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Tsuyoshi Tanaka

Publications and source records attributed to Tsuyoshi Tanaka.

At least 19 recordsLinked to original sources
PubMed2026

Genome analysis of the glycosphingolipid-producing green alga tetraselmis sp. NKG400013.

Microalgae are gaining attention as sustainable resources for the production of valuable compounds, including biofuels, pigments, and bioactive metabolites. To support metabolic engineering and genome editing approaches aimed at enhancing these traits, high-quality genome assemblies are essential; however, genomic information remains limited for many microalgal lineages. Tetraselmis sp. NKG400013 is a green alga known for high glycosphingolipid accumulation with distinctive structural features. Here, we report a draft genome assembly of this strain generated using PacBio HiFi sequencing and transcriptome-supported annotation. The assembled genome spans 423.7 Mbp, with 74.5% repetitive sequences and 15,322 predicted protein-coding genes. Comparative analyses across 11 green algal species revealed a positive correlation between genome sizes and repeat contents, indicating that transposable element expansion, particularly long terminal repeat retrotransposons, has substantially contributed to genome enlargement in Tetraselmis. Genome-wide functional annotation and ortholog inference identified core enzymes required for glycosylceramide biosynthesis. Both sphingolipid Δ4 and Δ8 desaturases were identified in Tetraselmis and their coexistence suggests an expanded capacity for long-chain base modification that may underlie its distinctive glycosphingolipid profile. These results establish a genomic framework for understanding the high glycosphingolipid-producing capacity of NKG400013 and provide insights into the evolutionary diversification of sphingolipid metabolism in green algae.

Chlorophyta↗
PubMed2005

Electrochemical probe for on-chip type flow immunoassay: immunoglobulin G labeled with ferrocenecarboaldehyde.

Labeling of ferrocenecarboaldehyde (Fc-CHO) to immunoglobulin G (IgG) via formation of Schiff-base and its reduction was investigated for construction of an electrochemical probe for miniaturized amperometric flow immunoassay. Approximately eight molecules of Fc-CHO were labeled to IgG and the reversible redox property of ferrocene was observed. Labeling efficiency improved by over three times as compared to the conventional method using ferrocenemonocarboxylic acid (Fc-COOH). Also, binding affinity of IgG labeled with Fc-CHO to its antigen, IgE, was investigated by enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance assay. IgG labeled with Fc-CHO that retained eight ferrocene moiety showed sufficient binding affinity to its antigen and the current response obtained in the flow electrochemical detection system increased by 14-fold as compared with IgG labeled with Fc-COOH when applying the potential of 390 mV vs. Ag/AgCl. The minimum detectable concentration of IgG labeled with Fc-CHO was 0.06 microM. IgG labeled with Fc-CHO demonstrate biochemical and electrochemical properties that are useful for electrochemical immunosensors.

Animals↗
PubMed2004

Fabrication of amino silane-coated microchip for DNA extraction from whole blood.

A simple microchip device for DNA extraction was constructed based on electrostatic interactions between surface amine groups and DNA. Microchannel was fabricated on silicon wafer by photolithography and coated with 3-aminopropyltriethoxysilane (APTES) or 3-[2-(2-aminoethylamino)-ethylamino]-propyltrimethoxysilane (AEEA) to introduce amine groups on the surface. Determination of the number of surface amine groups and optimization of DNA capture condition were demonstrated to characterize the microchip. Capacities of capturing DNA were approximately 97 ng/cm2 in APTES and 194 ng/cm2 in AEEA modified microchips, respectively. The amount of DNA captured in the microchip increased depending on surface amine density. Furthermore, DNA extraction using amine-coated microchip from whole blood was examined. Quantification of DNA and proteins in washing or eluting fraction indicates that proteins were removed at washing steps and only DNA was effectively eluted by changing alkalinity of buffer from pH 7.5 to 10.6. The amount of DNA extracted from whole blood was approximately 10 ng and its recovery ratio was 27-40%. Performance of PCR for the eluted fraction indicates that DNA extracted from whole blood was well purified using amine-coated microchip.

Adsorption↗
PubMed2004

Detection of biomolecular interaction between biotin and streptavidin on a self-assembled monolayer using magnetic nanoparticles.

For developing a magnetic bioassay system, an investigation to determine the presence of a specific biomolecular interaction between biotin and streptavidin was done using magnetic nanoparticles and a silicon substrate with a self-assembled monolayer. Streptavidin was immobilized on the magnetic particles, and biotin was attached to the monolayer-modified substrate. The reaction of streptavidin-modified magnetic particles on the biotin-modified substrate was clearly observed under an optical microscope. The magnetic signals from the particles were detected using a magnetic force microscope. The results of this study demonstrate that the combination of a monolayer-modified substrate with biomolecule-modified magnetic particles is useful for detecting biomolecular interactions in medical and diagnostic analyses.

Binding Sites↗
PubMed2004

Magnetic cell separation using antibody binding with protein a expressed on bacterial magnetic particles.

Bacterial magnetic particles (BacMPs) are efficient platforms of proteins for surface display systems. In this study, mononuclear cells from peripheral blood were separated using BacMPs expressing protein A on the BacMP membrane surface (protein A-BacMPs), which were complexed with the Fc fragment of anti-mouse IgG antibody. The procedure of positive selection involves incubation of mononuclear cells and mouse monoclonal antibodies against different cell surface antigens (CD8, CD14, CD19, CD20) prior to treatment with protein A-BacMP binding with rabbit anti-mouse IgG secondary antibodies. Flow cytometric analysis showed that approximately 97.5 +/- 1.7% of CD19(+) and CD20(+) cells were involved in the positive fraction after magnetic separation. The ratio of the negative cells in the negative fraction was approximately 97.6 +/-1.4%. This indicates that CD19(+) and CD20(+) cells can be efficiently separated from mononuclear cells. Stem cell marker (CD34) positive cells were also separated using protein A-BacMP binding with antibody. May-Grunwald Giemsa stain showed a high nuclear/cytoplasm ratio, which indicates a typical staining pattern of stem cells. The separated cells had the capability of colony formation as hematopoietic stem cells. Furthermore, the inhibitory effect of magnetic cell separation on CD14(+) cells was evaluated by measurement of cytokine in the culture supernatant by ELISA when the cells were cultured with or without lipopolysaccharide (LPS). The induction of IL1-beta, TNFalpha, and IL6 was observed in the presence of 1 ng/mL LPS in all fractions. On the other hand, in the absence of LPS, BacMPs had little immunopotentiation to CD14(+) cells as well as that of artificial magnetic particles, although TNFalpha and IL6 were slightly induced in the absence of LPS in the positive fraction.

Binding Sites, Antibody↗
PubMed2004

Evolution of vitamin B6 (pyridoxine) metabolism by gain and loss of genes.

Vitamin B(6) (VB6) functions as a cofactor of many diverse enzymes in amino acid metabolism. Three metabolic pathways for pyridoxal 5'-phosphate (PLP; the active form of VB6) are known: the de novo pathway, the salvage pathway, and the fungal type pathway. Most unicellular organisms and plants biosynthesize VB6 using one or two of these three biosynthetic pathways. However, animals such as insects and mammals do not possess any of the pathways and, thus, need to intake VB6 in their diet to survive. It is conceivable that breakdowns of these pathways occurred in the evolutionary lineages of insects and mammals, and one of the major reasons for this would be the loss of pertinent genes. We studied the evolution of VB6 biosynthesis from the view of the gain and loss of 10 pertinent genes in 122 species whose genome sequences were completely determined. The results revealed that each gene in the pathways was lost more than once in the entire evolutionary lineages of the 122 species. We also found the following three points regarding the evolution of PLP biosynthesis: (1) the breakdown of the PLP biosynthetic pathways occurred independently at least three times in animal lineages, (2) the de novo pathway was formed by the generation of pdxB in gamma-proteobacteria, and (3) the order of the gene loss in VB6 metabolism was conserved among different evolutionary lineages. These results suggest that the evolution of VB6 metabolism was subject to gains and frequent losses of related genes in the 122 species examined. This dynamic nature of the evolutionary changes must have been responsible for the breakdowns of the pathways, resulting in profound differentiation of heterotrophy among the species.

Animals↗
PubMed2004

Single nucleotide polymorphism detection in aldehyde dehydrogenase 2 (ALDH2) gene using bacterial magnetic particles based on dissociation curve analysis.

Single nucleotide polymorphism (SNP) detection for aldehyde dehydrogenase 2 (ALDH2) gene based on DNA thermal dissociation curve analysis was successfully demonstrated using an automated system with bacterial magnetic particles (BMPs) by developing a new method for avoiding light scattering caused by nanometer-size particles when using commercially available fluorescent dyes such as FITC, Cy3, and Cy5 as labeling chromophores. Biotin-labeled PCR products in ALDH2, two allele-specific probes (Cy3-labeled detection probe for ALDH2*1 and Cy5-labeled detection probe for ALDH2*2), streptavidin-immobilized BMPs (SA-BMPs) were simultaneously mixed. The mixture was denatured at 70 degrees C for 3 min, cooled slowly to 25 degrees C, and incubated for 10 min, allowing the DNA duplex to form between Cy3- or Cy5-labeled detection probes and biotin-labeled PCR products on SA-BMPs. Then duplex DNA-BMP complex was heated to 58 degrees C, a temperature determined by dissociation curve analysis and a dissociated single-base mismatched detection probe was removed at the same temperature under precise control. Furthermore, fluorescence signal from the detection probe was liberated into the supernatant from completely matched duplex DNA-BMP complex by heating to 80 degrees C and measured. In the homozygote target DNA (ALDH2*1/*1 and ALDH2*2/*2), the fluorescence signals from single-base mismatched were decreased to background level, indicating that mismatched hybridization was efficiently removed by the washing process. In the heterozygote target DNA (ALDH2*1/*2), each fluorescence signals was at a similar level. Therefore, three genotypes of SNP in ALDH2 gene were detected using the automated detection system with BMPs.

Aldehyde Dehydrogenase↗
PubMed2004

Spontaneous integration of transmembrane peptides into a bacterial magnetic particle membrane and its application to display of useful proteins.

An antimicrobial peptide, temporin L, and its derivative (TL-A2) were employed as anchor peptides and displayed streptavidin on a bacterial magnetic particle (BMP) membrane. The ribotoxin L3 loop (L3) and the arginine-chain peptide (R(12)), which are carrier peptides permeable to eukaryotic cell membranes, were also used. The peptides were labeled with a fluorescent dye, 4-fluoro-7-nitrobenzofurazan (NBD), at the N-terminal region (NBD-peptides) and mixed with BMPs. A specific integration of NBD-temporin L into a BMP membrane was observed. The basic amino acids in temporin L played an important role in the integration into BMPs. Biotin conjugated to the N-terminus of temporin L was integrated into a BMP membrane. The C-terminus of temporin L was incorporated into a BMP membrane, and the N-terminus was located on the BMP membrane surface. The present study shows that temporin L is a stable molecular anchor on BMPs by the binding of soluble protein to the N-terminus.

Antimicrobial Cationic Peptides↗
PubMed2004

Rapid and sensitive detection of 17beta-estradiol in environmental water using automated immunoassay system with bacterial magnetic particles.

A fully automated immunoassay of 17beta-estradiol (E2) was performed using anti-E2 monoclonal antibody immobilized on bacterial magnetic particles (AntiE2-BMPs) and alkaline phosphatase-conjugated E2 (ALP-E2). E2 concentration in environmental water samples was evaluated by decrease in luminescence based on competitive reaction. A linear correlation between the luminescence intensity and E2 concentration was obtained between 0.5 and 5 ppb. The minimum detectable concentration of E2 was 20 ppt. All measurement steps were done within 0.5 h. The analysis of environmental water samples by a commercially available ELISA kit and the BMP-based immunoassay gave good correlation plots with a correlation efficient of 0.992. These results suggest that the fully automated system using the BMP-based immunoassay has some advantages in the high rapidity and sensitivity of the measurement. This system will enable us to determine low E2 concentrations without sample condensation.

Bacterial Proteins↗
PubMed2003

Development and evaluation of an automated workstation for single nucleotide polymorphism discrimination using bacterial magnetic particles.

We designed an automated workstation for magnetic particle-based single nucleotide polymorphism (SNP) discrimination of ALDH genotypes. Bacterial magnetic particles (BMPs) extracted from Magnetospirillum magneticum AMB-1 were used as DNA carriers. The principle for SNP discrimination in this study was based on fluorescence resonance energy transfer (FRET) between FITC (donor) and POPO-3 (acceptor) bound to double-stranded DNA. The workstation is equipped with a 96-way automated pipetter which collects and dispenses fluids as it moves in x- and z-directions. The platform contains a disposable tip rack station, a reagent vessel serving as a stock for POPO-3 and FITC-labeled probes and a reaction station for a 96-well microtiter plate. BMPs were collected by attaching a neodymium iron boron sintered (Nd-Fe-B) magnet on the bottom of the microtiter plate. This system permits the simultaneous heating and magnetic separation of 96 samples per assay. The genotypes ALDH2*1 and ALDH2*2 were discriminated by calculating the relative fluorescence intensities on BMPs.

Aldehyde Dehydrogenase↗
PubMed2003

Microbulbifer arenaceous sp. nov., a new endolithic bacterium isolated from the inside of red sandstone.

An endolithic bacterium, strain RSBr-1, was isolated from the inside of a piece of red sandstone from coastal areas of Scotland. RSBr-1 was gram negative, oxidase and catalase positive, and cells were non-motile rods. Sodium was required for growth. The optimum sodium chloride concentration and pH for growth were 4% and pH 8.0, respectively. Eumelanin was produced in marine broth and in BY medium. RSBr-1 hydrolyzes chitin, esculin, gelatin, and starch, but not agar. Nitrate reduction is positive. Taxonomic characterization of this strain indicated that it belongs to the genus Microbulbifer. The difference between the aligned 16S rDNA sequences of RSBr-1 and the closest relative, M. elongata, is greater than the difference between the 16S rDNA sequences of M. hydrolyticus and M. elongata. On the basis of the phenotypic and genotypic comparison of this isolate with the other strains, RSBr-1 is proposed as a new species, Microbulbifer arenaceous, with type strain RSBr-1.

Alteromonadaceae↗
PubMed2003

Single nucleotide polymorphism genotyping of aldehyde dehydrogenase 2 gene using a single bacterial magnetic particle.

A single nucleotide polymorphism (SNP) genotyping for aldehyde dehydrogenase 2 gene (ALDH2) has been developed by using a nano-sized magnetic particle, which was synthesized intracellularly by magnetic bacteria. Streptavidin-immobilized on bacterial magnetic particles (BMPs) were prepared using biotin labeled cross-linkers reacting with the amine group on BMPs. ALDH2 fragments from genomic DNA were amplified using a TRITC labeled primer and biotin labeled primer pair, and conjugated onto BMP surface by biotin-streptavidin interaction. PCR product-BMP complex was observed at a single particle level by fluorescence microscopy. These complexes were treated with restriction enzyme, specifically digesting the wild-type sequence of ALDH2 (normal allele of ALDH2). The homozygous (ALDH2*1/*1), heterozygous (ALDH2*1/*2), and mutant (ALDH2*2/*2) genotypes were discriminated by three fluorescence patterns of each particle. SNP genotyping of ALDH2 has been successfully achieved at a single particle level using BMP.

Aldehyde Dehydrogenase↗
PubMed2003

Single nucleotide mismatch analysis using oligonucleotide probes synthesized on bacterial magnetic particle.

An approach to analyze mismatches using short and specific oligonucleotide probes directly synthesized on bacterial magnetic particles (BMPs) by phosphoramidite methods was exploited. Approximately 126 molecules of 4-mer oligonucleotides/particle were synthesized on BMPs with high reaction efficiencies. Hybridization between FITC-labeled oligonucleotides and chemically synthesized oligonucleotides on BMPs was performed. Perfect matched and mismatched hybridizations were successfully discriminated by using the oligonucleotide probes on BMPs.

Bacterial Proteins↗
PubMed2002

Pancreatic cancer with chylous ascites demonstrated by lymphoscintigraphy: successful treatment with peritoneovenous shunting.

A 60-year-old woman was admitted to our hospital because of abdominal distension caused by massive chylous ascites. Six months earlier, she was diagnosed with inoperative pancreatic head cancer and biliary tract stenting and radiation therapy were performed due to obstructive jaundice. The chylous ascites was observed by lymphoscintigraphy with 99mTc-labeled human serum albumin. It showed upper abdomen accumulation and leakage into the abdominal cavity, and a diagnosis of chylous ascites caused by obstruction or rupture of lymphocele due to pancreatic cancer was made. Chylous ascites did not improve with a conservative therapy, including low-fat diet and administration of flousemide, but was successfully treated with a peritoneovenous shunt using a Denver shunting tube. In association with the placement of the shunt, she lived for 15 months after the first diagnose of pancreatic cancer without pancreatoduodenectomy.

Chylous Ascites↗
PubMed2002

Cadmium recovery by a sulfate-reducing magnetotactic bacterium, Desulfovibrio magneticus RS-1, using magnetic separation.

Cadmium recovery by a sulfate-reducing magnetotactic bacterium, Desulfovibrio magneticus strain RS-1, was investigated. D. magneticus precipitated >95% of cadmium at an initial concentration of 1.3 ppm in the growth medium. Electron microscopic analysis revealed that D. magneticus formed electron-dense particles on its surface when cultivated in the presence of cadmium ions (Cd2+). Sulfide was also found in the precipitate, and the composition ratio of sulfide/cadmium was 0.7. Sixty percent of viable RS-1 cells was recovered by a simple magnetic separation revealing the removal of 58% cadmium from the culture medium.

Cadmium↗
PubMed2002

[CT evaluation of infectious colitis].

Computed tomography (CT) is useful for evaluating the diagnosis of gastrointestinal disease, such as infectious colitis, in patients with severe pain and bloody diarrhea. During the 7 years between November 1993 and October 2000, 34 patients with infectious colitis (18 male, 16 female; mean age 42 +/- 19 yrs), received emergency CT and colonoscopy because of severe abdominal pain and dysentery. The following organisms were isolated: pathogenic Escherichia coli (12), 6 of which were O157:H7 (O-157), Salmonella species (11), Campylobacter species (5), Vibrio parahaemolyticus (3), Yersinia enterocolotica (2) and Shigella species (1). Thickening of the intestinal wall greater than 10 mm was seen in the ascending colon in the 6 cases with E. coli O 157, in 5/11 cases with Salmonella, 4/5 with Campylobacter and 1/6 with non-O157 pathogenic E. Coli. Marked intestinal wall thickening, greater than 20 mm, was seen in the ascending colon of the 4 of the patients with an O-157 infection. In all patients with O-157 colitis, slight ascites was noted in the pelvic space. In additions, ascites was also seen in 3/13 patients with Salmonella and 1/5 patients with Campylobacter colitis. The CT findings, in the patients with infectious colitis, are non-specific but knowledge and recognition of the findings will help in patient evaluation and proper treatment.

Adult↗