大导致种群内部互相竞争残杀时的增长模型［1];（1b）描述种内既有竞争又有协作的所谓ALLEE效应的增长模型［2]．事实上,若β＞0,由（1b）式知,当N（t-r）充分小时,当N（t—r）充分大时,这正是ALLEE效应．文献…