Recent advances in soybean transformation and their application to molecular breeding and genomic analysis

Herbicide-resistant transgenic soybean plants hold a leading market share in the USA and other countries, but soybean has been regarded as recalcitrant to transformation for many years. The cumulative and, at times, exponential advances in genetic manipulation have made possible further choices for soybean transformation. The most widely and routinely used transformation systems are cotyledonary node–Agrobacterium-mediated transformation and somatic embryo–particle-bombardment-mediated transformation. These ready systems enable us to improve seed qualities and agronomic characteristics by transgenic approaches. In addition, with the accumulation of soybean genomic resources, convenient or promising approaches will be requisite for the determination and use of gene function in soybean. In this article, we describe recent advances in and problems of soybean transformation, and survey the current transgenic approaches for applied and basic research in Japan.     

Essential thrombocythemia in a dog

A two-year old male Welsh Corgi was referred for persistent thrombocytosis and occasional seizure. Hematological findings indicated marked thrombocytosis, eosinophilia, basophilia and moderate anemia. Bone marrow examination revealed marked megakaryocytic hyperplasia with morphologic abnormality. A diagnosis of essential thrombocythemia was made and the treatment was initiated with combination chemotherapy and maintained by prednisolone and busulfan. The dog successfully achieved complete remission on 100 days after initial presentation and has been good in health without chemotherapy since then.     

Effect of growth hormone on estrogen receptor binding properties in the chick oviduct in vivo

The equilibrium dissociation constant (K_d) and the maximum binding capacity (B_max) of estrogen receptor in cytosolic and nuclear fractions in oviduct of pullets following various daily injections (1–10 times) of growth hormone (GH: 50 µg/chick) were examined by Scathchard analysis of specific [³H]estradiol‐17β (E₂) binding. The K_d values of receptor in both fractions decreased after three times of GH‐injection. In the B_max values, three times of the injection caused a marked decrease in that value in the cytosolic fraction with a concomitant increase in that in the nuclear fraction, whereas the total B_max (sum of B_max in the cytosolic and nuclear fractions) did not change. A similar relationship between the K_d values in the binding of the two fractions was also observed in 4–10 times of GH‐injection. However, in 4–10 times of GH‐injection the B_max of the both fractions and total B_max was greater than that in vehicle‐injection (control). When the chicks were injected with 6–10 times of GH‐injection, the weight of oviduct was increased. No change in the plasma concentrations of E₂ was found following GH‐ and vehicle‐injections into the chicks. The results suggest that growth hormone stimulates the growth of the chick oviduct by increasing the binding affinity and capacity of estrogen receptor.     

Lipophilic,hydrolytic and photolytic properties and fungicidal activity against Botrytis cinerea of 1-(4-substituted phenoxymethyl)-2, 2-dimethylpropyl imidazole-1-carboxylates and -thiocarboxylates

The sterol 14α-demethylation inhibitors 1-(4-substituted phenoxymethyl)-2, 2-dimethylpropyl imidazole-1-carboxylates and their corresponding thiocarboxylates were optimized for maximum in-vitro activity against Botrytis cinerea Pers. ex Fr. in terms of the lipophilic parameter π. The activity of both carboxylates and thiocarboxylates was strongly related with π and predicted to be maximum at π 1.38 and 0.72 respectively. However, the preventive efficacy of the carboxylates against B. cinerea on Solanum melongena L. grown in a greenhouse did not correlate with the parameter. Despite the unfavourable lower lipophilicity, compounds with p-alkoxy substituents were superior to those with any other substituent. Moreover, the p-methoxy-substituted thiocarboxylate 46 was much less effective in greenhouse tests than the corresponding carboxylate 15, despite their equivalent in-vitro activity. To clarify these discrepancies, hydrolytic and photolytic stabilities of several representative compounds including 15 and 46 were investigated. The carboxylates examined were much more labile to hydrolysis than the thiocarboxylate 46; however, the four thiocarboxylates studied were less stable to light than the carboxylates. Consequently, the above discrepancies were attributable mainly to the superior stability of the carboxylates to photolysis as compared to the thiocarboxylates.     

Effect of a novel rice fermented extract on gastric ulcers in horses

Gastric ulcers cause appetite loss, poor body condition, and colic in horses. This study investigated the protective effect of a rice fermented extract on the gastric mucosa in 17 healthy Thoroughbreds. For one month, horses in the rice fermented extract (nine horses) and control (eight horses) groups were orally administered a rice fermented extract (100%; 0.2 ml/kg, SID) and tap water (0.2 ml/kg), respectively. Gastric endoscopic images were obtained before and one month after rice fermented extract administration. The gastric ulcer score was lower after administration (median, 1; maximum, 2; minimum, 1) than before administration (median, 4; maximum, 4; minimum, 3) in the rice fermented extract group (P<0.05). In conclusion, the administration of a rice fermented extract for one month improves gastric mucosal lesions in Thoroughbreds with gastric ulcers.     

Karnovsky’s fixative prevents artifacts appearing as vacuolation derived from tissue processing in kidneys treated with antisense oligonucleotide

Antisense oligonucleotide (ASO) therapies have been identified as a new treatment modality for intractable diseases. In kidneys treated with ASOs, vacuoles, in addition to basophilic granules, are often observed in the proximal tubules. Some reports have described that these vacuoles are likely to be a secondary phenomenon resulting from the extraction of ASOs during tissue processing. In this study, we compared renal morphology after fixation with Karnovsky’s fixative or 4% paraformaldehyde phosphate buffer (PFA) with that of 10% neutral-buffered formaldehyde solution (NBF). Female Sprague-Dawley rats, intravenously treated four times with 50 mg/kg locked nucleic acid containing antisense oligonucleotides (LNA-ASOs) for 1 or 2 weeks, were examined. Microscopically, vacuoles and basophilic granules in the proximal tubules were observed in the kidneys fixed with NBF. Basophilic granules are indicative of the accumulation of ASOs. Moreover, some of the vacuoles also contained faint basophilic granules, suggesting that the vacuoles were relevant to the accumulation of ASOs. Although moderate vacuolation was observed in the proximal tubules, the majority of the vacuolated epithelia were negative for kidney injury molecule-1 on immunohistochemical staining. Vacuoles in the proximal tubules were not observed in samples subjected to Karnovsky’s fixation, although basophilic granules were observed. In samples subjected to PFA fixation, vacuoles and basophilic granules were observed in the proximal tubules, similar to those in samples subjected to NBF fixation. Overall, our findings demonstrated the possibility of overestimation of vacuolation due to artifacts during tissue processing when using conventional NBF fixation. Karnovsky’s fixative is considered a useful alternative for distinguishing artificial vacuoles from true nephrotoxicity.     

Effect of Inositol Hexaphosphate (IP6) on Serum Uric Acid in Hyperuricemic Subjects: a Randomized,Double-Blind,Placebo-Controlled,Crossover Study

Plant Foods for Human Nutrition - Inositol hexaphosphate (IP6), a food constituent with various health benefits, has been shown to suppress postprandial elevations of serum uric acid (SUA) levels...     

Species identification of imported and Japanese commercial green algal products based on phylogenetic analyses using the nrITS2 and 5S rDNA spacer regions

The identification of imported green algal products represents a critical challenge to the Japanese Customs Service in terms of ensuring effective control of imported quota items (species of genera Monostroma and Enteromorpha and their products). To assist officials in this difficult task, we tested the applicability of phylogenetic analyses using the nuclear-encoded ribosomal DNA internal transcribed spacer 2 (nrITS2) and 5S ribosomal DNA (rDNA) spacer regions to identify green algal products at the species level. Our nrITS2-based phylogenetic analysis was able to roughly separate the Ulva-related samples into three clades: the Ulva linza–procera–prolifera (LPP) complex and two other clades, and identified the Monostroma-related samples as M. nitidum. Subsequent detailed analysis within the LPP complex using the 5S rDNA spacer region divided the samples into two groupings (the U. linza clade and U. prolifera clade), while it detected possible hybrid samples of U. linza and U. prolifera from a number of commercial green algal products.     

Quantification of the distributions of gluten, starch and air bubbles in dough at different mixing stages by fluorescence fingerprint imaging

A method combining fluorescence fingerprint (FF) measurement with an imaging technique was developed to visualize the distributions of gluten and starch in dough. The fluorescence images of dough at the under-, optimum- and over-mixing stages were acquired in multiple excitation and emission wavelengths constructing the FF. This data containing the FF data of each pixel in the sample image was colored according to the similarity of the FF of each pixel and the FF of gluten or starch. Furthermore, the areas corresponding to air bubbles in the dough were masked in black. The resulting image, referred to as the FF pseudocolor image, showed the distributions of gluten and starch in the dough with the incorporated bubbles. Quantitative parameters concerning gluten and starch distributions and bubble area were extracted from the dough images at each mixing stage, showing the homogenization of gluten and starch and the incorporation of bubbles with mixing. This imaging method is suggested to have an advantage over conventional imaging methods since there is no need to preprocess the sample, and any constituent in the sample can be visualized as long as there is information about the FF of the pure target constituent.