酵母菌支序分类研究

本文应用支序分类法(Cladistics)对16个属的酵母进行了系统演化研究,酵母菌的分类已有多年历史,但由于形态差异很大,所以在分类上一直存在着不同的看法,亦即存在着不同的分类体系。本文研究的隐球酵母等七个属在分类地位上也有争议,有的学者认为应属于子囊菌纲,有的学者认为应属于半知菌类。本文应用支序分类方法,通过诸多遗传性状的综合系统分析,认为上述7个属的酵母从系统进化和亲缘关系上,应该与子囊菌酵母同属一个发育系统,是与子囊菌酵母平行发展的一类酵母。     

Assessment of Cyto‐ and Genotoxic Effects of a Variety of Chemicals Using Saccharomyces cerevisiae

Saccharomyces cerevisiae as the most simple eukaryotic organism is broadly accepted as a laboratory model organism. For the detection of potential toxic effects of pure compounds and complex composed samples like wastewater a miniaturised short‐term in vitro cyto‐ and genotoxicity screening assay was developed. The assay based on genetically modified S. cerevisiae cells deleted in the prominent drug efflux transporters Pdr5, Snq2, and Yor1 that facilitate pleiotropic drug resistance. The yeast strain devoid of these proteins that mediate the efflux of structurally diverse hydrophobic compounds exhibited an increased sensitivity to a variety of organic compounds. The DNA damage inducible RAD54 promoter fused to a yeast optimized derivative of the GFP (green fluorescent protein) gene from the jelly fish Aequorea victoria served as an indicator of DNA damage in this strain. Various pure compounds including the direct‐acting genotoxins methyl‐N‐nitro‐N‐nitrosoguanidine (MNNG), 4‐nitroquinoline‐N‐oxide (4‐NQO), methyl methanesulfonate (MMS) and hydroxyurea as well as the heavy metals cadmium and chromium(VI), the insecticide lindane and the central nervous system stimulant caffeine were tested exhibiting dose dependent induction of green fluorescence. All compounds were in parallel examined for chronic toxicity. A bioassay detecting simultaneously geno‐ and cytotoxic effects of potential toxicants in a single assay can be an important tool with a variety of applications in environmental monitoring and aquatic ecotoxicology. By partial automation and miniaturisation to microtitration scale this bioassay enables sensitive and fast biomonitoring for a multitude of samples.     

草鱼呼肠孤病毒096vp7基因生物信息学分析及其酵母表达载体的构建

草鱼呼肠孤病毒（Grass carp reovirus, GCRV）是草鱼出血病的病原.从患病草鱼体内分离到一株草鱼呼肠孤病毒GCRV 096,经反转录PCR,克隆得GCRV 096 长度为855 bp 的vp7 基因,并分析该基因编码蛋白的特性.结果表明,同一基因型分离株VP7 蛋白间的差异不大,但不同基因型分离株VP7 蛋白间存在很大的差异.对GCRV 096 VP7 蛋白生物信息学分析表明,信号肽位点位于20 氨基酸处,且VP7 含有4个潜在的抗原决定簇.构建GCRV 096 vp7 基因的酵母表达载体pGBKT7-S10,并成功转化至酵母中.     

酵母双杂交系统的改进与研究进展

酵母双杂交系统是一种新兴的体内研究蛋白质之间相互作用以及筛选cDNA文库的技术手段,自建立以来得到了不断的改进与发展。在此基础上又相继出现了单杂交、三杂交、反向双杂交以及SOS富集系统等多种衍生系统,它们在功能基因组学、蛋白质组学以及药物开发研究中发挥了重要作用。本文拟就双杂交系统的原理、改进、发展及应用做一综述。     

酵母双杂交系统的应用及发展前景

酵母双杂交系统有利于检测蛋白质与蛋白质、蛋白质与DNA 、蛋白质与RNA和蛋白质与小分子之间的相互作用,因而对分子调控体系的研究起巨大的反 作用。文中系叙述了基于Ga14系统的双杂交技术的应用及其局限性,还介绍了单杂交系统、 双杂交系统、三杂交系统以及Sos恢复系统(SRS)。与其它领域一样,酵母双杂交系统将会在 海洋分子生物学研究中取得成功。     

Preliminary study on plasma membrane fluidity of Psychrophilic Yeast Rhodotorula sp.NJ298 in low temperature

The ability of cell to modulate the fluidity of plasma membrane was crucial to the survival of microorganism at low temperature. Plasma membrane proteins, fatty acids and carotenoids profiles of Antarctic psychrophilc yeast Rhodotorula sp. NJ298 were investigated at -3 ℃, 0 ℃ and 8 ℃. The results showed that plasma membrane protein content was greater at -3 ℃ than that at 8 ℃, and a unique membrane polypeptide composition with an apparent molecular mass of 94.7 kDa was newly synthesized with SDS-PAGE analysis; GC analysis showed that the main changes of fatty acids were the percentage of unsaturated fatty acids (C18∶ 1 and C18∶ 2) and shorter chain saturated fatty acid (C10∶ 0) increased along with the decrease of the culture temperature from 8 ℃ to -3 ℃; HPLC analysis indicated that astaxanthin was the major functional carotenoids of the plasma membrane, percentage of which increased from 54.6±1.5% at 8 ℃ to 81.9±2.1% at -3 ℃. However the fluidity of plasma membrane which was determined by measuring fluorescence anisotropy was similar at -3 ℃, 0 ℃ and 8 ℃. Hence these changes in plasma membrane's characteristics were involved in the cellular cold-adaptation by which NJ298 could maintain normal plasma membrane fluidity at near-freezing temperature.     

Food Type Effects on the Population Growth Patterns of Littoral Rotifers and Cladocerans

Littoral rotifers and cladocerans feed on bacteria, phytoplankton, and detritus. In comparison to planktonic organisms their species diversity is high, yet little is known about their biology. In this study we present data on the functional response of the cladocerans Macrothrix triserialis and Alona rectangula, and the rotifer Brachionus macracanthus to different levels of the alga Chlorella vulgaris. Based on the incipient limiting level, we chose the food concentration and tested the population growth on four types of diets for three cladoceran species — Macrothrix triserialis, Alona rectangula, and Chydorus sphaericus — and three rotifer species — Brachionus macracanthus, Lecane quadridentata, and Platyias quadricornis. We studied growth rates of each of these species on diets of live Chlorella vulgaris, heat‐killed Chlorella vulgaris, baker's yeast (Saccharomyces cerevisiae), and field collected detritus. The incipient limiting level was around 0.9 · 10⁶ cells mL^?1 of Chlorella for the tested zooplankton species. The population growth patterns showed that, in general, the cladocerans took 25…50 d to reach their peak population densities, which ranged from 10…75 ind. mL^?11. Among the rotifers L. quadridentata and B. macracanthus had equally high growth rates on live and heat‐killed forms of C. vulgaris while P. quadricornis grew best on detritus. The growth rates ranged from 0.03…0.21 d^?1. The similarities and differences between littoral and planktonic organisms in relation to food type and availability have been discussed.     

Occurrence of estrogen-like substances in the marine environment of the Northern Mediterranean Sea

Evidence has been presented that a number of environmental agents perturbs the function of the sex hormone signalling pathways in marine animals.

In this work the estrogenic/antiestrogenic activity of coastal marine water samples from five sites of the Northern Mediterranean Sea was assayed using a recombinant Saccharomyces cerevisiae yeast strain (RMY326 ER-ERE) transfected with the human estrogen receptor . Extraction of potential EDCs from seawater was performed in columns packed with XAD-2 resins.

The estrogenic activity was calculated as percentage of the activity of 17β-estradiol (10 nM) and it ranged from 4.8% and 59.03%, significantly depending on sampling site (p = 0.0013) and season (p < 0.05). Antagonistic activity of extracts was also detected and the percentage of inhibition of estradiol-dependent β-galactosidase induction ranged to 52.8%. These results point the reliability of the yeast assay as a first level screening test to assess the quality of aquatic environments.