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Biomedical subjects

R Terauchi

Publications and source records attributed to R Terauchi.

17 recordsLinked to original sources
PubMed2001

Production of antimicrobial defensin in Nicotiana benthamiana with a potato virus X vector.

A recombinant plasmid, pTXS.TH, was constructed to express the gene-encoding wasabi (Wasabia japonica) defensin with the potato virus X (PVX) vector. pTXS.TH allows the expression of defensin in the host Nicotiana benthamiana, and the defensin protein WT1 can be purified from virus-infected leaves by heat treatment and affinity chromatography. WT1 exhibits strong antifungal activity toward the phytopathogenic fungi Magnaporthe grisea (50% inhibitory concentration [IC50] = 5 microg/ml) and Botrytis cinerea (IC50 = 20 microg/ml) but is weakly active against the phytopathogenic bacterium Pseudomonas cichorii. This virus-mediated expression system is a rapid and efficient method to produce and characterize antimicrobial proteins in plants. It is particularly useful for the study of proteins that are difficult to produce with other expression systems.

Anti-Infective Agents↗
PubMed2000

Requirement of regulated activation of Ras for response of MDCK cells to hepatocyte growth factor/scatter factor.

Hepatocyte growth factor/scatter factor (HGF/SF) induces cell scattering, migration, and branching tubule formation of MDCK cells. To examine the role of the Ras protein in the HGF/SF-induced responses, we constructed MDCK cell clones expressing either inducible dominant-negative Ras or constitutively activated Ras and analyzed their effects on responses of cells to HGF/SF. Induced expression of dominant-negative Ras prevented cell dissociation required for cell scattering, migration, and cystic formation as well as branching morphology required for branching tubule formation. Constitutively activated Ras induced cell dissociation, but not a scattered fibroblastic morphology even in the presence of HGF/SF. MDCK cells expressing constitutively activated Ras migrated at a level similar to that of wild-type MDCK cells stimulated by HGF/SF. MDCK cells expressing constitutively activated Ras showed disorganized growth in three-dimensional culture and did not form the branching tubule structures. These results indicate that activation of the Ras protein is essential for the cell scattering, migration, and branching tubule formation of MDCK cells induced by HGF/SF, and a properly regulated activation is required for some stages of the HGF/SF-induced responses of MDCK cells.

Animals↗
PubMed2000

Rapid isolation of promoter sequences by TAIL-PCR: the 5'-flanking regions of Pal and Pgi genes from yams (Dioscorea).

Using a modified TAIL-PCR technique, the 5'-flanking regions of the phenylalanine ammonia lyase (Pal) genes of a yam species, Dioscorea bulbifera, and the phosphoglucose isomerase (Pgi) gene of D. tokoro were successfully isolated. Two novel modifications of the TAIL-PCR procedure introduced here, namely (1) the use of a battery of random 10-mers (RAPD primers) as short arbitrary primers, and (2) the use of a total of five nested, gene-specific primers, allow the rapid isolation of the 5'-flanking region of any gene from organisms with large genomes. Isolated 5'-flanking regions were fused to the gus gene, and tested for transient expression in tobacco BY2 cells. All the isolated 5'-flanking regions were shown to drive reporter gene expression. Three Pal promoters responded to salicylic acid, presumably as a result of the binding of a MYB transcriptional activator to the multiple MREs (Myb Recognition Elements) present in these regions.

Base Sequence↗
PubMed2000

Radial magnetic resonance imaging and pathological findings of acetabular labrum in dysplastic hips.

Radial MRI findings and pathological changes were comparatively examined in the acetabular labrum of 11 hips of 11 patients, who underwent total hip arthroplasty for osteoarthritis due to acetabular dysplasia. Diffuse high signal pattern on the radial MR images corresponded to histological degeneration of the labrum. High signal pattern which was equivalent to the synovial fluid, showed an intralabral tear. In the obscure areas of MR images, severe impairment of the labrum such as rupture, detachment, and displacement were found.

Journal Article↗
PubMed2000

cDNA cloning and molecular analysis of papaya small GTP-binding protein, pgp1.

In the course of papaya EST collection, one clone (pRA4-3) encoding partial sequence of papaya small GTP-binding protein gene, pgp1, was obtained. Based on the sequence information of pRA4-3, the entire coding region of pgp1 was cloned using the 3'RACE PCR technique. ORF of pgp1 is 636bp long and deduced molecular weight of the protein is 23,311. Phylogenetic analysis showed that PGP1 belongs to YPT/RAB group of the small GTP-binding protein and is a homologue of RAB2. Southern analysis showed that there are several pgp1-related genes in papaya genome. Northern analysis showed that pgp1 was expressed equally in stems of seedlings that were grown under light and dark conditions. This result shows that PGP1 is not involved in the phytochrome-mediated signal transduction as an auxin signal transducer in stems of papaya seedlings.

Amino Acid Sequence↗
PubMed1999

Technical advance: transcript profiling in rice (Oryza sativa L.) seedlings using serial analysis of gene expression (SAGE)

Serial analysis of gene expression (SAGE) was applied for profiling expressed genes in rice seedlings. In the SAGE method, a 9-11 bp fragment (tag) represents each transcript, and frequency of a tag in the sample directly reflects the abundance of the respective mRNA. We studied 10 122 tags derived from 5921 expressed genes in rice (Oryza sativa L.) seedlings, among which only 1367 genes (23.1%) matched the rice cDNA or EST sequences in the DNA database. SAGE showed that most of the highly expressed genes in rice seedlings belong to the category of housekeeping genes (genes encoding ribosomal proteins or proteins responsible for metabolism and cell structure). Unexpectedly, the most highly expressed gene in rice seedlings was a metallothionein (MT) gene, and together with three other messages for MT, it accounts for 2.7% of total gene expression. To our knowledge, this is the first quantitative study of global gene expression in a higher plant. We further applied the SAGE technique to identify differentially expressed genes between anaerobically treated and untreated rice seedlings. Additionally, we show that a longer cDNA fragment can be easily recovered by PCR using the SAGE tag sequence as a primer, thereby facilitating the analysis of unknown genes identified by tag sequence in SAGE. In combination with micro-array analysis, SAGE should serve as a highly efficient tool for the identification and isolation of differentially expressed genes in plants.

Journal Article↗
PubMed1999

A method for estimating nucleotide diversity from AFLP data.

A method for estimating the nucleotide diversity from AFLP data is developed by using the relationship between the number of nucleotide changes and the proportion of shared bands. The estimation equation is based on the assumption that GC-content is 0.5. Computer simulations, however, show that this method gives a reasonably accurate estimate even when GC-content deviates from 0.5, as long as the number of nucleotide changes per site (nucleotide diversity) is small. As an example, the nucleotide diversity of the wild yam, Dioscorea tokoro, was estimated. The estimated nucleotide diversity is 0.0055, which is larger than estimations from nucleotide sequence data for Adh and Pgi.

DNA Fingerprinting↗
PubMed1997

Microsatellite polymorphism in natural populations of the wild plant Arabidopsis thaliana.

Variation in repeat number at 20 microsatellite loci of Arabidopsis thaliana was studied in a worldwide sample of 42 ecotypes to investigate the pattern and level of polymorphism in repetitive sequences in natural plant populations. There is a substantial amount of variation at microsatellite loci despite the selfing nature of this plant species. The average gene diversity was 0.794 and the average number of alleles per locus was 10.6. The distribution of alleles was centered around the mean of repeat number at most loci, but could not be regarded as normal. There was a significantly positive correlation between the number of repeats and the amount of variation. For most loci, the observed number of alleles was between the expected values of the infinite allele and stepwise mutation models. The two models were rejected by the sign test. Linkage disequilibrium was detected in 12.1% of the pairwise comparisons between loci. In phylogenetic tree, there was no association between ecotype and geographic origin. This result is consistent with the recent expansion of A. thaliana throughout the world.

Alleles↗
PubMed1997

DNA polymorphism at the Pgi locus of a wild yam, Dioscorea tokoro.

To study the origin and maintenance mechanisms of the PGI allozyme polymorphism of a wild plant, Dioscorea tokoro, DNA sequences of the entire coding region (1701 bp) and two intronic regions (total 2049 bp) of the Pgi gene as well as a part of the Adh gene (590 bp) were analyzed. Two replacement substitutions were revealed to be responsible for the differentiation of three allozymes alleles (Pgi-a, Pgi-b and Pgi-c) that occur in natural population in intermediate frequencies. Interspecific comparison of DNA sequences identified Pgi-b as the oldest allele, from which two other alleles were derived probably within the last 150,000 years. The level of DNA polymorphism at D. tokoro Pgi locus was low. No elevated level of DNA polymorphism was detected in the close vicinity of the two replacement sites differentiating the three allozymes. Departures from the neutral mutation hypothesis were detected by Fu and Li's and MK tests. The observed patterns of DNA polymorphism are explainable by both (1) the neutral mutation hypothesis with an assumption of small effective size of D. tokoro population, and (2) the positive selection hypothesis that the allele frequencies of Pgi-a and Pgi-c have increased in a short time by their selective advantages.

Alcohol Dehydrogenase↗
PubMed1997

Nucleotide polymorphism in the acidic chitinase locus (ChiA) region of the wild plant Arabidopsis thaliana.

To investigate DNA variation in natural plant populations, a 1.8-kb region of the acidic chitinase locus (ChiA)was analyzed for 17 ecotypes of Arabidopsis thaliana sampled worldwide and 3 Arabis species in Japan. As in the Adh region, dimorphism was detected throughout the investigated ChiA region, suggesting the possibility that dimorphic DNA variation exists in the entire nuclear genome of A. thaliana. The ChiA region was divided into two blocks by an intragenic recombination between two parental sequence types, which diverged 7.4 MYA under the assumption that nucleotide mutation rate per site per year is mu = 10(-9). Nucleotide diversity in the entire ChiA region was 0.0104. Tajima's test was significantly negative for both nucleotide and indel variations, which was manifested as an excess of unique polymorphisms. However, the level and pattern of polymorphism in the ChiA region were inconsistent with simple theoretical explanations. The HKA test detected no difference in the levels of intra- and interspecific variations between the ChiA and Adh regions. In the ChiA coding region, no difference in the patterns of synonymous and replacement variation was found in intra- and interspecific comparisons by the MK test. Although it was difficult to determine the exact genetic mechanism acting on the ChA locus, these results suggested that the ChA locus region was under the same genetic mechanism before and after the establishment of A. thaliana as a species.

Alcohol Dehydrogenase↗
PubMed1996

Intragenic recombination in the Adh locus of the wild plant Arabidopsis thaliana.

Nucleotide variation in the Adh region of the wild plant Arabidopsis thaliana was analyzed in 17 ecotypes sampled worldwide to investigate DNA polymorphism in natural plant populations. The investigated 2.4-kb Adh region was divided into four blocks by intragenic recombinations between two parental sequence types that diverged 6.3 million years (Myr) ago, if the nucleotide mutation rate mu = 10(-9) is assumed. Within each block, dimorphism of segregating variations was observed with intermediate frequencies, which caused a substantial amount of nucleotide variation in A. thaliana at the species level. The first recombination introduced the divergent variation that resulted in dimorphism in this plant species approximately 3.3 Myr ago, and three subsequent intragenic combinations have occurred sporadically in approximately 1.1-Myr intervals. It was shown that there was only a limited number (six) of sequence types in this species and that no clear association was observed between sequence type and geographic origin. Taken together, these results suggest that A. thaliana has spread over the world only recently. It can be concluded that recombination played an important role in the evolutionary history of A. thaliana, especially through the generation of DNA polymorphism in the natural populations of this plant species.

Alcohol Dehydrogenase↗
PubMed1995

Molecular systematics of the Trilliaceae sensu lato as inferred from rbcL sequence data.

In this paper, we examine the phylogenetic affinities of Trilliaceae sensu lato. DNA sequences of the rbcL gene were analyzed for 8 species from Trilliaceae sensu lato (Trillium kamtschaticum, T. sessile, Daiswa polyphylla, Kinugasa japonica, Paris tetraphylla, Scoliopus bigelovii, S. hallii, and Medeola virginiana) and 10 other species from Liliaceae sensu lato. The analysis indicates that Trilliaceae sensu stricto (Trillium, Daiswa, Kinugasa, and Paris) is monophyletic and that Trilliaceae sensu lato is not monophyletic. Scoliopus and Medeola do not form a clade with Trilliaceae sensu stricto. Instead, they form a clade with Uvulariaceae and Liliaceae sensu stricto, respectively. Relative to the rbcL genes of Trilliaceae sensu stricto, Scoliopus and Medeola displayed approximately 14-24 synonymous substitutions per 100 sites. In similar comparisons with Streptopus and Clintonia (Uvulariaceae), these displayed approximately 1-4 synonymous substitutions per 100 sites. The rbcL gene analysis supports the previous suggestion that Scoliopus and Medeola should be moved from Trilliaceae to Liliaceae sensu stricto. The analysis also indicated that Veratrum (Melanthiaceae) may be related to Trilliaceae sensu stricto. For reconstructing the phylogeny of lower taxonomic level as such, it was found that data on nonsynonymous base substitution as well as weighting in the first and second positions of codons are not useful.

Base Sequence↗
PubMed1995

DNA fingerprinting study on the intraspecific variation and the origin of Prunus yedoensis (Someiyoshino).

In order to investigate the intraspecific variation of Prunus yedoensis (Someiyoshino) and interspecific relationship among P. yedoensis, P. lannesiana (Oshimazakura) and P. pendula (Edohigan), DNA fingerprinting study was conducted by using two different kinds of probes, M13 repeat sequence and (GACA)4 synthetic oligonucleotide. In this study, 68 plants of P. yedoensis grown in 46 prefectures in Japan were investigated. All the P. yedoensis individuals investigated showed the completely same banding pattern, indicating their clonal origin from a single plant. On the other hand, each of P. lannesiana and P. pendula individuals investigated showed a unique banding pattern, suggesting a considerable amount of genetic variation in these two species. About 90% of bands in DNA fingerprints of P. yedoensis were detected in either P. lannesiana or P. pendula. This result supports the hypothesis that P. yedoensis is an interspecific hybrid between P. lannesiana and P. pendula. From those results, it is concluded that P. yedoensis was produced only once through hybridization between P. lannesiana and P. pendula, and that this particular hybrid plant has been spread vegetatively all over Japan.

Base Sequence↗
PubMed1994

Microsatellite polymorphism in Dioscorea tokoro, a wild yam species.

Six microsatellite loci were characterized in Dioscorea tokoro, a wild yam species in East Asia. All six loci were polymorphic in a sample of 23 individuals from natural populations in Japan. The microsatellite loci displayed many alleles (6.2 alleles per locus on average), and the observed heterozygosity (Ho = 0.54) as well as expected heterozygosity (He = 0.68) were high. The heterozygosities were far more than that previously detected by allozyme analysis of D. tokoro (Ho = 0.23, He = 0.28). Five microsatellite loci were sufficient to provide a paternity exclusion rate (Q) of Q = 0.98, which enables monitoring of the pollen-mediated gene flow between plants in a population. Microsatellite loci are abundant and highly polymorphic in D. tokoro and other plants and are therefore ideal markers for plant population genetic studies.

Alleles↗
PubMed1994

A polymorphic microsatellite marker from the tropical tree Dryobalanops lanceolata (Dipterocarpaceae).

Di-nucleotide microsatellites were isolated from a genomic library of a tropical tree species, Dryobalanops lanceolata, in Sarawak, for the purpose of using them as hypervariable genetic markers to study the pollen-mediated gene flow. Among 1600 recombinant clones, in total 20 clones gave positive signals when hybridized with oligonucleotides with the three different repeat motifs, GT, CA and CT. Estimations of abundance of (GT)n/(CA)n and (GA)n/(CT)n dinucleotide repeats in D. lanceolata genome revealed to be one in every 84 kb and 80 kb, respectively. Among six sequenced microsatellite loci, one was selected to synthesize PCR primers to amplify the microsatellite. PCR product size of the locus was variable among different individuals, which is attributed to the different number of di-nucleotide repeats. The same microsatellite genotype was detected in the trunk and canopy of a single large tree, indicating the utility of trunk tissue as the source of DNA for the population genetic study of tropical tree species, the canopy of which is usually difficult to approach.

Base Sequence↗