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Seed coat cracking in soybeans [Glycine max (L). Merr.] leads to commercial and agronomic losses. The Japanese elite soybean cultivar ‘Fukuyutaka’ is often used as a parent for breeding, but its high rate of seed coat cracking is an obstacle to its further use in breeding programs. To establish a DNA marker-assisted selection system for seed coat cracking, genetic factors related to high rates of seed coat cracking were surveyed, and a quantitative trait locus (QTL) with a stable effect on seed coat cracking in both years of a two-year replication experiment was detected on chromosome 20. Comparison of a set of near-isogenic lines (NILs) around this locus verified that the presence of the ‘Fukuyutaka’ allele significantly increased seed coat cracking in the kernel. The locus is located in a genomic region spanning 3.2 Mb. Marker-assisted selection for the locus will improve the selection efficiency of ‘Fukuyutaka’-derived breeding populations.

Soyasaponins are specialized metabolites present in soybean seeds that affect the taste and quality of soy-based foods. The composition of the sugar chains attached to the aglycone moiety of soyasaponins is regulated by genetic loci such as sg-1, sg-3 and sg-4. Here, we report the cloning and characterization of the Sg-3 gene, which is responsible for conjugating the terminal (third) glucose (Glc) at the C-3 sugar chain of soyasaponins. The gene Glyma.10G104700 is disabled in the sg-3 cultivar, 'Mikuriya-ao', due to the deletion of genomic DNA that results in the absence of a terminal Glc residue on the C-3 sugar chain. Sg-3 encodes a putative glycosyltransferase (UGT91H9), and its predicted protein sequence has a high homology with that of the product of GmSGT3 (Glyma.08G181000; UGT91H4), which conjugates rhamnose (Rha) to the third position of the C-3 sugar chain in vitro. A recombinant Glyma.10G104700 protein could utilize UDP-Glc as a substrate to conjugate the third Glc to the C-3 sugar chain, and introducing a functional Glyma.10G104700 transgene into the mutant complemented the sg-3 phenotype. Conversely, induction of a premature stop codon mutation in Glyma.10G104700 (W270*) resulted in the sg-3 phenotype, suggesting that Glyma.10G104700 was Sg-3. The gmsgt3 (R339H) mutant failed to accumulate soyasaponins with the third Rha at the C-3 sugar chain, and the third Glc and Rha conjugations were both disabled in the sg-3 gmsgt3 double mutant. These results demonstrated that Sg-3 and GmSGT3 are non-redundantly involved in conjugation of the third Glc and Rha at the C-3 sugar chain of soyasaponins, respectively.

Soyasaponins can be classified into two groups as A saponins and DDMP saponins. Fully-acetylated group A saponins, located on the seed hypocotyls, are the major cause of undesirable characteristics in soy foods. Apart from the fully-acetylated and deacetylated group A saponins, other saponin components have not been studied. This research aimed to identify 105 saponins in the seed hypocotyls and cotyledons of nine saponin phenotype soybean varieties. Soyasapogenol A glucosides (SAGs) including fully-acetylated SAGs (FSAGs), partially-acetylated SAGs (PSAGs), deacetylated SAGs (DSAGs), and DDMPs were identified by LC-PDA-MS/MS and quantified by UV-chromatograms. The complexity of soyasaponin compositions depended on the seed structure (hypocotyls and cotyledons) and variety. Other than in seed hypocotyls, FSAGs were detected in seed cotyledons ranging from 2.1 to 34.9 μmol/100 g dry basis. Minor components as PSAGs were also detected including monoacetyl, diacetyl, and triacetyl ranging between 2.2–19.8 and 1.6–44.0 μmol/100 g dry basis for hypocotyls and cotyledons, respectively. The degree of acetylation affects the intensity of undesirable characteristics in soy products and this issue requires further study. All saponin compositions require clarification using tandem mass spectrometry.

Calibration of the molecular rate is one of the major challenges in marine population genetics. Although the use of an appropriate evolutionary rate is crucial in exploring population histories, calibration of the rate is always difficult because fossil records and geological events are rarely applicable for rate calibration. The acceleration of the evolutionary rate for recent coalescent events (or more simply, the time dependency of the molecular clock) is also a problem that can lead to overestimation of population parameters. Calibration of demographic transition (CDT) is a rate calibration technique that assumes a post-glacial demographic expansion, representing one of the most promising approaches for dealing with these potential problems in the rate calibration. Here, we demonstrate the importance of using an appropriate evolutionary rate, and the power of CDT, by using populations of the sandy beach amphipod Haustorioides japonicus along the Japanese coast of the northwestern Pacific Ocean. Analysis of mitochondrial sequences found that the most peripheral population in the Pacific coast of northeastern Honshu Island (Tohoku region) is genetically distinct from the other northwestern Pacific populations. By using the two-epoch demographic model and rate of temperature change, the evolutionary rate was modeled as a log-normal distribution with a median rate of 2.2%/My. The split-time of the Tohoku population was subsequently estimated to be during the previous interglacial period by using the rate distribution, which enables us to infer potential causes of the divergence between local populations along the continuous Pacific coast of Japan.

Small cryptic animals utilize the interstices of coral rubble as a refuge from predators and form a significant part of the diversity and productivity of coral reef ecosystems. In the present study, we examined the composition of cryptic mobile animals along an estuarine gradient within a mangrove-seagrass-coral reef seascape off Ishigaki Island, southwest Japan. We used mesh traps with defaunated coral rubble to sample cryptic mobile animals quantitatively at four sites: upper-estuary mangrove, lower-estuary mangrove, seagrass bed, and coral lagoon. Sixty-five species of decapod crustaceans, gastropods, fishes, and other mobile animals were identified. The assemblage composition of these identified species was significantly different among the four sites. Low-resolution taxa including copepods, gammarids, and other taxa also showed significant differences in composition among the four sites. Two types of rubble (branch and lump) were used to examine the possible effects of microhabitat structure, but no significant effect on assemblage composition was detected. Results indicate that, despite the close proximity and high degree of water mass mixing among the sites, cryptic animals colonized the trays from the surrounding habitats, and that the assemblage of cryptic animals varied considerably along the estuarine gradient.

In soybean, the I gene inhibits pigmentation over the entire seed coat, resulting in yellow seeds. It is thought that this suppression of seed coat pigmentation is due to naturally occurring RNA silencing of chalcone synthase genes (CHS silencing). Fully pigmented seeds can be found among harvested yellow seeds at a very low percentage. These seed coat pigmented (scp) mutants are generated from yellow soybeans by spontaneous recessive mutation of the I gene. A candidate for the I gene, GmIRCHS, contains a perfect inverted repeat (IR) of a CHS pseudogene (pseudoCHS3) and transcripts of GmIRCHS form a double-stranded CHS RNA that potentially triggers CHS silencing. One CHS gene, ICHS1, is located 680 bp downstream of GmIRCHS. Here, the GmIRCHS-ICHS1 cluster was compared in scp mutants of various origins. In these mutants, sequence divergence in the cluster resulted in complete or partial loss of GmIRCHS in at least the pseudoCHS3 region. This result is consistent with the notion that the IR of pseudoCHS3 is sufficient to induce CHS silencing, and further supports that GmIRCHS is the I gene.

Determination of the growth pattern of a harvested species is an important step toward a proper assessment and suitable management of the stock. Although coconut crab Birgus latro populations in most regions have been severely depleted, few reliable biological data are available. We assessed the growth of B. latro using a mark-recapture method based on the use of passive integrated transponder (PIT) tags on Hatoma Island, located southwest of Okinawa, Japan. Of 781 crabs tagged (337 males, 444 females; 14.89 to 58.03 mm thoracic length (ThL)), 17 males and 20 females were recaptured. The application of PIT tags provided field data for growth of individuals for up to 2326 d and confirmed that coconut crabs molt during the winter dry season on Hatoma Island. The curves relating ThL with age, estimated from data of 29 crabs that had spent one or multiple winter dry seasons at liberty, showed that the coconut crab is an extremely slow growing species, with males showing much higher mean asymptotic ThL (69.87 mm) than females (42.79 mm). The Brody growth coefficient (K)was 0.061 yr �1 for males and 0.091 yr �1 for females. This study is the first to provide growth estimates of wild coconut crabs based on mark- recapture over multiple years. Potentially long-lived, slow growing species such as coconut crabs may be susceptible to overharvesting, and a cautious approach is recommended to the determina- tion of resource management regimes for B. latro.

The objective of this study was to identify quantitative trait loci (QTL) affecting fitness of hybrids between wild soybean (Glycine soja) and cultivated soybean (Glycine max). Seed dormancy and seed number, both of which are important for fitness, were evaluated by testing artificial hybrids of G. soja × G. max in a multiple-site field trial. Generally, the fitness of the F1 hybrids and hybrid derivatives from self-pollination was lower than that of G. soja due to loss of seed dormancy, whereas the fitness of hybrid derivatives with higher proportions of G. soja genetic background was comparable with that of G. soja. These differences were genetically dissected into QTL for each population. Three QTLs for seed dormancy and one QTL for total seed number were detected in the F2 progenies of two diverse cross combinations. At those four QTLs, the G. max alleles reduced seed number and severely reduced seed survival during the winter, suggesting that major genes acquired during soybean adaptation to cultivation have a selective disadvantage in natural habitats. In progenies with a higher proportion of G. soja genetic background, the genetic effects of the G. max alleles were not expressed as phenotypes because the G. soja alleles were dominant over the G. max alleles. Considering the highly inbreeding nature of these species, most hybrid derivatives would disappear quickly in early self-pollinating generations in natural habitats because of the low fitness of plants carrying G. max alleles.

Triterpenoid saponins are major components of secondary metabolites in soybean seeds and are divided into two groups: group A saponins, and 2,3-dihydro-2,5-dihydroxy-6-methyl-4H-pyran-4-one (DDMP) saponins. The aglycone moiety of group A saponins consists of soyasapogenol A (SA), which is an oxidized β-amyrin product, and the aglycone moiety of the DDMP saponins consists of soyasapogenol B (SB). Group A saponins produce a bitter and astringent aftertaste in soy products, whereas DDMP saponins have known health benefits for humans. We completed map-based cloning and characterization of the gene Sg-5, which is responsible for SA biosynthesis. The naturally occurring sg-5 mutant lacks group A saponins and has a loss-of-function mutation (L164*) in Glyma15g39090, which encodes the cytochrome P450 enzyme, CYP72A69. An enzyme assay indicated the hydroxylase activity of recombinant CYP72A69 against SB, which also suggested the production of SA. Additionally, induced Glyma15g39090 mutants (R44* or S348P) lacked group A saponins similar to the sg-5 mutant, indicating that Glyma15g39090 corresponds to Sg-5. Endogenous levels of DDMP saponins were higher in the sg-5 mutant than in the wild-type lines due to the loss of the enzyme activity that converts SB to SA. Interestingly, the genomes of palaeopolyploid soybean and the closely related common bean carry multiple Sg-5 paralogs in a genomic region syntenic to the soybean Sg-5 region. However, SA did not accumulate in common bean samples, suggesting that Sg-5 activity evolved after gene duplication event(s). Our results demonstrate that metabolic switching of undesirable saponins with beneficial saponins can be achieved in soybean by disabling Sg-5.

The genus Macridiscus Dall, 1902 contains a few species present in warm temperate to tropical faunas of the western Pacific. Macridiscus was widely accepted as a subgenus of Gomphina until a recent suggestion that it should be separated from Gomphina and elevated to an independent genus, based on morphology and molecular data. The taxonomy of the genus Macridiscus has in the past been based solely on shell characters and there has been no agreement about the number of valid species. In this study, we explore the taxonomy and phylogeny of Macridiscus species in order to resolve the systematics of the genus, based not only on shell characters but also on mitochondrial and nuclear DNA sequences. The morphological characters, the sequences of the cytochrome c oxidase subunit I of mitochondrial DNA and the first internal transcribed spacer region (ITS1) between the 18S and 5.8S ribosomal DNA were highly concordant and clearly suggested that three species should be recognized in the genus Macridiscus: M. multifarius new species, M. semicancellata (Koch, in Philippi

Saponins are sterols or triterpene glycosides that are widely distributed in plants. The biosynthesis of soybean saponins is thought to involve many kinds of glycosyltransferases, which is reflected in their structural diversity. Here, we performed linkage analyses of the Sg-3 and Sg-4 loci, which may control the sugar chain composition at the C-3 sugar moieties of the soybean saponin aglycones soyasapogenols A and B. The Sg-3 locus, which controls the production of group A saponin Af, was mapped to chromosome (Chr-) 10. The Sg-4 locus, which controls the production of DDMP saponin βa, was mapped to Chr-1. To elucidate the preference of sugar chain formation at the C-3 and C-22 positions, we analyzed the F(2) population derived from a cross between a mutant variety, Kinusayaka (sg-1(0)), for the sugar chain structure at C-22 position, and Mikuriya-ao (sg-3), with respect to the segregation of the composition of the group A saponins, and found that the formation of these sugar chains was independently regulated. Furthermore, a novel saponin, predicted to be A0-γg, 3-O-[β-d-galactopyranosyl (1→2)-β-d-glucuronopyranosyl]-22-O-α-l-arabinopyranosyl-soyasapogenol A, appeared in the hypocotyl of F(2) individuals with genotype sg-1(0)/sg-1(0)sg-3/sg-3.

The peanut stunt virus (PSV) causes yield losses in soybean and reduced seed quality due to seed mottling. The objectives of this study were to determine the phenotypic reactions of soybean germplasms to inoculation with two PSV isolates (PSV-K, PSV-T), the inheritance of PSV resistance in soybean cultivars, and the locus of the PSV resistance gene. We investigated the PSV resistance of 132 soybean cultivars to both PSV isolates; of these, 73 cultivars exhibited resistance to both PSV isolates. Three resistant cultivars (Harosoy, Tsurunotamago 1 and Hyuga) were crossed with the susceptible cultivar Enrei. The crosses were evaluated in the F(1), F(2) and F(2:3) generations for their reactions to inoculation with the two PSV isolates. In an allelism test, we crossed Harosoy and Tsurunotamago 1 with the resistant cultivar Hyuga. The results revealed that PSV resistance in these cultivars is controlled by a single dominant gene at the same locus. We have proposed Rpsv1, as the name of the resistance gene in Hyuga. We also constructed a linkage map using recombinant inbred lines between Hyuga × Enrei using 176 SSR markers. We mapped Rpsv1 near the Satt435 locus on soybean chromosome 7.

Among commonly applied molecular markers, simple sequence repeats (SSRs, or microsatellites) possess advantages such as a high level of polymorphism and codominant pattern of inheritance at individual loci. To facilitate systematic and rapid genetic mapping in soybean, we designed a genotyping panel comprised 304 SSR markers selected for allelic diversity and chromosomal location so as to provide wide coverage. Most primer pairs for the markers in the panel were redesigned to yield amplicons of 80–600 bp in multiplex polymerase chain reaction (PCR) and fluorescence-based sequencer analysis, and they were labelled with one of four different fluorescent dyes. Multiplex PCR with sets of six to eight primer pairs per reaction generated allelic data for 283 of the 304 SSR loci in three different mapping populations, with the loci mapping to the same positions as previously determined. Four SSRs on each chromosome were analysed for allelic diversity in 87 diverse soybean germplasms with four-plex PCR. These 80 loci showed an average allele number and polymorphic information content value of 14.8 and 0.78, respectively. The high level of polymorphism, ease of analysis, and high accuracy of the SSR genotyping panel should render it widely applicable to soybean genetics and breeding.

Benthos such as Amphipoda living in coastal sandy sediments are important food organisms for fish, but, factors controlling their distributions remain poorly understood. Here, we aim to solve this question by introducing our new approach encompassing ecology, geophysics and hydraulics / geotechnics. We performed sets of integrated observations / surveys and analyses concerning the in situ intertidal geoenvironments and the benthos distributions at three natural coasts. The results demonstrate that the waterfront suction governs the variety of the habitat environments, involving a range of the degree of saturation, stiffness and void state, and the manifestation of the peak and critical area of densities for the three dominant species are uniquely determined by the magnitudes of such suction which are found to be particular to species throughout the coasts. These findings will effectively contribute to the conservation and management of fishery resources at sandy coasts.

Saponins are a diverse group of secondary metabolites widely distributed in plants. Some saponins in soybean seeds have medicinal properties, but the terminal acetylated sugar at the C-22 position of group A saponins in seed hypocotyls causes a bitter and astringent taste. We used cultivated and wild soybean accessions classified into four different group A saponin phenotypes, including non-acetylated A0-αg and deacetyl-Af, as parents to obtain four F2 populations and a population of recombinant inbred lines to test their genetic relationship. The gene controlling the phenotype of the group A saponins in each line was mapped near the simple sequence repeat marker Satt336 on soybean chromosome 7 (linkage group M). An allelism test of the A0-αg and deacetyl-Af variants revealed no segregation of Aa and Ab phenotypes in the progeny, although the genes controlling the two phenotypes have been assigned to two different loci, Sg-1 and Sg-2. These results suggest that the four group A saponin phenotypes are controlled by multiple alleles at the single locus Sg-1.

A well-saturated molecular linkage map is a prerequisite for modern plant breeding. Several genetic maps have been developed for soybean with various types of molecular markers. Simple sequence repeats (SSRs) are single-locus markers with high allelic variation and are widely applicable to different genotypes. We have now mapped 1810 SSR or sequence-tagged site markers in one or more of three recombinant inbred populations of soybean (the US cultivar ‘Jack’ × the Japanese cultivar ‘Fukuyutaka’, the Chinese cultivar ‘Peking’ × the Japanese cultivar ‘Akita’, and the Japanese cultivar ‘Misuzudaizu’ × the Chinese breeding line ‘Moshidou Gong 503’) and have aligned these markers with the 20 consensus linkage groups (LGs). The total length of the integrated linkage map was 2442.9 cM, and the average number of molecular markers was 90.5 (range of 70–114) for the 20 LGs. We examined allelic diversity for 1238 of the SSR markers among 23 soybean cultivars or lines and a wild accession. The number of alleles per locus ranged from 2 to 7, with an average of 2.8. Our high-density linkage map should facilitate ongoing and future genomic research such as analysis of quantitative trait loci and positional cloning in addition to marker-assisted selection in soybean breeding.

Coral rubble is one of the characteristic bottom habitats in tropical and subtropical coral reefs. Many mobile organisms inhabit the interstices among coral rubble. Because coral rubble habi- tats are frequently subjected to physical disturbances, the process of assemblage recolonization is important for maintaining the diversity and productivity in coral reefs. The present study examined the colonization patterns of these mobile organisms by using experimental traps with defaunated coral rubble at 3 areas (Back reef, Seagrass bed, and Nearshore) in a fringing coral reef on Ishigaki Island, Japan. Sixteen mesh trays filled with coral rubble were set at each of the 3 areas; 4 trays were collected after 1, 2, 4, and 8 wk. The experiment was repeated in winter and spring. The number of species that occurred in the trays generally increased with the time period, but detailed examination was difficult because of significant interaction factors. Higher order interactions (season, area, and time period) were also significant for variations in assemblage composition. Temporal and spatial variations in abundances of 15 numerically dominant species (gastropods and decapods) were analyzed by a general linear model with an Akaike's information criterion model selection. Although the patterns of colonization were species specific, initial (

Sessile organisms on PVC panels suspended inside and outside coral reef at Ishigaki Island, Yaeyama Islands, Japan, was monitored for 20 days just after the wide-spread incident of coral bleaching in September, 1998. The settlement of a total of 16 species of algae and invertebrates was recorded. Generally, Enteromorpha sp. (Chlorophyceae) dominated the panels. The percentage area cover, number of species, biomass and algal growth were significantly greater in waters outside the reef.

The environment of the habitat is important in the organization of the intertidal community. In a boulder shore habitat, accumulation of boulder layers increases shade and the number of interstices. This study examined whether experimental shade increased the density of mobile animals and modified the community structure at 2 intertidal zones on a moderately sheltered boulder shore in Amakusa, Japan. Artificial shade reduced light intensity and surface temperature of boulders, and subsequently increased microalgal abundance. Mobile animals were sampled quantitatively at 2 zones (high and mid) for 4 treatments: single layer boulders without a roof as a control, double layer boulders without a roof, single layer boulders shaded by a roof for 6 wk and for 2 wk. Statistical analyses (MANOVA and ANOVA) showed strong effects of treatments on community diversity and the density of 15 abundant species in both tidal zones. Generally, the diversity was lower in the single layer boulders than the double layer boulders and increased with the experimental shade. Two carnivorous species (Hemigrapsus sanguineus and Japeuthria cingulata) and some herbivorous species (e.g. Nerita japonica and Nipponacmea nigrans) increased in density with the shade, while other herbivores (e.g. Monodonta labio and Littorina brevicula) were unaffected. In conclusion, a reduction of environmental stress produced by the shade increased community diversity on the boulder shore, but the responses of mobile animals were species specific. As these responses to a change in the environment occurred within a very short period (< 2 wk), migration may play an important role in the short-term variation of community structure on boulder shores.

Growth of Monodonta lablo (Linne) (Gastropoda: Trochidae) was investigated bimonthly for a period of 1 yr by quadrat sampling and mark-recapture on a moderately exposed boulder shore in Amakusa, South Japan. Linear regression was calculated for shell width at recapture on shell width at release. Growth in the high zone was slower than in the mid and low zones and decreased as shell width increased. Estimated growth curves fit well with the observed growth curves of individual snails. Seasonal growth patterns of juvenile (58 mm) snails may be affected by seasonal fluctuation of micro- algal abundance. Due to reproduction, growth of large snails decreased in summer. Tidal level varia- tion of growth rate caused large variation in growth curves in middle-sized snails (ca 10 rnm shell width), because of their wide range of distribution, which was related to vertical migration on the shore. Simulated growth curves showed that 8 mm snails in the high zone reached asymptotic size (ca 13 mm) 1 yr later than snails in the mid and low zones. In small (ca 6 mm) and large (ca 13 mm) snails, less van- ability in observed growth curves was explained due to their limited ranges of distribution. This study demonstrates that variation in growth rate is attributable to sue-specific distribution on a shore and indicates how migration expands size variation within a single age group.

Triterpene saponins are a diverse group of biologically functional products in plants. Saponins usually are glycosylated, which gives rise to a wide diversity of structures and functions. In the group A saponins of soybean (Glycine max), differences in the terminal sugar species located on the C-22 sugar chain of an aglycone core, soyasapogenol A, were observed to be under genetic control. Further genetic analyses and mapping revealed that the structural diversity of glycosylation was determined by multiple alleles of a single locus, Sg-1, and led to identification of a UDP-sugar-dependent glycosyltransferase gene (Glyma07g38460). Although their sequences are highly similar and both glycosylate the nonacetylated saponin A0-αg, the Sg-1(a) allele encodes the xylosyltransferase UGT73F4, whereas Sg-1(b) encodes the glucosyltransferase UGT73F2. Homology models and site-directed mutagenesis analyses showed that Ser-138 in Sg-1(a) and Gly-138 in Sg-1(b) proteins are crucial residues for their respective sugar donor specificities. Transgenic complementation tests followed by recombinant enzyme assays in vitro demonstrated that sg-1(0) is a loss-of-function allele of Sg-1. Considering that the terminal sugar species in the group A saponins are responsible for the strong bitterness and astringent aftertastes of soybean seeds, our findings herein provide useful tools to improve commercial properties of soybean products.