Genome Analysis of Lotus japonicus

Lotus japonicus , a model legume plant, was reviewed and compared with Medicago truncatula and soybean. Several mutant libraries are being analyzed, focusing on the nodulation mechanism. The first plant nodulation gene nin was cloned by Ac-transposon tagging. Soybean remains as the most studied legume, especially in relation to the disease resistance genes. However, Lotus japonicus offers several advantages for molecular genetics, and the remained lackings were recently filled up, namely 1) an appropriate crossing partner for Gifu, accession Miyakojima, was proposed for its 4% polymorphism and smooth recombining ability; 2) a genome library with long inserts, average of 140 kb, and 8.2 genome equivalents of library size, has been established; and 3) the rather low polymorphic rate between Gifu and Miyakojima can be overcome with the HEGS (High Efficiency Genome Scanning). With this infrastructure, positional cloning of the causative genes of several mutant libraries will be accomplished in a short term. Genome sizes of L. japonicus acc. Gifu and Miyakojima were determined with high accuracy, to be 494±0 MB and 512±1 MB, respectively. The feasibility of constructing a physical map of the entire genome, for functional genomics, was discussed. Received 5 September 2000/ Accepted in revised form 11 October 2000     

Large Scale Structural Analysis of cDNAs in the Model Legume, Lotus japonicus

Lotus japonicus possesses certain characteristics suited to molecular genetic and genomic analyses and has been adopted as a model species in the study of legume plants. To make a catalogue of genes expressed in L. japonicus and understand biological processes specific to legume plants, large scale EST analyses have been performed. To date, more than 26,000 EST sequences of L. japonicus have been deposited in the public databases. These sequences were developed by five laboratories using different organs. In this review, information obtained from two EST projects carried out in Japan is presented. Some 7137 non-redundant EST groups from young plants and 718 groups from flower buds were generated. A similarity search revealed that homologues of nodulin genes in other legume plants, as well as genes related to secondary metabolism, seed development and the reproductive process, were included in the EST collection, indicating the usefulness of the EST clones in the study of biological phenomena distinctive to legume plant species. Received 23 August 2000/ Accepted in revised form 22 September 2000     

Snails can survive passage through a bird’s digestive system

Aim Predation is generally viewed as a factor that limits the distribution of animal prey species. However, in certain instances, such as seed dispersal, predation may enhance the dispersal capability of prey species. In a prior study, we found that land snails are preyed upon by the Japanese white‐eye (Zosterops japonicus) and the brown‐eared bulbul (Hypsipetes amaurotis) in the Ogasawara Islands. In this paper we provide experimental and field evidence indicating that land snails could potentially be dispersed through bird predation. Location Hahajima Island of the Ogasawara Islands in the western Pacific. Methods Experimentation was first performed to test whether the land snail Tornatellides boeningi could remain alive after being swallowed and passed through the bird digestive system. Next, in order to investigate the potential role of internal bird transport and dispersal of this snail, we investigated the relationship between the distribution of population genetic diversity in the snail and the regional geographical abundance of predatory birds. The population genetic structure of T. boeningi and isolation by distance were inferred with Arlequin . The association between nucleotide diversity in T. boeningi populations and population density of predators was examined using a generalized linear mixed model. We conducted a likelihood ratio test for the full model and for another model that removed the fixed effect. Results Of the 119 snails fed to Japanese white‐eyes and 55 snails fed to brown‐eared bulbuls, 14.3% and 16.4% of the snails, respectively, passed through the gut alive. Additionally, one snail gave birth to juveniles after emerging from a bird’s gut. Significant heterogeneity among the populations of T. boeningi on Hahajima was indicated using AMOVA; however, there was no evidence of isolation by distance. A positive correlation was found between levels of mitochondrial DNA variation among and within T. boeningi populations and the density of Japanese white‐eyes in the wild. Main conclusions Bird predation appears to be a method of dispersal for T. boeningi, and our results suggest that bird‐mediated dispersal plays a role in land snail population structure.     

Classification,inhibition, and specificity studies of the vitelline coat lysin from toad sperm

The sonicated supernatant of the sperm of the toad, Bufo japonicus, can digest easily the vitelline coat (VC) of uterine eggs, and to a lesser extent the VC of coelomic eggs, but not that of activated eggs. The VC lysis and fertilization were competitively inhibited in the presence of t-butyloxycarbonyl-L-Gln-L-Arg-L-Arg-4-methylcoumaryl-7-amide (Boc-Gln-Arg-Arg-MCA), suggesting the involvement of proteases in the fertilization process. Starting from a sonicated supernatant, a potent VC lysin, possessing hydrolytic activity on Boc-Gln-Arg-Arg-MCA, was obtained by anion-exchange chromatography and gel filtration. The activity of the partially purified lysin was inhibited by diisopropyl fluorophosphate (DFP) and by such trypsin inhibitors as soybean trypsin inhibitor, leupeptin, and (p-amidinophenyl) methanesulfonyl fluoride hydrochloride, but not by chymostatin, E-64, and ethylene glycol bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid. The molecular weight of the lysin was estimated to be 32K, based on the fluorographic image of ³H-DFP binding to the lysin on sodium dodecyl sulfate gel electrophoresis. The VC lysin was most active at pH 7.0–7.6 and under low ionic strength equivalent to fresh water. The release of the VC lysin was induced upon incubation of sperm with the contents of oviducal pars recta granules (PRG), which are known to induce the acrosome reaction. We conclude that the protease studied here represents the VC lysin of toad sperm that is involved in fertilization by digesting the VC of uterine eggs, probably released as a result of the acrosome reaction induced by PRG.     

Effects of interspecific competition on development and reproduction in two giant water bugs,Diplonychus japonicus vuillefroy andDiplonychus major esaki (Hemiptera: Belostomatidae)

We studied the interpecific competition between 2 species of predatory aquatic bugs, Diplonychus japonicus and D. major by conducting a field experiment. We set up 3 types of experimental plots in the paddy fields where D. major predominated. The two plots contained single species of either D. japonicus or D. major, respectively, and one plot had both species in equal number. We compared the development and the reproductive performance between plots in each species. In D. japonicus, the number of eggs and early instar nymphs were significantly smaller in the plots containing both species than in the monospecific plots. However, the numbers of late instar nymphs and newly emerged adults were not significantly different between plots. The proportions of starved nymphs in both plots were larger than those in the D. japonicus's natural habitats. The final densities of adults in both plots were lower than those in the natural habitats. These results suggest that lower density of D. japonicus in these paddy fields is due to the lack of available food for nymphs rather than the effects of interspecific competition with D. major. In D. major, significant differences were not found in the number of eggs, each instar nymphs and adults. These results suggest that the effects of interspecific competition did not affect the reproductive performance of D. major.     

识别未衍生化的13—羟化GAs及其葡萄糖苷的单克隆抗体

抗ＧＡ３ 及其葡萄糖苷的ＭＡＢ１０单克隆抗体源于以ＧＡ３ 中的３ 位羟基（３－ＯＨ）为偶联位点,人血清白蛋白（ＨＳＡ）为载体合成的ＧＡ３－３－ＨＳＡ 免疫原． 该抗体对１３－羟化ＧＡｓ（１３－ＯＨＧＡｓ、ＧＡ１、ＧＡ３、ＧＡ５ 等）和ＧＡ３ 葡萄糖苷具有高亲和力． ７ 位羧基的甲酯化可显著降低ＭＡＢ１０ 对１３－ＯＨ ＧＡｓ的亲和力,而３－ＯＨ 的糖苷化却未降低其亲和力． 用该抗体建立的两种分别用于ＧＡ３ 及其葡萄糖苷测定的酶联免疫吸附法（ＥＬＩＳＡ）,其检测线性范围均为０．２～２０ ｐｍ ｏｌ． 借助这两种ＥＬＩＳＡｓ,研究了羊蹄（Ｒｕｍ ｅｘ ｊａｐｏｎｉｃｕｓ）叶片中ＧＡ３ 及其类似ＧＡｓ和葡萄糖苷的动态变化．结果表明,叶片衰老与游离态ＧＡｓ的糖苷化有关;而６－ＢＡ 延缓衰老则可能与其减缓ＧＡｓ的糖苷化有关     

Leucocyanidin reductase activity and accumulation of proanthocyanidins in developing legume tissues

Proanthocyanidin (PA) and anthocyanin accumulation and location in developing leaves, flowers, and seeds of the legumes Medicago sativa, Lotus japonicus, Lotus uliginosus, Hedysarum sulfurescens, and Robinia pseudacacia were investigated by quantitative measurements and by histological analysis after staining with 1% vanillin/HCl, butanol/HCl, or 50% HCl. M. sativa leaves and flowers, L. japonicus leaves, and R. pseudacacia flowers do not contain PAs, but seeds of all investigated species contain PAs. Anthocyanins are absent in the seed coats of all five species and in leaves of L. japonicus. PA content generally increases as a function of development in leaves, but declines in flowers. With the exception of H. sulfurescens, flower PAs are synthesized in the parenchyma cells of the standard petal, while anthocyanins are located in the neighboring epidermal cells. Leucocyanidin reductase (LCR) catalyzes the conversion of 2,3-trans-3,4-cis-leucocyanidin to (+)-catechin and is the first enzyme in the PA-specific pathway. LCR activity was only detected in PA-containing tissues and generally declined during tissue development.     

Hatch date‐dependent differences in early growth and development recorded in the otolith microstructure of Trachurus japonicus

Otolith microstructure of 308 juvenile jack mackerel Trachurus japonicus sampled in the southern Sea of Japan between June and September 2002 were examined by light microscopy. Hatch‐date distribution indicated a protracted spawning season from 16 January to 30 May 2002. The first secondary primordium (SP) was formed at ages 19–54 days, with a mean ±  s . d . of 30·4 ± 6·1. The number of SP in an otolith ranged from two to 15, with a mean of 6·1 ± 1·7. Age of the first SP formation, number of SP and increment width varied with hatch date. In general, late‐hatched fish were younger when the first SP formed, and had a higher number of SP and wider increments, than early‐hatched fish. As increment width is a linear function of somatic growth and formation of the first SP is associated with metamorphosis in jack mackerel, these variations in otolith microstructure indicated that rates of growth (as inferred from wider increments) and development (as inferred by younger age of first SP formation) were higher in late‐hatched fish relative to early‐hatched fish. Higher temperature experienced by late‐hatched fish during early life stages may contribute critically to their higher growth and development rates compared to early‐hatched fish.     

浙麦冬块根与须根主要成分比较

为比较浙麦冬(Ophiopogon japonicus)块根和须根中的主要成分,采用GC-MS对2年生块根及须根中的有效成分和挥发油成分进行了分析。结果表明,浙麦冬块根的总糖含量高于须根,而须根的可溶性糖、黄酮含量皆略高于块根。从浙麦冬根的挥发油中共检出73种化合物,其中块根50种,须根48种,共同含有的物质有香桧烯、莰酮、愈创木醇等25种,含量较多的成分有β-榄香烯、长叶烯及绿花白千层醇等。低沸点的醇类及烯烃为麦冬根部挥发油的主要成分。浙麦冬须根的化学成分在种类和含量上皆与块根相似,有些成分甚至高于块根,可以作为块根的替代品应用于保健品及药物研发等领域。