• The Plant Pathology Journal
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• v.20 no.3
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• pp.229-233
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• 2004

The bacterial leaf blight, which is caused by Xantho-monas oryzae pv. oryzae, is the most damaging and intractable disease of rice. To identify the genes involved in the virulence mechanism of transposon TnS complex, which possesses a linearized transposon and transposase, was successfully introduced into X. oryzae pv. oryzae by electroporation. The transposon mutants were selected and confirm the presence of transposition in X. oryzae pv. oryzae by the PCR amplification of transposon fragments and the Southern hybridization using these mutants. Furthermore, transposon insertion sites in the mutant bacterial chromosome were deter-mined by direct genomic DNA sequencing using transposon-specific primers with ABI 3100 Genetic Analyzer. Efficiency of transposition was influenced mostly by the competence status of X. oryzae pv. oryzae cells and the conditions of electroporation. These results indicated that the insertion mutagenesis strategy could be applied to define function of uncharacterized genes in X. oryzae pv. oryzae.

• Asian-Australasian Journal of Animal Sciences
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• v.16 no.7
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• pp.1066-1070
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• 2003

In order to identify differentially expressed mRNAs (which represent possible candidates for significant phenotypic variances of muscle growth, meat quality between introduced European and Chinese indigenous pigs) in the longissimus dorsi muscle tissue between adult Duroc and Erhualian pigs, mRNA differential display was performed. Five 3' anchor primers in combination with 20 different 5' arbitrary primers (100 primer sets) were used and nearly 5,000 cDNA bands were examined, among which 10 differential display cDNAs were obtained, cloned and sequenced. Six of the 10 cDNAs showed similarity to identified genes from GenBank and the other 4 had no matches in GenBank. Differential expression was tested by Northern blot hybridization and could be confirmed for 2 cDNAs. The method used in this study provides a useful molecular tool to investigate genetic variation that occurs at the transcriptional level between different breeds.

• Biotechnology and Bioprocess Engineering:BBE
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• v.10 no.5
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• pp.432-443
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• 2005

Microarray technology has contributed Significantly to the understanding of bacterial genetics and transcriptional regulation. One neglected aspect of this technology has been optimization of microarray-generated signals and quality of generated information. Full genome microarrays were developed for Clostridium acetobutylicum through spotting of PCR products that were designed with minimal homology with all other genes within the genome. Using statistical analyses it is demonstrated that Signal quality is significantly improved by increasing the hybridization volume. possibly increasing the effective number of transcripts available to bind to a given spot, while changes in labeled probe amounts were found to be less sensitive to improving signal quality. In addition to Q-RT-PCR, array validation was tested by examining the transcriptional program of a mutant (M5) strain lacking the pSOL1 178-gene megaplasmid relative to the wildtype (WT) strain. Under optimal conditions, it is demonstrated that the fraction of false positive genes is 1% when considering differentially expressed genes and 7% when considering all genes with signal above background. To enhance genomic-scale understanding of organismal physiology, using data from these microarrays we estimated that $40{\sim}55%$ of the C. acetobutylicum genome is expressed at any time during batch culture, similar to estimates made for Bacillus subtilis.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2001.10a
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• pp.18-24
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• 2001

To identify genes implicated in the control of pluripotency as well as characteristics of stem cells, we analyzed expression profiles of genes derived from mouse morulas, blastocysts, embryonic stem cells, mesenchymal stem cells, and uterus tissue cDNA microarray. Comparative analyses of their expression profiles identified putative clones that expressed specifically in specific samples or not in a specific sample. The expression pattern of these candidate clones was analyzed using RT-PCR and non-radioactive in situ hybridization. Functional annotation of these clones on pluripotency and stem cells and molecular mechanisms underlying many facets of mammalian development and differentiation.

• International Journal of Industrial Entomology and Biomaterials
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• v.2 no.1
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• pp.61-64
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• 2001

Ubiquitin can be covalently attached to cellular proteins as a post-translational modification rind is involved in metabolic stresses, such as bent shock and immune response. We have isolated and sequenced a cDNA encoding ubiquitin from the silkworm, Bombyx mori. The insert in the clone is 533 nucleotide long with an open reading frame of 387 nucleotides that encodes a protein of 129 amino acids with a molecular weight of 14.8 kDa. The amino acid sequence shared high homology with the ubiquitins known so far, The result of dot blot hybridization showed that the B. mori ubiquitin gene is up-regulated upon f. rofi infection, suggesting that the B. mori ubiquitin plays an immune-related role.

• Parasites, Hosts and Diseases
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• v.40 no.3
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• pp.131-138
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• 2002

The cyclophilins (Cyps) are family members of proteins that exhibit peptidylprolyl cis-trans isomerase (PPIase, EC 5.2.1.8) activity and bind the immunosuppressive agent cyclosprin A (CsA) in varying degrees. During the process of random sequencing of a cDNA library made from Giardia intestinalis WB strain, the cyclophilin gene (gicypl) was isolated. An open reading frame of gicyp1 gene was 576 nucleotides, which corresponded to a translation product of 176 amino acids (Gicypl). The identity with other Cyps was about 58-71%. The 13 residues that constituted the CsA binding site of human cyclophilin were also detected in the amino acid sequence of Gicypl, including tryptophan residue essential for the drug binding. The single copy of the gicypl gene was detected in the G. intestinalis chromosome by southern hybridization analysis. Recombinant Gicyp 1 protein clearly accelerated the rate of cis ${\rightarrow}$ trans isomerization of the peptide substrate and the catalysis was completely inhibited by the addition of $0.5{\;}{\mu}M$ CsA.

• Journal of Plant Biotechnology
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• v.4 no.4
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• pp.155-161
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• 2002

The HCV gene was expressed in potato plants under the control of the constitutive CaMV 355 promoter or tuber-specific patatin promoter. Solanum tuberosum plants carrying a plant expression vector harboring the encoding region of HCV gene were generated by Agrobacterium tumefaciens-mediated in vitro transformation methods. The presence of HCV gene in the plant genome was detected by PCR and DNA hybridization experiments. We obtained the 5 lines of transgenic potato with the pMBPHCV construct and 4 lines of transgenic potato with the pATHCV construct. The HCV transgenic stably integrated into the potato genome, as well as their transcription. HCV mRNA was identified in leaf and tuber tissues of transgenic plants by Northern blot analysis. The transgenic potato plants produced the expected transcript, and the corresponding HCV protein accumulated in individual transgenic plants.

• Journal of Photoscience
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• v.12 no.2
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• pp.75-82
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• 2005

Rumex acetosa L. is a dioecious flowering plant with well developed sex chromosome system: 2n = 12 + XX in the female plants and 2n = 12 + XY1Y2 in the male plants. To isolate sex-linked DNA, we carried out chromosome micromanipulation, followed by DOP-PCR, AFLP of the PCR products, reverse Southern hybridization and sequence analysis. From 500 AFLP specific clones, 13 X-chromosome and 5 Y-chromosome specific clones were obtained. Except one clone RADAX-239 ($\underline{R}umex\;\underline{a}-\underline{D}OP-PCR-\underline{A}FLP-\underline{Y}-chromosome\;specific$), all clones appear to be R. acetosa plant-specific sequences and non-coding sequences. Southern blot analysis using these clones could not discriminate genomic DNAs either from male or female plants. Results of this study imply that both autosome-origin and degeneration of sex chromosomes are prevalent in plant systems.

• Korean Journal of Environmental Biology
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• v.21 no.1
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• pp.52-55
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• 2003

There is considerable evidence that ionizing radiation (IR) mediates checkpoint control, repair and cell death. In this study, we have used a high density microarray hybridization approach to characterize the transcriptional response of human breast carcinoma MCF-7 cell line to ${\gamma}$-radiation, such as 4 Gy 4 hr, 8 Gy 4 hr, and 8 Gy 12 hr. We found that exposure to ${\gamma}$-ray alters by at least a $log_2$ factor of 1.0 the expression of 115 known genes. Of the 66 genes affected by ${\gamma}$-radiation, 49 are down-regulated. In our results, the cellular response to irradiation includes induction of the c-jun and EGR1 early response genes. The present work has examined potential cytoplasmic signaling cascades that transduce IR-induced signals to the nucleus. 40S ribosomal protein s6 kinase modulates the activities of the mitogen activated protein kinase (MAPK) and c-Jun $NH_2$-terminal kinase (JNK1) cascades in human monocytic leukemia (U937/pREP4) cells. 14-3-3 family members are dimeric phosphoserine -binding proteins that participate in signal transduction and checkpoint control pathways.

• Microbiology and Biotechnology Letters
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• v.26 no.5
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• pp.387-392
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• 1998

Brevibacterium lactofermentum, a gram-positive bacteria, has both the diaminopimelate (DAP) pathway and meso-DAP-dehydrogenase (DDH) pathway for L-lysine biosynthesis. To investigate importance of DDH pathway and the related ddh gene in lysine production, we introduced site-specific mutagenesis technique. A 300 bp DNA fragment central to the meso-DAP-dehydrogenase gene (ddh) of B. lactofermentum was used to inactive chromosomal ddh gene via homologous recombination. Southern hybridization analysis confirmed that the chromosomal ddh gene was disrupted by the vector sequence. The B. lactofementum ddh mutant obtained have an inactive DDH pathway. The results reveal that inactivation of the ddh gene in B. lactofermentum leads to dramatic reduction of lysine production as well as decrease of the growth rate, indicating that the DDH pathway is essential for high-level lysine production as well as biosynthesis of meso-DAP.