Total and subcellular distribution of trace elements in the liver of a mother-fetus pair of Dall's porpoises (Phocoenoides dalli)

Total and subcellular hepatic Zn, Cu, Se, Mn, V, Hg, Cd, and Ag were determined in a mother-fetus pair of Dall's porpoises (Phocoenoides dalli). Except for higher fetal Cu concentration, all maternal elements were higher. Elements existed mostly in the cytosol of both animals except in the case of maternal Ag in the microsome and fetal Cu and Ag in the nuclei and mitochondria. In the maternal cytosol, Zn, Mn, Hg, and Ag were present in the high-molecular-weight substances (HMW); Se and V were present in the low-molecular-weight substances (LMW); Cu and Cd were mostly sequestered by metallothionein (MT). In the fetal cytosol, Zn, Se, Mn, Hg, Cd, and Ag were present in the HMW and V in the LMW, while Cu and Ag were mostly associated with MT. MT isoforms were characterized using the HPLC/ICP-MS. Two and four obvious peaks appeared in the maternal and fetal MT fractions, respectively. The highest elemental ion intensities were at a retention time of 7.8 min for the mother, and for the fetus the peak elemental ion intensities occurred at a retention time of 4.3 min, suggesting that different MT isoforms may be involved in elemental accumulation in maternal and fetal hepatocytosols. (C) 2003 Elsevier Ltd. All rights reserved.

Two unisexual artificial polyploid clones constructed by genome addition of common carp (Cyprinus carp) and crucian carp (Carassius auratus)

A polyploid hybrid fish with natural gynogenesis can prevent segregation and maintain their hybrid vigor in their progenies. Supposing the reproduction mode of induced polyploid fish being natural gynogenesis, allopolyploid hybrid between common carp and crucian carp into allopolyploid was performed. The purpose of this paper is to describe a lineage from sexual diploid carp transforming into allotriploid and allotetraploid unisexual clones by genome addition. The diploid hybrid between common carp and crucian carp reproduces an unreduced nucleus consisting of two parental genomes. This unreduced female pronucleus will fuse with male pronucleus and form allotriploid zygote after penetration of related species sperms. Allotriploid embryos grow normally, and part of female allotriploid can produce unreduced mature ova with three genomes. Mature ova of most allotriploid females are provided with natural gynogenetic trait and their nuclei do not fuse with any entrance sperm. All female offspring are produced by gynogenesis of allotriploid egg under activation of penetrating sperms. These offspring maintain morphological traits of their allotriploid maternal and form an allotetraploid unisexual clone by gynogenetic reproduction mode. However, female nuclei of rare allotriploid female can fuse with penetrating male pronuclei and result in the appearance of allotetraploid individuals by means of genome addition. All allotetraploid females can reproduce unreduced mature eggs containing four genomes. Therefore, mature eggs of allotetraploid maintain gynogenetic trait and allotetraploid unisexual clone is produced under activation of related species sperms.

Comparative studies on in vitro sperm decondensation and pronucleus formation in egg extracts between gynogenetic and bisexual fish

A cell-free system based upon the egg extracts from gynogenetic gibel carp (Carassius auratus gibelio) or bisexual red common carp (Cyprinus carpio red variety) was developed to investigate developmental behaviors of the demembranated sperm nuclei. Both red common carp and gibel carp sperm nuclei could decondense fully and form pronuclei in the red common carp egg extracts. Gibel carp sperm nuclei could also decondense fully and form pronuclei in the gibel carp egg extracts, but red common carp sperm nuclei could not decondense sufficiently in the same extracts. The significant differences of morphological changes were further confirmed by ultrastructural. observation of transmission electron microscopy. The data further offer cytological evidence for gonochoristic reproduction in the gynogenetically reproducing gibel carp. In addition, the sperm nuclei in vitro decondensation is dependent on the pH in the extracts, and the decondensed efficiency is optimal at pH 7. However, no DNA replication was observed in the two kinds of egg extracts during the incubation period of the sperm nuclei. It is suggested that the egg extracts prepared from the gynogenetic gibel carp should be a valid in vitro system for studying molecular mechanism on gynogenesis and reproduction mode diversity in fish.

Factors affecting the efficiency of somatic cell nuclear transplantation in the fish embryo

Procedures to improve somatic cell nuclear transplantation in fish were evaluated. We reported effects of nonirradiated recipient eggs, inactivated recipient eggs, different combinations between recipient eggs and donor cells, duration of serum starvation, generation number, and passage number of donor cells on developmental rates of nuclear transplant (NT) embryos. Exposure to 25,000 R of gamma-rays inactivated recipient eggs. Single nucleus of cultured, synchronized somatic cell from gynogenetic bighead carp (Aristichthys nobilis) was transplanted into nonirradiated or genetically inactivated unfertilized egg of gibel carp (Carassius auratus gibelio). There was no significant difference in developmental rate between nonirradiated and inactivated recipient eggs (27.27% vs. 25.71%, respectively). Chromosome count showed that 70.59% of NT embryos contained 48 chromosomes. It showed that most NT embryos came from donor nuclei of bighead carp, which was supported by microsatellite analysis of NT embryos. But 23.53% of NT embryos contained more than 48 chromosomes. It was presumed that those superfluous chromosomes came from nonirradiated recipient eggs. Besides, 5.88% of NT embryos were chimeras. Eggs of blunt-snout bream (Megalobrama amblycephala) and gibel carp were better recipient eggs than those of loach (Misgurnus anguillicaudatus) (25% and 18.03% vs. 8.43%). Among different duration of serum starvation, developmental rate of NT embryos from somatic nuclei of three-day serum starvation was the highest, reaching 25.71% compared to 14.14% (control), 20% (five-day), and 21.95% (seven-day). Cultured donor cells of less passage facilitated reprogramming of NT embryos than those of more passage. Recloning might improve the developmental rate of NT embryos from the differentiated donor nuclei. Developmental rate of fourth generation was the highest (54.83%) and the lowest for first generation (14.14%) compared to second generation (38.96%) and third generation (53.01%). (C) 2002 Wiley-Liss, Inc.

Nuclear transplantation in different strains of zebrafish

Single later blastula nuclei from AB strain of zebrafish (Danio rerio) were transplanted into enucleated unfertilized eggs of Long fin strain. Of 1119 cloning embryos, 14 reconstructed embryos developed into fry. DNA fingerprinting systems of the cloned fish were similar to those of the nuclear donor fish, but were distinctly different from those of the unclear recipient fish. It confirmed that the genetic material originated from nuclear donor cell other than from nuclear recipient egg. The research suggested that the basic technique for nuclear transplantation performed with different strains of zebrafish has made a breakthrough. It should be helpful for the study of some important developmental problems such as gene function, the regulation of gene expression during animal development, the developmental potential of a nucleus and the interactions between the donor nucleus and the recipient cytoplasm, etc.

Long-term toxic impact of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the reproduction, sexual differentiation, and development of different life stages of Gobiocypris rarus and Daphnia magna

The sexual ratio of Gobiocypris rarus exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and 17 beta -estradiol from embryo to sexually mature revealed feminization and overdevelopment of connective tissue in male fish gonad in 2-30 pg/L TCDD concentration range. Daphnia magna was not sensitive to the high dose of TCDD (0.1-1000 ng/ml), but the reproduction of D. magna treated with TCDD decreased after the 8th day. 7-Ethoxyresorufin-O-deethylase (EROD) activities in newly fertilized eggs of G. rarus exposed to TCDD dosage groups (1000-100,000 pg/L) were significantly induced and increased with TCDD concentrations at the early life stage, while no difference was found between low TCDD dosage groups (<100 pg/L), but a good relationship between the EROD activity and the TCDD concentration was observed during a long-term developmental stage. There was a pericardial edema formed in a 2-week yolk-sac at the concentration of 1000 pg/L TCDD. But in the exposure group (2 pg/L TCDD for 120 days), the cell nuclei of hepatocytes was far from the center and packed toward the cell membrane; the cristae of most mitochondria in the cell dropped and collapsed; the rough endoplasmic reticulum broke into fragments; and numerous lipid droplets formed in the cell. (C) 2001 Academic Press.

The onset of foreign gene transcription in nuclear-transferred embryos of fish

The transcriptional onset of hGH-transgene in fish was studied in the following three cases: the first is in MThGH-transgenic F-4 common carp (Cyprinus carpio) embryos, the second is in nuclear-transferred embryos supported by the transgenic F-4 embryonic nuclei, and the third is in nuclear-transferred embryos supported by the transgenic F-4 tail-fin nuclei. RT-PCR results show that the hGH-transgene initiates its transcriptional activity from early-gastrula stage, the early blastula stage and even 16-cell stage in the first, second and third cases, respectively. it looks like that fish egg cytoplasm could just offer a very restricted reprogramming on transcriptional activity of specific gene in differentiated cell nuclei by nuclear transplantation.

Nuclear transplantation with early-embryonic cells of transgenic fish

Like other transgenic animals, transgenic fishes produced by microinjection are transgenic mosaics. In order to produce homogenous transgenic fish, the transgenic blastula or gastrula cells were dissociated from Carassius auratus, Pengze var, and Cyprinus carpio, Huanghe var., and the nuclei were transferred into the mature eggs of the same species via microinjection or electro-fusion. Five nuclear-transferred Carassius auratus, Pengze var. and one Cyprinus carpio, Huanghe var. were obtained and the existence of the transgene was detected. The possibility of generating homogenous strain of transgenic fish by nuclear transplantation with transgenic early-embryonic cells is discussed.

Embryonic and genetic manipulation in fish

Fishes, the biggest and most diverse community in vertebrates are good experimental models for studies of cell and developmental biology by many favorable characteristics. Nuclear transplantation in fish has been thoroughly studied in China since 1960s. Fish nuclei of embryonic cells from different genera were transplanted into enucleated eggs generating nucleo-cytoplasmic hybrids of adults. Most importantly, nuclei of cultured goldfish kidney cells had been reprogrammed in enucleated eggs to support embryogenesis and ontogenesis of a fertile fish. This was the first case of cloned fish with somatic cells. Based on the technique of microinjection, recombinant MThGH gene has been transferred into fish eggs and the first batch of transgenic fish were produced in 1984. The behavior of foreign gene was characterized and the onset of the foreign gene replication occurred between the blastula to gastrula stages and random integration mainly occurred at later stages of embryogenesis. This eventually led to the transgenic mosaicism. The MThGH-transferred common carp enhanced growth rate by 2-4 times in the founder juveniles and doubled the body weight in the adults. The transgenic common carp were more efficient in utilizing dietary protein than the controls. An "all-fish" gene construct CAgcGH has been made by splicing the common carp beta-actin gene (CA) promoter onto the grass carp growth hormone gene (gcGH) coding sequence. The CAgcGH-transferred Yellow River Carp have also shown significantly fast-growth trait. Combination of techniques of fish cell culture, gene transformation with cultured cells and nuclear transplantation should be able to generate homogeneous strain of valuable transgenic fish to fulfil human requirement in 21(st) century.

Kinetically controlled InN nucleation on GaN templates by metalorganic chemical vapour deposition

This paper presents a study on the nucleation and initial growth kinetics of InN on GaN, especially their dependence on metalorganic chemical vapour deposition conditions. It is found that the density and size of separated InN nano-scale islands can be adjusted and well controlled by changing the V/III ratio and growth temperature. InN nuclei density increases for several orders of magnitude with decreasing growth temperature between 525 and 375 degrees C. At lower growth temperatures, InN thin films take the form of small and closely packed islands with diameters less than 100 nm, whereas at elevated temperatures the InN islands grow larger and become well separated, approaching an equilibrium hexagonal shape due to enhanced surface diffusion of adatoms. The temperature dependence of InN island density gives two activation energies of InN nucleation behaviour, which is attributed to two different kinetic processes related to In adatom surface diffusion and desorption, respectively.

The influence of growth temperature and input V/III ratio on the initial nucleation and material properties of InN on GaN by MOCVD

The effects of growth temperature and V/III ratio on the InN initial nucleation of islands on the GaN (0 0 0 1) surface were investigated. It is found that InN nuclei density increases with decreasing growth temperature between 375 and 525 degrees C. At lower growth temperatures, InN thin films take the form of small and closely packed islands with diameters of less than 100 nm, whereas at elevated temperatures the InN islands can grow larger and well separated, approaching an equilibrium hexagonal shape due to enhanced surface diffusion of adatoms. At a given growth temperature of 500 degrees C, a controllable density and size of separated InN islands can be achieved by adjusting the V/III ratio. The larger islands lead to fewer defects when they are coalesced. Comparatively, the electrical properties of the films grown under higher V/III ratio are improved.

Spin Relaxation of Electrons in Single InAs Quantum Dots

By using polarization-resolved photoluminescence spectra, we study the electron spin relaxation in single InAs quantum dots (QDs) with the configuration of positively charged excitons X+ (one electron, two holes). The spin relaxation rate of the hot electrons increases with the increasing energy of exciting photons. For electrons localized in QDs the spin relaxation is induced by hyperfine interaction with the nuclei. A rapid decrease of polarization degree with increasing temperature suggests that the spin relaxation mechanisms are mainly changed from the hyperfine interaction with nuclei into an electron-hole exchange interaction.

Surface morphology of AlN buffer layer and its effect on GaN growth by metalorganic chemical vapor deposition

The in situ optical reflectivity measurements are employed to monitor the GaN epilayer growth process above low-temperature AlN buffer layer on c-plane sapphire substrate by metalorganic chemical vapor deposition. It is found that the lateral growth of GaN islands and their coalescence is promoted in the initial growth stage if the AlN buffer layer is treated with a long annealing time and has an optimal thickness: As confirmed by atomic force microscopy observations, the quality of GaN epilayers is closely dependent on the surface morphology of AlN buffer layer, especially the grain size and nuclei density after the annealing treatment. (C) 2004 American Institute of Physics.

Evolution of mosaic structure in InN grown by metalorganic chemical vapor deposition

Mosaic structure in InN layers grown by metalorganic chemical vapor deposition at various temperatures has been investigated by X-ray diffraction (XRD). With a combination of Williamson-Hall measurement and fitting of twist angles, it was found that variation of growth temperature from 450 to 550 degrees C leads to the variation of the lateral coherence length, vertical coherence length, tilt and twist of mosaic blocks in InN films in a, respectively, monotonic way. In particular, mosaic tilt increases whereas mosaic twist decreases with elevating temperature. Atomic force microscopy shows the morphological difference of the InN nucleation layers grown at 450 and 550 degrees C. Different coalescence thickness and temperature-dependent in-plane rotation of InN nuclei are considered to account for the XRD results. (c) 2006 Elsevier B.V. All rights reserved.

Influence of growth pressure of a GaN buffer layer on the properties of MOCVD GaN

The influence of growth pressure of GaN buffer layer on the properties of MOCVD GaN on alpha-Al2O3 has been investigated with the aid of a home-made in situ laser reflectometry measurement system. The results obtained with in situ measurements and scanning electron microscope show that with the increase in deposition pressure of buffer layer, the nuclei increase in size, which roughens the surface, and delays the coalescence of GaN nuclei. The optical and crystalline quality of GaN epilayer was improved when buffer layer was deposited at high pressure.