Confirmation of Novel Quantitative Trait Loci for Seed Dormancy at Different Ripening Stages in Rice

Seed dormancy contributes resistance to pre-harvest sprouting.Effects on respective quantitative trait loci (QTLs) for dormancy should be assessed by using fresh seeds before germinability altered through storage.We investigated QTLs related to seed dormancy using backcross inbred lines derived from a cross between Nipponbare and Kasalath.Four putative QTLs for seed dormancy were detected immediately after harvest using composite interval mapping.These putative QTLs were mapped near C1488 on chromosome 3 (qSD-3.1),R2171 on chromosome 6 (qSD-6.1),R1245 on chromosome 7 (qSD-7.1) and C488 on chromosome 10 (qSD-10.1).Kasalath alleles promoted dormancy for qSD-3.1,qSD-6.1 and qSD-7.1,and the respective proportions of phenotypic variation explained by each QTL were 12.9%,9.3% and 8.1%.We evaluated the seed dormancy harvested at different ripening stages during seed development using chromosome segment substitution lines (CSSLs) to confirm gene effects.The germination rates of CSSL27 and CSSL28 substituted with the region including qSD-6.1 were significantly lower than those of Nipponbare and other CSSLs at the late ripening stage.Therefore,qSD-6.1 is considered the most effective novel QTL for pre-harvest sprouting resistance among the QTLs detected in this study.     

Bovine hepatocyte growth factor and its receptor c-Met: cDNA cloning and expression analysis in the mammary gland

Hepatocyte growth factor/scatter factor (HGF/SF) is a pleiotropic cytokine that plays a crucial role in the embryonic and postnatal development of various organs including the mammary gland. We cloned bovine HGF and its c-Met receptor cDNAs, and examined their expression during mammary gland development in dairy cows. The 2.5-kbp HGF cDNA clone contained a 2190 bp open reading frame coding a 730 amino acid protein, while the 4.8-kbp c-Met cDNA clone contained a 4152 bp open reading frame coding a 1384 amino acid protein. The bovine HGF and c-Met sequences exhibited more than 87% identity with those of other mammals. RT-PCR analysis revealed ubiquitous expression of both HGF and c-Met mRNAs in various bovine tissues tested. HGF mRNA was detected only in the inactive stage of bovine mammary gland development and not in the developing, lactating, and involuting stages, while c-Met mRNA was detected in the inactive and involuting stages. Immunohistochemical analysis demonstrated that the c-Met protein was found on mammary epithelial cells in the inactive, developing, and involuting stages, and on myoepithelial cells in all stages. These results suggest pivotal roles of HGF and c-Met in the development of bovine mammary gland.     

The complete mitochondrial genome of the domestic red deer (Cervus elaphus) of New Zealand and its phylogenic position within the family Cervidae

We determined the complete nucleotide sequence of the mitochondrial genome of the semidomestic red deer (Cervus elaphus) of New Zealand. The genome was 16 357 bp long and contained 13 protein‐coding genes, 12SrRNA, 16SrRNA, 22 tRNAs and a D‐loop as found in other mammals. Database homology searches showed that the mitochondrial DNA (mtDNA) sequence from the New Zealand semidomestic deer was similar to partial mtDNA sequences from the European, Norwegian (C. e. atlanticus) and Spanish red deer (C. e. hispanicus). Phylogenetic analysis of the mitochondrial protein‐coding regions revealed two well‐defined monophyletic clades in subfamilies Cervinae and Muntiacinae. However, red deer and Sika deer were not found to be close relatives. The analysis did identify the red deer as a sister taxon of a Samber/Sika deer clade, although it was more closely related to the Samber than the Sika group.     

Expression and subcellular localization of GSE protein in germ cells and preimplantation embryos

We previously identified a novel gonad-specific expression gene (Gse) and investigated its expression during gametogenesis in the mouse testis and ovary. In this study, we generated a polyclonal antibody to GSE protein and determined the profiles of the protein's expression in germ cells and preimplantation embryos in detail using immunocytochemical and immunofluorescence staining. In a Western blot analysis, the anti-GSE antibody recognized long and short isoforms (approximately 27.6 kDa and 23.1 kDa) of the protein in the mouse testis and the long isoform in the ovary. In the mouse testis, GSE protein was expressed in spermatocytes I in the pachytene stage, round spermatids, and elongated spermatids. In the mouse ovary, the protein was located in the cytoplasm and nucleus of all oocytes regardless of the stage of the ovarian follicles. In preimplantation embryos from the pronuclear to blastocyst stage, however, GSE protein was mainly detected in the nuclei of cells. At the blastocyst stage, the protein was confirmed to have accumulated in the inner cell mass (ICM), whereas it had mostly disappeared from the trophectoderm (TE). These findings suggest that GSE protein may play a role in the establishment of nuclear totipotency and may be associated with early lineage specification.     

Fine surface structure of bovine acrosome-intact and reacted spermatozoa observed by atomic force microscopy

The atomic force microscope (AFM) provides nanometer resolution, topographic data of the natural surface structure of materials. We studied the topology of the surface structure of bovine sperm heads during the acrosome reaction by AFM. In addition, we numerically analyzed the areas of the median sagittal plane of the sperm heads. Bovine frozen-thawed spermatozoa were washed, capacitated by heparin, and incubated with lysophosphatidylcholine (LPC) to induce the acrosome reaction, smeared on a cover glass, air-dried, and observed with AFM using the dynamic force (tapping) mode. AFM analysis of spermatozoa showed the clear surface structure of acrosomes, equatorial segments, postacrosomal regions and necks. Although AFM images of spermatozoa capacitated by heparin had complete acrosomes, most spermatozoa treated with LPC had no acrosomal caps as shown by AFM. These observations coincided with those obtained by light microscopy after staining with naphthol yellow S and erythrosin B. Furthermore, numerical analysis of AFM images indicated that areas of the median sagittal plane of the anterior portions of acrosome-reacted sperm heads (2679 +/- 616 pixels) were approximately 40% less than those of intact heads (4535 +/- 174 pixels, P<0.05). These results indicate that AFM can usefully observe and numerically analyze the fine surface structures of bovine spermatozoa.     

Factors maintaining species diversity in satoyama, a traditional agricultural landscape of Japan

Satoyama is a traditional agricultural landscape in Japan. It contains many kinds of organisms, including endangered species. Satoyama is composed of several habitat types, including paddy fields, secondary forests, secondary grasslands, ponds, and streams. High species richness has been sustained in most habitats by anthropogenic disturbances of intermediate impact, mainly activities related to agriculture. The variety of habitats and connectivity among them have contributed to the high species diversity within satoyama. These factors allow organisms to move among habitats and use different habitat types to obtain different resources. The connectivity among habitats is often more vulnerable than the specific habitats themselves under anthropogenic influences. In satoyama, species that require connectivity among habitats (e.g., grey-faced buzzard [Butastur indicus] and Genji firefly [Luciola cruciata]) have tended to decrease. The grey-faced buzzard is categorized as a “vulnerable” species in the Japanese Red List. It usually nests in forest trees and forages in habitats such as paddy fields, grasslands, and forests. Its foraging locations shift seasonally depending on food availability. It is thought that the degradation of either paddy fields or forests in a landscape may result in the disappearance of this species. The Genji firefly spends its larval period in stream water, pupates underground along streams, and flies near streams after emergence. This species requires not only the integrity of streams and nearby areas, but also connectivity between these areas. The ecology of these species suggests that the variety of habitats and connectivity among them are critical factors for their survival.