A histochemical method using a substrate of beta-glucuronidase for detection of genetically modified papaya

A histochemical assay for detecting genetically modified (GM) papaya (derived from Line 55-1) is described. GM papaya, currently undergoing a safety assessment in Japan, was developed using a construct that included a beta-glucuronidase (GUS) reporter gene linked to a virus coat protein (CP) gene. Histochemical assay was used to visualize the blue GUS reaction product from transgenic seed embryos. Twelve embryos per fruit were extracted from the papaya seeds using a surgical knife. The embryos were incubated with the substrate 5-bromo-4-chloro-3-indolyl-beta-D-glucuronide (X-Gluc) in a 96-well microtiter plate for 10-15 hours at 37 degrees C. Seventy-five percent of GM papaya embryos should turn blue theoretically. The histochemical assay results were completely consistent with those from a qualitative polymerase chain reaction (PCR) method developed by this laboratory. Furthermore, the method was validated in a five-laboratory study. The method for detection of GM papaya is rapid and simple, and does not require use of specialized equipment.     

Reaction between magnesium ammine complex compound and lithium hydride

The possibility of using ammonia as a hydrogen carrier is examined for the reaction between magnesium ammine complex MgCl₂(NH₃)₆ and lithium hydride LiH. Sample was milled at low temperature of −40 °C to avoid decomposition of MgCl₂(NH₃)₆ during the milling. The effects of milling time, milling speed (revolutions per minute), and catalysts on hydrogen storage properties were investigated by thermogravimetry, thermal desorption mass spectroscopy, and X-ray diffraction experiments. Experimental results indicated that a milled composite of Mg(NH₃)₆Cl₂ and catalyzed-LiH desorbed the ∼100% H₂ gas even at 125 °C in a closed system. The reverse reaction also proceeded by separately cooling MgCl₂ at lower temperature than 100 °C and heating LiNH₂ at 300 °C in the closed system.     

Detection of genetically modified organisms in foreign-made processed foods containing corn and potato

Investigations of the validity of labeling regarding genetically modified (GM) products were conducted using polymerase chain reaction (PCR) methods for foreign-made processed foods made from corn and potato purchased in the Tokyo area and in the USA. Several kinds of GM crops were detected in 12 of 32 samples of processed corn samples. More than two GM events for which safety reviews have been completed in Japan were simultaneously detected in 10 samples. GM events MON810 and Bt11 were most frequently detected in the samples by qualitative PCR methods. MON810 was detected in 11 of the 12 samples, and Bt11 was detected in 6 of the 12 samples. In addition, Roundup Ready soy was detected in one of the 12 samples. On the other hand, CBH351, for which the safety assessment was withdrawn in Japan, was not detected in any of the 12 samples. A trial quantitative analysis was performed on six of the GM maize qualitatively positive samples. The estimated amounts of GM maize in these samples ranged from 0.2 to 2.8%, except for one sample, which contained 24.1%. For this sample, the total amount found by event-specific quantitative analysis was 23.8%. Additionally, Roundup Ready soy was detected in one sample of 21 potato-processed foods, although GM potatoes were not detected in any sample.     

Performance tests of catalysts for the safe conversion of hydrogen inside the nuclear waste containers in Fukushima Daiichi

The safe decommissioning as well as decontamination of the radioactive waste resulting from the nuclear accident in Fukushima Daiichi represents a huge task for the next decade. At present, research and development on long-term safe storage containers has become an urgent task with international cooperation in Japan. One challenge is the generation of hydrogen and oxygen in significant amounts by means of radiolysis inside the containers, as the nuclear waste contains a large portion of sea water. The generation of radiolysis gases may lead to a significant pressure build-up inside the containers and to the formation of flammable gases with the risk of ignition and the loss of integrity.In the framework of the project “R&D on technology for reducing concentration of flammable gases generated in long-term waste storage containers” funded by the Japanese Ministry of Education, Culture, Sports, Science and Technology of Japan (MEXT), the potential application of catalytic recombiner devices inside the storage containers is investigated. In this context, a suitable catalyst based on the so-called intelligent automotive catalyst for use in a recombiner is under consideration. The catalyst is originally developed and mass-produced for automotive exhaust gas purification, and is characterized by having a self-healing function of precious metals (Pd, Pt and Rh) dissolved as a solid solution in the perovskite type oxides. The basic features of this catalyst have been tested in an experimental program. The test series in the REKO-4 facility has revealed the basic characteristics of the catalyst required for designing the recombiner system.     

Isolation and Characterization of Two Types of Xyloglucanases from a Phytopathogenic Fungus,Verticillium dahliae

Xyloglucan is a major hemicellulosic component in plant cell walls. Phytopathogenic fungi secrete cell wall-degrading enzymes on their infection to hosts, while the nature of the cell wall-lytic enzymes of such fungi are yet to be fully understood. Verticillium dahliae is a soil-borne fungus that causes vascular wilt diseases in a variety of commercially important crops worldwide. We purified two types of xyloglucanases, XEG12A and XEG74B, from the culture of naturally isolated Verticillium dahliae strain 2148. XEG12A showed a molecular size of 23 kDa with its maximal activity at pH 7.5. XEG12A specifically hydrolyzed xyloglucan with no activity on other β-glucans. XEG74B had a molecular size of 110 kDa with its optimum pH at 6.0. XEG74B primarily hydrolyzed xyloglucan, with a slight activity on β-1,3-1,4-glucan. Analysis of hydrolytic products of xyloglucanooligasaccharide (XXXGXXXG) by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) revealed that the both enzymes cleaved β-1,4-glucosidic linkage at the position of unbranched chain, while XEG74B showed a little fluctuation with the cleavage site. Both enzymes did not hydrolyzed xyloglucanoheptasaccharide (XXXG) at all. N-Terminal and internal amino acid sequencing of the enzymes revealed that XEG12A and XEG74B belonged to Glycoside Hydrolase (GH) Families 12 and 74, respectively. Based on these results we concluded that V. dahliae XEG12A and XEG74B were xyloglucan-specific endo-β-1,4-glucanases (EC 3.2.1.151).     

Levels of glutathione and related enzymes in yellowtail fish muscle subjected to ice storage in a modified atmosphere

We investigated the effects of ice storage in a modified atmosphere on levels of glutathione (GSH) and its related enzyme activities, the metmyoglobin to total myoglobin ratio (metMb%), and the K value (a freshness index) of yellowtail fish muscle. GSH in ordinary muscle (fast skeletal muscle) as well as in dark muscle (slow skeletal muscle) stored in air decreased. GSH in those muscles was almost unchanged during storage when packaged with an oxygen absorber or with an oxygen absorber-CO(2) generator. Glutathione disulfide in each type of packaging remained at low concentrations during storage. The GSH peroxidase activities of ordinary muscle and of dark muscle after 7 d of storage in air were lower than when packaged with the oxygen absorber or with the oxygen absorber-CO(2) generator. The GSH reductase (GR) activities of ordinary muscle at the 4th and 7th day of storage when packaged with the oxygen absorber showed a tendency to be lower than when stored in air. The GR activity of dark muscle in each type of packaging method was unchanged during storage. The packaging method did not influence the K values of either the ordinary muscle or the dark muscle during storage. The metMb% of dark muscle when packaged with the oxygen absorber was lower than in the other types of packaging during storage. Therefore, packaging with the oxygen absorber is an effective method to prevent the loss of GSH in fish meat as well as to reduce the discoloration during storage. PRACTICAL APPLICATION: This study examined whether modified atmosphere packaging preserves the level of GSH, which is unstable to oxidative stress, in fish muscle. The use of an oxygen absorber for packaging can allow us to take in a sufficient amount of the bioactive compound from fish meat after storage as well as fresh fish.