As a part of our effort to obtain a strong regulatory element for the construction of an autogenic mud loach transgenic vector, we isolated a genomic clone that contains an open reading frame encoding the beta-actin gene, then examined the transcriptional activity of the upstream sequences, including the first intron, in transiently transfected cell lines. It showed that the upstream region has substantially strong transcriptional activity, and that both the proximal promoter and distal region of intron 1 play a crucial role in the activity. A similar result, based on fish growth, was obtained with the expression vectors containing the growth hormone gene of mud loach. These were driven by the regulatory region of the mud loach beta-actin gene with various mutations, and were directly transferred into the trunk muscle of fish using an electrostimulation-mediated method. Fish weights were monitored over the next 4 weeks. These data suggest that the proximal promoter and the first intron enhancer of the mud loach beta-actin gene are useful for autogenic mud loach transgenesis.
The shortnose sturgeon Acipenser brevirostrum was revealed to have a larger number of chromosomes than previously reported for other sturgeon species. Its chromosome number ranged from 362 to 372 (of ten specimens examined), showing intraindividual variation. The karyotype of metaphase with the highest chromosome number (372) consisted of 89 pairs of macrochromosomes and 97 pairs of microchromosomes (fundamental number; NF ϭ 550). Although the microchromosomes were relatively shorter than the macrochromosomes, most of them had discernible arms and centromeres. Silver-stained nucleolar organizer regions (Ag-NORs) were localized on the telomeric regions of 5 pairs of chromosomes (Ag-NORs ϭ 10): 4 were made up of small meta/submetacentrics and 1 of acrocentrics. Polyploidy of A. brevirostrum should be hexaploid based on the karyotype, numerous chromosomes, Ag-NORs, and previously reported large genome size (ca. 13 pg DNA/cell).Supplementary material A color version of silver-stained metaphase of Acipenser brevirostrum (Fig. S1) is available on Springer's server at springerlink.com Materials and MethodsSpecimens were captive-bred and raised in the hatchery at University of Florida, Gainesville, USA. They (half-siblings) were given an intraperitoneal injection of 0.5% colchicin at a dose level of 10 µg/g body weight and then kept for 3-4 h in well-aerated water at 25°C. The kidneys were surgically removed, scissored, and subjected to hypotonic treatment (0.075 M KCl) for 20 min at room temperature. The cell suspension was fixed with cold methanol-glacial acetic acid (3 : 1). Chromosome slides were prepared using an airdrying method (Kim et al., 1995). The chromosomes were stained with Giemsa (Fluka) for karyotypic analysis. Countable metaphases were examined from ten speciemens.For the selective staining of Ag-NORs, Giemsa-stained metaphases were destained with alcohol-acetic acid mixture for 20 min followed by absolute ethanol for 1 h. NOR staining with AgNO 3 was carried out following the colloidal silver method of Howell and Black (1980). The best 6-10 metaphases exhibiting high NOR values per fish were examined for determining the number of active Ag-NORs. ResultsChromosome number and karyotype. The chromosome counts of Acipenser brevirostrum showed intraindividual IchthyologicalResearch
The population structure of olive flounder Paralichthys olivaceus was estimated using nine polymorphic microsatellite (MS) loci in 459 individuals collected from eight populations, including five wild and three hatchery populations in Korea. Genetic variation in hatchery (mean number of alleles per locus, A = 10.2-12.1; allelic richness, A(R) = 9.3-10.1; observed heterozygosity, H(O) = 0.766-0.805) and wild (mean number of alleles per locus, A = 11.8-19.6; allelic richness, A(R) = 10.9-16.1; observed heterozygosity, H(O) = 0.820-0.888) samples did not differ significantly, suggesting a sufficient level of genetic variation in these well-managed hatchery populations, which have not lost a substantial amount of genetic diversity. Neighbour-joining tree and principal component analyses showed that genetic separation between eastern and pooled western and southern wild populations in Korea was probably influenced by restricted gene flow between regional populations due to the barrier effects of sea currents. The pooled western and southern populations are genetically close, perhaps because larval dispersal may depend on warm currents. One wild population (sample from Wando) was genetically divergent from the main distribution, but it was genetically close to hatchery populations, indicating that the genetic composition of the studied populations may be affected by hydrographic conditions and the release of fish stocks. The estimated genetic population structure and potential applications of MS markers may aid in the proper management of P. olivaceus populations.
This article documents the addition of 205 microsatellite marker loci to the Molecular Ecology Resources Database. Loci were developed for the following species: Bagassa guianensis, Bulweria bulwerii, Camelus bactrianus, Chaenogobius annularis, Creontiades dilutus, Diachasmimorpha tryoni, Dioscorea alata, Euhrychiopsis lecontei, Gmelina arborea, Haliotis discus hannai, Hirtella physophora, Melanaphis sacchari, Munida isos, Thaumastocoris peregrinus and Tuberolachnus salignus. These loci were cross-tested on the following species: Halobaena caerulea, Procellaria aequinoctialis, Oceanodroma monteiroi, Camelus ferus, Creontiades pacificus, Dioscorea rotundata, Dioscorea praehensilis, Dioscorea abyssinica, Dioscorea nummularia, Dioscorea transversa, Dioscorea esculenta, Dioscorea pentaphylla, Dioscorea trifida, Hirtella bicornis, Hirtella glandulosa, Licania alba, Licania canescens, Licania membranaceae, Couepia guianensis and 7 undescribed Thaumastocoris species.
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