Protein synthesis measured in the whole body of chick embryos cultured in vitro

1. Whole body protein synthesis was measured in chick embryos cultured in vitro. On day 7 of incubation chick embryos were cultured for 60 min in synthetic serum‐free medium containing 4‐[³H]phenylalanine. Specific radioactivities in free and protein‐bound phenylaline in the whole embryo were measured, starting 2 min after commencement of the culture process.

2. The values for fractional synthesis rate (FSR) estimated in vitro at 20, 30, 45 and 60 min during the embryo culture agreed well, ranging from 35 to 40%/d, suggesting that the method would serve as a useful model for studying the effect of growth promoters in chick embryos.

3. Bovine insulin in the synthetic medium did not affect FSR of protein in chick embryos cultured in vitro.     

Basic characterization of avian β‐defensin genes in the Japanese quail,Coturnix japonica

In this study, we identified a cluster of 14 avian β‐defensins (AvBD; approximately 66 kbp) in the Japanese quail, Coturnix japonica. Except for AvBD12 (CjAvBD12) and ‐13, the CjAvBDs coding sequences exhibited greater than 78.0% similarity to the respective orthologous chicken AvBD genes (GgAvBD). The putative amino acid sequence encoded by each CjAvBD contained six cysteine residues and the GXC (X1‐2) motif considered essential for the β‐defensin family. Each CjAvBDs also formed a sub‐group with the respective orthologous genes of various bird species in a phylogenetic tree analysis. Synteny between the CjAvBD cluster and GgAvBD cluster was confirmed. The CjAvBD cluster was mapped on the long‐arm end of chromosome 3 by linkage analysis based on single nucleotide polymorphisms (SNPs) of CjAvBD1 and CjAvBD12 (approximately 46kbp), as well as GgAvBD cluster. We also confirmed that CjAvBD1, ‐4, ‐5, ‐9, and ‐10 are transcribed in 20 tissues, including immune and digestive tissues. However, our experimental data indicated that the CjAvBD cluster lacks the AvBD3 and ‐7 loci, whereas the CjAvBD101α, ‐101β, and ‐101θ loci arose from gene duplication of the AvBD6 orthologous locus in the CjAvBD cluster after differentiation between Coturnix ‐ Gallus.     

Preventive effects of CIDR-based protocols on premature ovulation before timed-AI in Ovsynch in cycling beef cows

Ovsynch is a program developed to synchronize ovulation for timed breeding. In this paper, the authors investigate whether controlled internal drug release (CIDR)-based protocols prevent premature ovulation before timed-artificial insemination (AI) when Ovsynch is started a few days before luteolysis in cycling beef cows. Nine beef cows at 16 days after oestrus were treated with (1) Ovsynch, i.e. gonadotropin releasing hormone (GnRH) analogue on day 0, prostaglandin (PG) F(2alpha) analogue on day 7 and GnRH analogue on day 9 with timed-AI on day 10, (n=3); (2) Ovsynch+CIDR (Ovsynch protocol plus a CIDR for 7 days from day 0, n=3), or (3) oestradiol benzoate (OB)+CIDR+GnRH (OB on day 0 in lieu of the first GnRH treatment, followed by the Ovsynch+CIDR protocol, n=3). In the Ovsynch group (1) plasma progesterone concentrations fell below 0.5 ng/mL earlier (day 5) than in both CIDR-treated groups (2) and (3), where this occurred on day 8. Plasma oestradiol-17beta concentrations peaked on day 8 in the Ovsynch group and on day 9 in both CIDR-treated groups. The dominant follicle ovulated on day 10 in the Ovsynch group and on day 11 in both CIDR-treated groups. Thus, both CIDR-based protocols prevented premature ovulation before timed-AI in Ovsynch when the protocol was started a few days before luteolysis. This reflects the fact that progesterone levels remained high until the beef cattle were treated with PGF(2alpha).     

Effects of synchronization of donor cell cycle on embryonic development and DNA synthesis in porcine nuclear transfer embryos

The relationship between donor cell cycle and the developmental ability of somatic cell nuclear transfer (SCNT) embryos has not fully been elucidated. Donor cells that are usually prepared by serum starvation or confluent-cell culture for SCNT represent a heterogeneous population that includes mainly G0 phase cells, other cells in different phases of the cell cycle and apoptotic cells. In this study, we compared the developmental ability of porcine SCNT embryos reconstructed from G0 phase cells (G0-SCNT embryos) and strictly synchronized-G1 phase cells (G1-SCNT embryos), and examined the developmental rates and timing of first DNA synthesis. The G0 phase cells were synchronized by confluent culture, and the G1 phase cells were prepared from actively dividing M phase cells. The G1-SCNT embryos showed a significantly higher (P<0.05) developmental rate to the blastocyst stage per cleaved embryo (59%) than the G0-SCNT embryos (43%). Moreover, initiation of first DNA synthesis and cleavage occurred significantly earlier in the G1-SCNT embryos than in the G0-SCNT embryos. Delay of initiation of first DNA synthesis in the SCNT embryos by aphidicolin resulted in decreased developmental rates to the blastocyst stage without any effect on cleavage rates. Our data demonstrates that synchronized-G1 phase cells can be used as donor cells for SCNT embryos and that earlier initiation of first DNA synthesis may be important for subsequent development of SCNT embryos. The SCNT system using G1-synchronized cells, in terms of their highly uniform and viable cell states, can be useful for studying the reprogramming processes and embryonic development of SCNT embryos.     

Quantitative assessment of muscle mass and gene expression analysis in dogs with glucocorticoid-induced muscle atrophy

The present study aimed to quantitatively evaluate muscle mass and gene expression in dogs with glucocorticoid-induced muscle atrophy. Five healthy beagles received oral prednisolone for 4 weeks (1 mg/kg/day), and muscle mass was then evaluated via computed tomography. Histological and gene expression analyses were performed using biopsy samples from the biceps femoris before and after prednisolone administration. The cross-sectional area of the third lumbar paraspinal and mid-femoral muscles significantly decreased after glucocorticoid administration (from 27.5 ± 1.9 to 22.6 ± 2.0 cm² and from 55.1 ± 4.7 to 50.7 ± 4.1 cm², respectively; P<0.01). The fast- and slow-twitch muscle fibers were both atrophied (from 2,779 ± 369 to 1,581 ± 207 μm² and from 2,871 ± 211 to 1,971 ± 169 μm², respectively; P<0.05). The expression of the growth factor receptor-bound protein 10 (GRB10) significantly increased after prednisolone administration (P<0.05). Because GRB10 suppresses insulin signaling and the subsequent mammalian target of rapamycin complex 1 activity, increased expression of GRB10 may have resulted in a decrease in protein anabolism. Taken together, 1 mg/kg/day oral prednisolone for 4 weeks induced significant muscle atrophy in dogs, and GRB10 might participate in the pathology of glucocorticoid-induced muscle atrophy in canines.     

Genetic studies for breeding of rice cultivars with superior grain appearance and lodging resistance from the rice cultivar ‘Emi-no-kizuna’

ABSTRACT

We conducted a quantitative trait locus (QTL) analysis of grain appearance (GA) and agronomic traits of rice, using 128 recombinant inbred lines derived from ‘Emi-no-kizuna’ and ‘Tomohonami’. We detected two promising QTLs associated with GA: qGA4 on chromosome 4 and qGA8 on chromosome 8. qGA4 contributed highly to the greater percentage of perfect grains of the Emi-no-kizuna genotype. In the same region, we detected other QTLs associated with panicle number and spikelet number per panicle. In near-isogenic lines (NILs) in which Emi-no-kizuna alleles were introgressed in the genomic region of only the semi-dwarf 1 (sd1) locus (NIL_1) and both the sd1 locus and qGA8 (NIL_2), respectively, the percentage of perfect grains was significantly higher and the percentages of milky white, basal white, and white back grains were significantly lower than in Tomohonami; and the percentages of milky white and white back grains of NIL_2 were significantly lower than those of NIL_1. These results suggest that introgression in the sd1 region could improve GA, and that the addition of qGA8 could further improve GA. The culm lengths of the NILs were significantly shorter than that of Tomohonami, indicating improved lodging resistance. Grain weight of NIL_2 was significantly smaller than that of NIL_1, suggesting that the effect of qGA8 could be pleiotropic, or the gene that underlies qGA8 could be linked with genes associated with grain weight.

Abbreviations: ANOVA: analysis of variance; AT20: mean air temperature in the 20 days after heading; BW: basal white grain; CL: culm length; DAH: days after heading; GA: grain appearance; GW: 1000-grain weight; LOD: logarithm of odds; MW: milky white grain; NIL: near-isogenic line; PG: perfect grain; PL: panicle length; PN: panicle number; PTSN: putative total spikelet number; PVE: percentage of phenotypic variation explained; QTL: quantitative trait locus; RIL: recombinant inbred line; SN: spikelet number per panicle; SNP: single nucleotide polymorphism; WB: white back grain     

Involvement of microaerophilic iron-oxidizing bacteria in the iron-oxidizing process at the surface layer of flooded paddy field soil

Journal of Soils and Sediments - To reveal whether microaerophilic Fe(II)-oxidizing bacteria (FeOB) participate in the Fe(II) oxidation at the oxic-anoxic interface in flooded paddy field soil,...