Development of a high-throughput screening method using a cell-based, lawn format assay for the identification of novel plant defense activators from combinatorial peptide libraries

Plants respond to attack by pathogens through various defense mechanisms. These defense responses are triggered by a variety of molecules derived from pathogenic microorganisms as well as host plants. In this study, we developed a high-throughput screening method using a cell-based lawn format assay for the identification of novel peptides that can induce plant defense responses from combinatorial peptide libraries. Solid-phase peptide libraries were synthesized using a photocleavable linker and immobilized using agarose gel. The peptides were partially cleaved from beads, and the agarose gel was layered on the tobacco cells. The defense response was then observed by detecting the generated H2O2 using a sensitive H2O2 indicator dye, N-(carboxymethylaminocarbonyl)-4,4'-bis(dimethylamino)diphenylamine sodium salt (DA-64). Using this assay format, a 6859-member peptide library based on the sequence of flagellin-derived peptides was screened, and several structural features important for the activity were obtained.     

Estimating harvest costs and projecting quantities of logging residues for small-scale forestry in Nasushiobara,Tochigi Prefecture,Japan

We used GIS on a regional scale to estimate and compare supply potentials and costs of small-scale logging systems, a mini-forwarder and a 4-ton truck operated by private logging contractors, and manual logging and a light truck operated by individual forest own- ers, with the mechanized operational system of the Forest Owners＇ Asso- ciation. Total potential yields of timber and logging residues were esti- mated as 418,895 m3 and 254,962 m3, respectively. The economic bal- ances were estimated and available amounts were projected as supply potentials from profitable sub-compartments. As a result, available amounts of timber and logging residues were estimated at 375,466 m3 （89.9%） and 203,850 m3 （80.0%）, respectively. Because their transport expenses were lower than for other systems the most profitable sub-compartments were operated by private logging contractors who sold logging residues at a plant. The profitable sub-compartments operated by individual forest owners were few because the extracting distances were usually greater than 20 m. Raising logging residue prices from 3,000 yen·m-3 to 4,080 yen·m-3 or 6,800 yen·m-3, and establishing forest roads, which reduced some extracting distances to less than 20 m, increased the number and area of profitable sub＊compartments, and increased available amounts of logging residues.     

Possible degradation of type I collagen in relation to yellowtail muscle softening during chilled storage

ABSTRACT:   In this paper, the detection of type I collagen degradation during the softening phenomenon of yellowtail muscle, was examined. Acid soluble collagen was isolated from dorsal ordinary muscle at death and after 24-h chilled storage. In the abundant ratio of subunit components, an increase in β12 chain (5.4 points) and a decrease in components with molecular weights larger than γ chain (7.0 points) after 24-h chilled storage, was found. Type I collagen was detected in the alkali-soluble fraction by SDS-PAGE. Its amount calculated from hydroxyproline contents in alkali-soluble fraction was increased from 0.097 mg/g muscle to 0.155 mg/g muscle during 24-h storage. The increased alkali-soluble collagen (0.058 mg/g muscle) was about 1.4% of whole collagen. These results suggest that a slight decomposition of type I collagen of yellowtail muscle may occur and subsequently becomes alkali-soluble corresponding to postmortem softening.     

Changes in myoglobin content and proximate compositions of the dorsal ordinary muscles of full-cycle cultured Pacific bluefin tuna Thunnus orientalis (Temminck et Schlegel) with body size

We investigated the changes in myoglobin (Mb) content and proximate compositions of the cephalad parts of the dorsal ordinary muscles (Ce‐DOM) of full‐cycle cultured (FC) Pacific bluefin tuna [body length: 42.6–115.4 cm, body weight (BW): 1.7–38.4 kg, killed from July 2004 to October 2005]. The Mb content of Ce‐DOM increased (P<0.01) until a BW of 7.0 kg was reached. However, the Mb content of Ce‐DOM had a wide variety above a BW of 7.0 kg. On the other hand, the moisture, protein and ash contents of Ce‐DOM of FC tuna decreased (P<0.05 and 0.01) with an increase in BW. However, the lipid content of Ce‐DOM increased gradually above about a BW of 20.0 kg. Furthermore, the Mb content in protein of the Ce‐DOM of FC tuna increased with an increase in BW. These results indicated that the increase in Mb content of Ce‐DOM of FC tuna was not related to changes in proximate composition between a BW of 1.7 and 38.4 kg.     

The effectiveness of a new physical property evaluation method of kamaboko which applied the principal component analysis to the puncture and stress relaxation tests

ABSTRACT:   The physical properties of 49 commercial kamabokos were measured by the puncture and stress relaxation tests. The principal component analysis was applied to the physical parameters of both tests, and their cumulative contribution ratios were over 90% with the first and second principal components, respectively. The comparison among the kamabokos was carried out using the synthetic physical parameters. The kamabokos produced in same area showed the peculiar distribution. The relative positional relation of kamabokos measured by the stress relaxation test was different from that by the puncture test for many of the kamabokos. Physical property evaluation using a principal component analysis is very effective for intensiveness of many measurement parameters. If much more kamabokos were measured by this method, regional characteristics of kamabokos would be clarified.     

Properties of scallop mantle collagen: its content,tissue distribution and thermal behavior

ABSTRACT:   To obtain fundamental information for the effective use of scallop Patinopecten yessoensis mantle, which is one of the underutilized marine resources. Some properties of collagen contained in the mantle were examined by chemical and histochemical techniques. Collagen content in the mantle varied annually, ranging from 0.98 to 1.72% of wet tissue, 7.7 to 12.6% of dry tissue and 13.5 to 26.5% of total protein, being relatively in high level of collagen content of invertebrate muscles. Collagen fiber was densely distributed in the inner connective tissue matrix of the mantle pallial, in contrast to the inner fold part which was rich in muscle fibers. The collagen contained in the crude collagen fraction (residue after alkali extraction), prepared from the mantle, was revealed to have considerably low solubility on hot-water extraction, constantly less than 20% of the total collagen at the temperatures in the range of 20–90°C.     

Maturity and reproduction of goneplacid crab <Emphasis Type="Italic">Carcinoplax vestita</Emphasis> (Decapoda,Brachyura) in Tokyo Bay

ABSTRACT:     Sexual maturity, morphological sexual dimorphism, and reproduction of the goneplacid crab Carcinoplax vestita were investigated in Tokyo Bay, Japan, from November 2002 to October 2003. The puberty molt in males was evidenced by changes in the relative size of the chelipeds and merus of the walking legs, and was estimated to occur at a size range of 13.20–18.85 mm carapace length. Post-pubertal females were identified by the relative size of the abdomen and puberty was estimated to occur at a size range of 12.81–15.46 mm carapace length. Sexual dimorphism in C. vestita was observed in all features that showed secondary sexual characteristics. Monthly changes of gonad index in males, and of seminal receptacle index and occurrence of sperm plugs in females were synchronized, and indicated that mating was intense in spring. Ovaries began developing in March. Ovigerous females were found in all months except December, but were clearly more abundant between August and October. Fecundity ranged from 7800 to 57 000 mature oocytes per female per batch and was highly correlated with body size. The results suggest that some females may spawn more than one batch per year.     

Application of ELISA-based kit for detecting AOZ and determining its clearance in eel tissues

Furazolidone, an antibacterial drug that was once widely used in the livestock industry and aquaculture, is now prohibited in numerous countries. It is difficult to detect residual furazolidone because it is readily metabolized in animal tissues but, by using and liquid chromatography coupled with tandem mass spectrometry, its metabolite, 3-amino-2-oxazolidinone (AOZ) can be detected. Here we describe the validity of an enzyme-linked immunosorbent assay (ELISA) kit to detect AOZ in Japanese eel Anguilla japonica tissue. ELISA is capable of detecting AOZ at 1.0 μg/kg in an eel sample with excellent accuracy and precision. Our results show that ELISA is suitable for regulatory purposes and for studying the fate of AOZ residues in eel treated with furazolidone. To measure the persistence of AOZ in eel tissues, eels (1.4–6.5g) were immersed in tanks containing 2 and 10 mg furazolidone/L for 3 h, and then maintained in a tank supplying well water for the next 160 days. The half-lives of AOZ, calculated from the linear terminal part of the excretion curve, were 25.0 days in muscle and 21.6 days in liver from fish exposed to 2 mg/L furazolidone. In the eels treated with 10 mg/L furazolidone, by contrast, high levels of AOZ were detected in liver and muscle, but the half-lives of AOZ were similar to those in fish treated with 2 mg/L furazolidone. The half-lives of AOZ in eel tissues were prolonged by the condition of low water temperature.