Stimulation of somatic embryo formation by mutagens and darkness in culture of immature zygotic embryos of Arabidopsis thaliana (L.) Heynh.

The aim of the study was to evaluate the influence of light conditions,physical state of the induction medium and the mutagenic treatment on theembryogenic ability of Arabidopsis thaliana (L.) immaturezygotic embryos differing in developmental stage. The efficiency of directsomatic embryogenesis (DSE) was analysed in a culture of immature zygoticembryos at an early (ES) and a late (LS) developmental stage. The efficiency ofDSE was scored as a percentage of the explants producing somatic embryos. Theexperiments indicated that the physical state of the induction medium (solid orliquid) did not influence the embryogenic ability of the cultured explants. Inthe cultures on both solid and liquid induction medium, the ES explantsproducedsomatic embryos with a frequency of 25.8–37.3% i.e. 2.5–3-timeslower than LS explants. However, an increase in the embryogenic ability of ESexplants (up to 69.8%) was observed when DSE was induced in darkness. Moreover,the stimulation of DSE efficiency in culture of ES explants was also observedafter mutagenic treatment. The chemical mutagens, MNH and EMS, applied forexplant treatment, both stimulated efficiency of somatic embryo formation inculture of ES explants. The most effective DSE induction was observed when MNHand EMS were applied in doses of 0.125–1.0 mM × 3h and0.05–0.2% × 18h, respectively. In these treatment combinations thefrequency of ES explants forming somatic embryos was found to be about 2 timeshigher than in the control culture.     

Isolation and characterisation of an antifolate insensitive (afi1) mutant of Arabidopsis thaliana

Antifolates can impair the synthesis and/or function of folates in living organisms. Mechanisms of resistance or tolerance to antifolates have been mainly described in plants using the drug methotrexate. In this work, the antifolate trimethoprim (TMP) was used with the aim of revealing a novel mechanism of resistance. EMS mutagenised seeds from Arabidopsis were screened to isolate individuals insensitive to TMP. Genetic analysis revealed a homozygous recessive mutation that segregates with the phenotype of tolerance to 50 μm TMP. Mapping analysis localised the mutation at the end of the short arm of chromosome 3. Preliminary characterisation demonstrated up‐regulation of several genes from the folate biosynthetic pathway in the TMP insensitive mutant, and a slight increase in total folate content in the mutant as compared with the Col‐0 control. Moreover, sequence analysis of the DHFR (dihydrofolate reductase) genes, which encode a known target for resistance to antifolates, did not reveal any changes. This study is the first report of a stable mutant insensitive (afi1) to the antifolate trimethoprim in plants, and suggests the existence of a novel mechanism of resistance to antifolates.     

Metronidazole (Flagyl): lack of induction of sister-chromatid exchanges in CHO-K1 cells

328 X-linked recessive lethal mutations induced in late spermatids by hycanthone methanesulfonate were tested for coverage by duplications that comprised, in total, about 24% of the euchromatic X chromosome; 78 lethals appeared to be covered. Crossover localization tests of a random sample of 38 non-covered lethals revealed 4 chromosomes carrying a lethal within a duplicated segment. Lethals localized to a particular region were crossed to reference deficiencies and single-locus mutations, and inter se, to ascertain their genetic extent. The proportion of multi-locus deletions among these 78 covered and 4 non-covered lethals was 3/48, 1/10 and 13/24 for the distal, medial and proximal regions, respectively. A storage period of 9 days did not noticeably influence these proportions. In the sample of 38 non-covered lethals, and among 17 of the covered single-site lethals, 4 cases of strong crossover suppression were detected. Comparison of these results with data obtained with other mutagens suggests that induction of multi-locus deletions, and possibly of other types of chromosome rearrangement, could in part depend on other mechanisms than those acting in the formation of translocations and chromosome loss. For the purpose of mutagen testing, these findings imply that, in Drosophila, results in the regular genetic tests for chromosome breakage events do not always accurately predict the capacity of a mutagen to induce multi-locus deletions. This is of importance since transmissible multi-locus deletions have been considered a significant source of genetic damage in man.     

景观生态学的实验研究方法综述

实验是生态学研究的基本途径之一 ,对其理论的检验和发展至关重要。 2 0世纪 80年代后期 ,实验研究途径开始应用于景观生态学 ,至今方兴未艾。近年来 ,“景观”的生态学内涵更加强调“不同尺度上的空间异质性”。而研究对象的尺度和空间异质性往往正是景观生态学实验设计和操作所面临的困难。自然界景观格局与过程中客观存在的自相似结构为景观生态实验设计提供了可行性。现有的景观生态学实验方法可分为 3类 :野外比较观测性实验仍是目前应用较多的方法 ;操作性实验设计更严密 ,结果更可靠 ,但受现实条件的限制更大 ;计算机模拟实验是克服实验条件的困难的一个替代途径 ,并对理论的检验与发展特别有用。这 3类实验方法各自存在不同的优势和局限 ,彼此难以替代。景观生态实验方法主要从种群、群落和生态系统实验中借鉴并发展而来 ,但其实验设计在科学问题和操作尺度上具有显著的特点。从空间范畴来讲 ,景观生态实验包含斑块、边界、景观、斑块景观 4种 ,其所对应的生物群体组织水平、实验设计所涉及的问题和解决的方法都有所不同。多尺度的对比实验有助于了解生态现象与机制的尺度推移规律 ,是目前实验研究的难点和焦点之一。实验模型系统 (EMS)途径来源于生态系统实验。由于它有对自然因素更多的保留和     

Induction of mutation to streptomycin resistance in Micrococcus radiodurans.

No detectable induction of mutation to streptomycin resistance could be used in wild-type Micrococcus radiodurans and its radiation-sensitive and super-resistant mutants by ionizing or UV-radiation. N-methyl-N'-nito-N-nitrosoguanidine (NTG) was mutagenically active. The results suggest that repair of radiation-damaged DNA in Micrococcus radiodurans is mutation-proof.     

Determination of radiation equivalence of ethyl methanesulphonate (EMS) for induction of gene conversion in diploid yeast.

A unit Rad-Equivalent Chemical (REC) has been suggested for purposes of quantitating the mutagenic hazards of chemicals. The usefulness of this approach is demonstrated by the establishment of a constant relationship between the forward mutation frequency and haploid genome size in various organisms for both radiation and chemical EMS. However, it is necessary to determine the radiation equivalence of chemicals in as many organisms and for as many end-points as possible. For end-points we are limited to forward mutations. Another relevant genetic end-point of interest in this regard is gene conversion which can also monitor any kind of DNA damage in a suitable diploid system. Hence, we have determined the REC value for EMS in diploid yeast with gene conversion as the end-point. This agrees well with the REC values estimated in a number of organisms with forward mutation as the end-point. This finding further underlines the generality of the REC concept.     

The use of different test systems with yeasts for the evaluation of chemically induced gene conversions and gene mutations

The wide variety of genetic alterations that can be induced in human populations when exposed to chemical genotoxic substances present in our environment may be predictable using laboratory organisms such as yeasts.In the present paper methodologies are described for analysing the genetic effects induced by a well known chemical mutagen, ethyl methanesulfonate (EMS). Haploid or diploid yeast cells have been treated in vitro, in buffer or in the presence of mouse liver microsomes, and in vivo, in the peritoneum of the mouse (host-mediated assay).With these different methods of assaying the genetic activity of a compound, its metabolic activation occuring in the mammalian body is taken into account: this might lead to a more reliable extrapolation of data from laboratory experiments to man.The relationships between doses and frequencies of the induced genetic effects are described by equations obtained after regression analysis of the data, thus allowing a quantitative comparison among different methodologies and different genetic systems.One genetic system analyzed is represented by forward-mutations scored phenotypically on a non selective medium. Mutations induced in five loci with different sensitivity and average data of mutation-induction per locus have been derived. The second genetic system was provided by scoring on a selective medium mitotic gene-conversions induced in two loci with different kinetics.Haploid cells of Schizosaccharomyces pombe and diploid cells of Saccharomyces cerevisiae were submitted to analysis for the evaluation of gene-mutations and gene-conversions respectively.     

TILLING在水稻育种中的应用前景

TILLING(Targeting Induced Local Lesions in Genomes)是功能基因组研究中应用的一种反向遗传学技术。它能高通量低成本地在EMS诱变群体中鉴定出发生在特定基因上的点突变。在其基础上发展出的EcoTILLING技术则可发现种质资源中的SNP位点及小插入或缺失多态性位点。水稻是非常重要的粮食作物, 也是已经完成了全基因组序列测定,有丰富的生物信息学资源可以利用的基因组研究模式植物。水稻的分子标记辅助育种将在育种中扮演越来越重要的角色。在这样的背景下,本文从基于特定基因的种质资源鉴定、EMS诱变育种、及水稻功能标记开发等方面论述了其在水稻育种中的应用前景。     

低双乙酰啤酒酵母菌株BEZ112的选育

以啤酒酿造生产菌株啤酒酵母(Saccharomyces cerevisiae)FB作为出发菌株,用甲基磺酸乙酯(EMS)诱变,经分离筛选得到一株优良的啤酒酵母菌株BEZ112。该菌株的絮凝性、发酵度、酒精度、发酵液的总酯和总高级醇的含量等特性保持了亲株的优良性状。但以12°Bx麦芽汁为培养基用500mL三角瓶在12℃下发酵,该菌株发酵至第4d,发酵液中的双乙酰含量达到峰值(0.291mg/L),比出发菌株FB发酵4d的峰值降低了30％,发酵至第8d,BEZ112发酵液中的双乙酰含量比出发菌株FB的降低了23％。以12°Bx麦芽汁为培养基用500L罐在12℃下发酵8d,BEZ112发酵液中的双乙酰含量(0.091mg/L)比出发菌株FB的(0.124mg/L)降低了27％。发酵得到的啤酒口感纯正清爽。