Potential application of simple easy-to-use insertion-deletion (InDel) markers in citrus cultivar identification

We previously developed insertion-deletion (InDel) markers that distinguish three genotypes (two homozygous and one heterozygous) of diverse citrus cultivars. These InDel markers were codominant and could be clearly detected by using simple agarose gel electrophoresis. We sought to establish a method for cultivar identification using these 28 InDel markers to genotype 31 citrus cultivars. The results revealed that a minimum of 6 markers were required to identify individuals using the three-genotype classification method. Furthermore, we found that a simple method for distinguishing between two genotypes (homozygous and heterozygous) could be used to identify individuals using a minimum of 7 markers. Our findings provide a basis for the development of simple and rapid citrus cultivar identification methods.     

Dehydroeffusol Rescues Amyloid β25–35-Induced Spatial Working Memory Deficit

Plant Foods for Human Nutrition - Amyloid β (Aβ) peptides produced from the amyloid precursor protein, a transmembrane protein, are neurotoxic and blocking the neurotoxicity may lead to...     

Ventilation of seawater during bleeding suppresses the occurrence of burnt meat in yellowtail <Emphasis Type="Italic">Seriola quinqueradiata</Emphasis>

Fifteen farmed yellowtail Seriola quinqueradiata were randomly selected from sea cages. Five fish were slaughtered instantly by cutting the spinal bulb (control group). The remaining ten fish were left for 20 min in a narrow net and then they were slaughtered by cutting the spinal bulb instantly. These ten fish were bled in two water tanks. In one tank, the water was aerated (ventilation group), while the other tank was not aerated (burnt group). The ATP ratio in the control group (83.8 ± 4.1%) was higher than the ventilation group (37.2 ± 22.1%) and the burnt group (5.8 ± 6.0%) at 3 h after slaughter. The K value of the burnt group increased remarkably at 3 h after slaughter, and was significantly higher than those for the other groups. The ventilation group showed a recovery in available energy for physiological maintenance to the same level as the control group during bleeding, as well as a significantly slowed loss of muscular firmness compared to the burnt group. This study shows that the yellowtail continued to respire and take up oxygen until complete physiological death, and elevated oxygen levels during bleeding slowed the postmortem changes and suppressed the occurrence of burnt meat.     

Assemblages of vertical migratory mesopelagic fish in the transitional region of the western North Pacific

Assemblages of vertical migratory mesopelagic fish are described for the nighttime upper 20 m layer of the transitional and adjacent waters in the western North Pacific. Twenty-three mesopelagic fish species belonging to six families and 16 genera were collected during June 1996. Family Myctophidae was most speciose, represented by 17 species, followed by Sternoptychidae (two spp.) and Bathylagidae (one spp.). Six assemblages are recognized based on their species composition: Subarctic, Northern Transition, Southern Transition, Subtropical, Slope Water and Seamount Assemblages. These assemblages are also characterized by size composition and biomass of the catches. The distributions of the Subarctic, Northern Transition, Southern Transition and Subtropical Assemblages are approximately defined by the Kuroshio and Oyashio fronts and the Subarctic Boundary. Slope Water and Seamount Assemblages are closely related to the slope water around the shelf edge off Japan and waters around the Shatsky Rise and the Emperor Seamount chain, respectively. The importance of the Southern Transition Assemblage area between the Kuroshio front and Subarctic Boundary as spawning and nursery grounds of many subarctic, transitional and subtropical fishes is elucidated, based on the geographical distribution patterns of their larvae and juveniles.     

The TLR Expression Pattern on Monocyte-Derived Macrophages for Lipopolysaccharid Stimulation of Calves

In this paper, toll-like receptor expression pattern in monocytes-derived macrophages by lipopolysaccharid （LPS） stimulation was examined. Jugular venous blood samples from 4 Japanese calves were obtained and the peripheral blood mononuclear cells （PBMC） were isolated. The PBMC were cultured for 7 d so as to collect monocytes-derived macrophages in Repcell. The PBMC were stimulated by LPS for 24 h and the mRNA expression pattern of TLR and cytokines in monocytes-derived macrophages （Mod-Mφ） was analyzed. Results showed that LPS stimulation of Mod-Mφ could increase the mRNA levels of the genes of TNF-α, IL-6, and IL-8. In addition, the mRNA levels of the genes of TNF-α and IL-6 in the group of LPS stimulation were most significantly （P 〈 0.01） higher than those in control group and the mRNA levels of TLR1, 3, 5, 8, and 10 were significantly （P 〈 0.05） decreased after LPS stimulation. There was no difference in the mRNA expressions of TLR2, 4, 6, and 7 between the groups of the control and LPS stimulation. Besides, expression of TLR9 was not found. It suggested that monocytes-derived macrophages could respond to LPS and they might take an important role in the innate immunity. The important function of the cells might contribute to better disease treatment.     

Nephrocalcinosis formation by soy isoflavones in male rats

Isoflavones (IFs), found in the form of both aglycones and glucosides in soybean foods, induce weak estrogenic activities. Although IFs have a number of health benefits, it was previously reported that IFs cause nephrocalcinosis (NC) in the kidney of male Fischer 344 (F344) rats. The present study aims to elucidate the safety of IFs by focusing on IF-induced NC formation in rats. Fermented soybean extract (FSE) containing 420 mg/g isoflavone aglycones was orally administered to male F344 and Sprague-Dawley (SD) rats for 28 days. FSE induced NC formation in the kidney of F344 rats, but not in SD rats. However, absorption of IFs did not differ between F344 and SD rats. NC formation and its severity of FSE were histologically compared with those of soybean extract (SE) containing 518 mg/g isoflavone glucosides in F344 rats. There were no differences in the number of NC formations and the extent of calcium deposit between FSE and SE groups. To examine the dose effect of FSE on NC formation, doses of 20, 140, or 1000 mg/kg FSE were administered to F344 rats for 90 days. NC formation was observed in the 140 and 1000 mg/kg groups. These results indicated that a high dose of oral administration of IFs induced NC formation depending on the strain of rat.     

Anesthesia and acoustic stress-induced intra-uterine growth retardation in mice

Stress interferes with reproduction, adversely influencing implantation and fetal growth, and sometimes even leading to abortion. Here, we attempted to evaluate the early gestational effects of uncomfortable sound on pregnant mice and their offspring. Ten-week-old pregnant Jcl:ICR mice were exposed to sound (100 dB, random frequency between 9-34 kHz) for 8 hours on the 3(rd), 5(th) and 7(th) gestational days (GD). The effects of general anesthesia were also investigated, with or without acoustic stress. All groups were examined on the 18(th) GD for fetal growth. Fetal weight, number of ossified sacrococcygeal vertebrae and placental weight were all significantly reduced (P<0.0001) when stress was induced on the 7(th) GD, but not on the 3(rd) or 5(th) GD. This intra-uterine growth retardation (IUGR) was significantly inhibited by general anesthesia (P<0.0001), although general anesthesia alone induced significant IUGR (P<0.0001) when compared with control mice. This suggests that acoustic exposure indirectly exerts an effect on fetal growth, possibly via a psycho-maternal pathway. We also found that analysis of the number of ossified sacrococcygeal vertebrae is the most sensitive tool for the study of IUGR.     

An in vitro screening assay to discover novel inhibitors of 4-coumarate:CoA ligase

4-Coumarate:CoA ligase (4CL, EC 6.2.1.12) exists only in plants and plays an important role in the phenylpropanoid pathway. Identification of inhibitors targeting 4CL provides a novel approach for developing effective plant growth inhibitors (PGIs). The full-length gene of tobacco 4CL (Nt4CL1) was cloned and expressed in Escherichia coli Cast & Chalm. The recombinant 4CL protein was extracted and purified by several purification steps including gel-filtration and anion-exchange chromatography. 4CL activity assay was miniaturized and optimized using a 96-well microplate and a reader. Among 28 existing herbicides, propanil and swep strongly inhibited in vitro 4CL enzyme activity, and they were selected for further studies. The process of this assay can be developed into a high-throughput screening system of PGI targeting 4CL in the phenylpropanoid pathway.     

Transgenic Pigs with Pancreas-specific Expression of Green Fluorescent Protein

The development and regeneration of the pancreas is of considerable interest because of the role of these processes in pancreatic diseases, such as diabetes. Here, we sought to develop a large animal model in which the pancreatic cell lineage could be tracked. The pancreatic and duodenal homeobox-1 (Pdx1) gene promoter was conjugated to Venus, a green fluorescent protein, and introduced into 370 in vitro-matured porcine oocytes by intracytoplasmic sperm injection-mediated gene transfer. These oocytes were transferred into four recipient gilts, all of which became pregnant. Three gilts were sacrificed at 47–65 days of gestation, and the fourth was allowed to farrow. Seven of 16 fetuses obtained were transgenic (Tg) and exhibited pancreas-specific green fluorescence. The fourth recipient gilt produced a litter of six piglets, two of which were Tg. The founder Tg offspring matured normally and produced healthy first-generation (G1) progeny. A postweaning autopsy of four 27-day-old G1 Tg piglets confirmed the pancreas-specific Venus expression. Immunostaining of the pancreatic tissue indicated the transgene was expressed in β-cells. Pancreatic islets from Tg pigs were transplanted under the renal capsules of NOD/SCID mice and expressed fluorescence up to one month after transplantation. Tg G1 pigs developed normally and had blood glucose levels within the normal range. Insulin levels before and after sexual maturity were within normal ranges, as were other blood biochemistry parameters, indicating that pancreatic function was normal. We conclude that Pdx1-Venus Tg pigs represent a large animal model suitable for research on pancreatic development/regeneration and diabetes.