Analysis of Microsatellite DNA Polymorphisms in Rockfish Sebastes thompsoni and Application to Population Genetics Studies

Population differentiation and relationships among 6 natural rockfish populations collected from northern coastal seas around Japan were assayed using microsatellite DNA loci. Seven loci examined were polymorphic in all populations. The number of alleles per locus ranged from 6.7 to 9.3, and the average of observed and expected heterozygosity ranged from 0.63 to 0.68, and from 0.66 to 0.69, respectively. The observed genotype frequencies at each locus were almost in agreement with Hardy-Weinberg expectations with two exceptions (P < .05). The allele frequencies of 16 population pairs were significantly different (P < .05). Genetic distance (D _A ) between 6 populations ranged from 0.03 to 0.08. According to a neighbor-joining tree generated from the D _A values, the 6 populations fell into 3 clusters. These clusters were correlated with the geographical positions of each population; larval dispersions due to water current were also found to have an effect on these results. Received May 2, 2000; accepted July 17, 2000.     

A preliminary study of gut dysbiosis in children with food allergy

Gut microbiota of food allergic children was analyzed by high throughput 16S rRNA gene sequencing. Signs of gut dysbiosis, which is likely associated with gut inflammation, was observed in children with food allergies. For example, decreased abundance of genus Akkermansia but increased abundance of Veillonella was found in children with food allergy in comparison with healthy control children.     

The role of QTLs in the breeding of high-yielding rice

Food shortages have once again become a serious problem, not only because of world population growth but also as a result of escalating demand for crops as a substrate for biofuels. The production of improved plant varieties, especially major crops such as rice, is urgently required to increase yield. The completion of the sequencing of the rice genome has made it possible to clone and analyze quantitative trait loci (QTLs). Furthermore, the development of high-throughput sequencing and genotyping technologies has improved spectacularly the accuracy of QTL analysis. In this review article, we focus on the potential roles of major QTLs in the selection for agronomic traits in rice and discuss the application of high-throughput technologies for high-resolution genetic analysis.     

Backbone resonance assignments for the cytoplasmic region of the Mg2+ transporter MgtE in the Mg2+-unbound state

Magnesium ion (Mg²⁺) is an essential metal element for life, and has many cellular functions, including ATP utilization, activation of enzymes, and maintenance of genomic stability. The intracellular Mg²⁺ concentration is regulated by a class of transmembrane proteins, called Mg²⁺ transporters. One of the prokaryotic Mg²⁺ transporters, MgtE, is a 450-residue protein, and functions as a dimer. We previously reported that MgtE exhibits the channel-like electrophysiological property, i.e., it permeates Mg²⁺ according to the electrochemical potential of Mg²⁺. The Mg²⁺-permeation pathway opens in response to the decrease of the intracellular Mg²⁺ concentration, while it is completely closed at the intracellular Mg²⁺ concentration of 10 mM. The crystal structures of the MgtE dimer revealed that the Mg²⁺-sensing cytoplasmic region consists of the N and CBS domains. The Mg²⁺-bound state of MgtE adopts a compact, globular conformation, which is stabilized by the coordination of a number of Mg²⁺ ions between these domains. On the other hand, in the Mg²⁺-unbound state, these domains are far apart, and fixed by the crystal packing. Therefore, structural analyses in solution were awaited, in order to characterize the Mg²⁺-dependent alteration of the MgtE structure and dynamics relevant to its gating. In this paper, we report the backbone resonance assignments of the dimer of the cytoplasmic region of the MgtE from Thermus thermophilus with a molecular weight of 60 KDa, in the Mg²⁺-unbound state.     

Purification and characterization of a novel O-methyltransferase from Flammulina velutipes

An enzyme catalyzing the methylation of phenolic hydroxyl groups in polyphenols was identified from mycelial cultures of edible mushrooms to synthesize O-methylated polyphenols. Enzyme activity was measured to assess whether methyl groups were introduced into (?)-epigallocatechin-3-O-gallate (EGCG) using SAM as a methyl donor, and (?)-epigallocatechin-3-O-(3-O-methyl)-gallate (EGCG3″Me), (?)-epigallocatechin-3-O-(4-O-methyl)-gallate (EGCG4″Me), and (?)-epigallocatechin-3-O-(3,5-O-dimethyl)-gallate (EGCG3″,5″diMe) peaks were detected using crude enzyme preparations from mycelial cultures of Flammulina velutipes. The enzyme was purified using chromatographic and two-dimensional electrophoresis. The purified enzyme was subsequently analyzed on the basis of the partial amino acid sequence using LC–MS/MS. Partial amino acid sequencing identified the 17 and 12 amino acid sequences, VLEVGTLGGYSTTWLAR and TGGIIIVDNVVR. In database searches, these sequences showed high identity with O-methyltransferases from other mushroom species and completely matched 11 of 17 and 9 of 12 amino acids from five other mushroom O-methyltransferases.     

A simple method of markerless gene deletion in Staphylococcus aureus

Staphylococcus aureus is a Gram-positive pathogen that causes opportunistic infections and a wide variety of diseases. Methicillin-resistant S. aureus (MRSA) is frequently isolated as multidrug-resistant in nosocomial and community infections. Molecular genetic manipulation is an important tool for understanding the molecular mechanism of S. aureus infection. However the number of available antibiotic markers is limited due to multidrug resistance. In this study, we constructed two Escherichia coli-S. aureus shuttle vectors, pKFT and pKFC, that carry a temperature-sensitive origin of replication in S. aureus, lacZ(a) enabling a simple blue-white screening in E. coli, an ampicillin resistant gene, and either a tetracycline resistance gene or a chloramphenicol resistance gene. We report a simple technique using pKFT to construct a markerless gene deletion mutant in S. aureus by allelic replacement without the use of a counter-selection marker. Subculture twice at 25 °C was critical to promote an allelic exchange rate in S. aureus. This technique is very simple and useful to facilitate genetic research on S. aureus.     

Characterization of phospholipids in membrane vesicles derived from Pseudomonas aeruginosa

Many Gram-negative bacteria release membrane vesicles (MVs), but their phospholipid properties are poorly understood. Phosphatidylglycerol was present at high levels in MVs derived from Pseudomonas aeruginosa, but not in the cellular outer membrane. The ratio of stearic acid in MVs was high compared to that in the cellular outer membrane. These findings suggest that membrane rigidity is associated with MV biogenesis.     

Design and synthesis of new prostaglandin D₂ receptor antagonists

To identify new cost-effective prostaglandin D? (DP) receptor antagonists, a series of novel 3-benzoylaminophenylacetic acids were synthesized and biologically evaluated. Among those tested, some representative compounds were found to be orally available. Receptor selectivity and rat PK profiles were also evaluated. The structure-activity relationship (SAR) study is presented.