• Radiation Oncology Journal
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• v.19 no.4
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• pp.389-396
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• 2001

Purpose : To detect differentially expressed genes in the patients with uterine cervical cancer during the radiation therapy. Materials and Methods : In patients with biopsy proven uterine cervical cancer, we took tumor tissue just before radiation therapy and at 40 minutes after external irradiation of 1.8 Gy. Total RNAs isolated from non-irradiated and irradiated tumor tissue samples were analyzed using the differential-display reverse transcription-polymerase chain reaction (DDRT-PCR). Complementary DNA (cDNA) fragments corresponding to differentially expressed messenger RNAs(mRNAs) were eluted, and cloned. The differential expression of the corresponding mRNAs was confirmed by reverse northern blot. Differentially expressed cDNA bands were sequenced. Nucleotide sequence data were analyzed in the Gene Bank and EMBL databases via the BLAST network sewer to identify homologies to known genes or cDNA fragments. Expression pattern of down-regulated clone was examined using RT-PCR in S patients undergoing radiotherapy. Results : We identified 18 differentially expressed bands by DDRT-PCR, which were eluted and cloned. There were 10 up-regulated clones and 1 down-regulated clone in reverse northern blot. One cDNA fragment had homology to chemokine receptor CXCR4, four were identified as Human ESTs in the EMBL database in EST clones. Down-regulated CxCa-11 was also down regulated in all patients. Conclusion : Using the DDRT-PCR, we have identified 10 up-regulated and 1 down-regulated clone(s) in the patients with uterine cervical cancer during the radiation therapy. The clinical relevance and the functions of these genes will be further investigated.

• Journal of Life Science
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• v.20 no.12
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• pp.1743-1749
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• 2010

A research group demonstrated that the 37 kDA protein of Edwardsiella tarda, a causing causative agent of edwardsiellosis in fish, exhibited high antigenicity in Japanese flounder. The research group also showed that the N-terminus amino acid sequences of the 37 kDa protein were mapped to the N-terminus of GAPDH (glyceraldehyde-3-phosphate dehydrogenase). Using degenerated primer sets based on the known N-terminus sequence, the corresponding E. tarda DNA was amplified and cloned. The nucleotide sequences of the cloned gene revealed high homology with a bacterial gene for GAPDH, as we was expected. The amino acid sequence of E. tarda GAPDH (etGAPDH) revealed a <70% similarity with GAPDH proteins in other Enterobacteriaceae. With the application of artificial protein overexpression system in Escherichia coli, the recombinant etGAPDH (rGAPDH) was produced and purified. In this study, Using the purified rGAPDH, the etGAPDH specific polyclonal antibody has been was generated using the purified rGAPDHin this study. The immunoblotting analyses demonstrated that the location of the GAPDH protein is located with the association of is associated with the envelops of E. tarda. The rGAPDH was administrated into Japanese flounder via IP route for evaluation of the protective ability. Although the specific antibody titer against etGAPDH was increased about 3-fold after 4 weeks post-vaccination, the survival rates of vaccinated Japanese flounder and the control group with wild type E. tarda was were 12.5% and 0%, respectively. Our results indicated that rGAPDH is immunoreactive antigen but that it will not generate protective immunity in Japanese flounder.

• Journal of Plant Biotechnology
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• v.2 no.2
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• pp.61-71
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• 2000

By screening a cDNA library of auxin-treated mung bean (Vigna radiata L.) hypocotyls, we have isolated two full-length cDNA clones, pVR-ACS6 and pVR-ACS7, for 1-aminocyclopropane-1-carboxylate (ACC) synthase, the rate-limiting enzyme in the ethylene biosynthetic pathway. While PVR-ACS6 corresponds to the previously identified PCR fragment pMBA1, pVR-ACS7 is a new cDNA clone. A comparison of deduced amino acid sequences among auxin-induced ACC synthases reveal that these enzymes share a high degree of homology (65-75%) to VR-ACS6 and VR-ACS7 polypeptides, but only about 50% to VR-ACS1 polypeptide. ACS6 and ACS7 are specifically induced by auxin, while ACS1 is induced by cycloheximide, and to lesser extent by excision and auxin treatment. Results from nuclear run-on transcription assay and RNA gel blot studies revealed that all three genes were transcriptionally active displaying unique patterns of induction by IAA and various hormones in etiolated hypocotyls. Particularly, 24-epibrassinolide (BR), an active brassinosteroid, specifically enhanced the expression of VR-ACS7 by distinct temporal induction mechanism compared to that of IAA. In addition, BR synergistically increased the IAA-induced VR-ACS6 and VR-ACS7 transcript levels, while it effectively abolished both the IAA- and kinetin-induced accumulation of VR-ACS1 mRNA. In light-grown plants, VR-ACS1 was induced by IAA in roots, whereas W-ACS6 in epicotyls. IAA- and BR-treatments were not able to increase the VR-ACS7 transcript in the light-grown tissues. These results indicate that the expression of ACC synthase multigene family is regulated by complex hormonal and developmental networks in a gene- and tissue-specific manner in mung bean plants. The VR-ACS7 gene was isolated, and chimeric fusion between the 2.4 kb 5'-upstream region and the $\beta$-glucuronidase (GUS) reporter gene was constructed and introduced into Nicotiana tobacum. Analysis of transgenic tobacco plants revealed the VR-ACS7 promoter-driven GUS activity at a highly localized region of the hypocotyl-root junction of control seedlings, while a marked induction of GUS activity was detected only in the hypocotyl region of the IAA-treated transgenic seedlings where rapid cell elongation occurs. Although there was a modest synergistic effect of BR on the IAA-induced GUS activity, BR alone failed to increase the GUS activity, suggesting that induction of VR-ACS7 occurs via separate signaling pathways in response to IAA and BR.

• The Korean Journal of Mycology
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• v.29 no.1
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• pp.34-40
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• 2001

RAPD test and the observation of morphological, cultural characteristics of fourteen selected entomogenous fungi were conducted to investigate the analysis of their internal relationships. Paecilomyces tenuipes showed snow flower form attached to numerous white conidiophores, produced globular and semi-egg types on the club types of phialides. Cordyceps militaris formed globosely conidiophores, dark yellow fruiting body on pupa. The phialide as on Acremonium-type in global conidiophores. Beauveria bassiana covered with conidia was not formed fruiting body and adhered conidia on conidiophore of zigzag type. The PDA and SDAY medium were confirmed as an optimum growth of them. P. tenuipes showed to velvet and plane types in several media whereas C. militaris was belong to centrally raised and floccose in the morphological type. In contrast, B. bassiana covered with conidia on velvet shape. The size of amplified products were analyzed by RAPD using URP primer and were from 100 bp to 2.0 kb with $10{\sim}14$ geuomic DNA. Total similarities of two groups were by dendrogram of UPGMA analysis. The homology of P. tenuipes groups was 94.8 to 100%. It also showed 70.1 to 96.6% in C. militaris group and B. bassiana was higher similarity than any other. The internal change of C. militaris, produced telemorph fruiting body, was higher seperated in species than P. tenuipes and B. bassiana in the RAPD.

• The Korean Journal of Mycology
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• v.27 no.1 s.88
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• pp.27-31
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• 1999

The internal transcribed spacer II regions (ITS II) of the ribosomal DNA gene repeat from Coprinus spp. were amplified using polymerase chain reaction (PCR) and sequenced. Sequences from 11 species including Coprinus comatus, C. atramentarius, C. micaceus, C. lagopus, C. cinereus, C. rhizophorus, C. flocculosus, C. radians, and C. echinosporus were compared. The spacer regions of them were $253{\sim}275$ nucleotide in length and partially contained 5.8S and 25S. The reciprocal homologies of each ITS II sequence among these strains were in the range of $50.6{\sim}100%$. According to the analysis of ITS II sequences, Coprinus spp. were classified into three clusters. Cluster I consisted of Coprinus lagopus, C. cinereus, C. echinosporus, C. rhizophorus, C. niveus, and C. atramentarius. Cluster II comprised C. micaceus, C. flocculosus, C. radians, and C. disseminatus. On the other hand C. comatus is in Cluster III even though this species is belonging to the section Coprinus in morphological aspect. These results suggest that Coprinus comatus, which was considered as a type species of the genus Coprinus in morphological classification, gives a doubt of monophyletic evolution and is assumed to be paraphyletic or polyphyletic.

• BMB Reports
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• v.38 no.6
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• pp.739-747
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• 2005

Full-length cDNA sequences of four novel SPATA4 genes in chimpanzee, cow, chicken and ascidian were identified by bioinformatic analysis using mouse or human SPATA4 cDNA fragment as electronic probe. All these genes have 6 exons and have similar protein molecular weight and do not localize in sex chromosome. The mouse SPATA4 sequence is identified as significantly changed in cryptorchidism, which shares no significant homology with any known protein in swissprot databases except for the homologous genes in various vertebrates. Our searching results showed that all SPATA4 proteins have a putative conserved domain DUF1042. The percentages of putative SPATA4 protein sequence identity ranging from 30% to 99%. The high similarity was also found in 1 kb promoter regions of human, mouse and rat SPATA4 gene. The similarities of the sequences upstream of SPATA4 promoter also have a high proportion. The results of searching SymAtlas (http://symatlas.gnf.org/SymAtlas/) showed that human SPATA4 has a high expression in testis, especially in testis interstitial, leydig cell, seminiferous tubule and germ cell. Mouse SPATA4 was observed exclusively in adult mouse testis and almost no signal was detected in other tissues. The pI values of the protein are negative, ranging from 9.44 to 10.15. The subcellular location of the protein is usually in the nucleus. And the signal peptide possibilities for SPATA4 are always zero. Using the SNPs data in NCBI, we found 33 SNPs in human SPATA4 gene genomic DNA region, with the distribution of 29 SNPs in the introns. CpG island searching gives the data about CpG island, which shows that the regions of the CpG island have a high similarity with each other, though the length of the CpG island is different from each other.This research is a fundamental work in the fields of the bioinformational analysis, and also put forward a new way for the bioinformatic analysis of other genes.

• The Korean Journal of Pesticide Science
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• v.10 no.1
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• pp.56-65
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• 2006

An antagonistic bacterial isolate, that inhibits the growth of plant pathogens, was selected and identified from 5,000 isolates screened from the rhizosphere of various crop plants. An isolate Bacillus sp. N32, tested against Colletotrichum gloeosporioides causing anthracnose disease in hot pepper, produced both a heat resistant antifungal protein and a heat sensitive antifungal protein. The heat resistant protein was partially purified by Ammonium sulfate fractionation and gel filtration chromatography. The bioautography showed that the proteins possessed high antifungal activity. The biosynthetic gene cluster responsible for the heat resistant antifungal protein was cloned from cosmid library using DNA probe obtained from PCR product with the primers targeting the conserved nucleotide sequence of the synthetic genes reported earlier, Most of the clones obtained showed higher homology to fengycin antibiotic synthetic gene family reported earlier. On the other hand, the heat sensitive protein was isolated from SDS-PAGE and electroblotting to determine the N-terminal amino acid sequences. The heat sensitive antifungal protein gene was cloned from the ${\lambda}-ZAP$ libraries using a DNA probe based on the N-terminal amino acid sequences of the heat sensitive protein. We are contemplating to clone and sequence the whole gene cluster encoding the heat sensitive protein for further analysis.

• BMB Reports
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• v.39 no.5
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• pp.502-510
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• 2006

2C-methyl-D-erythritol 2, 4-cyclodiphosphate synthase (MECPS, EC: 4.6.1.12) is the fifth enzyme of the non-mevalonate terpenoid pathway for isopentenyl diphosphate biosynthesis and is involved in the methylerythritol phosphate (MEP) pathway for ginkgolide biosynthesis. The full-length mecps cDNA sequence (designated as Gbmecps) was cloned and characterized for the first time from gymnosperm plant species, Ginkgo biloba, using RACE (rapid amplification of cDNA ends) technique. The full-length cDNA of Gbmecps was 874 bp containing a 720 bp open reading frame (ORF) encoding a peptide of 239 amino acids with a calculated molecular mass of 26.03 kDa and an isoelectric point of 8.83. Comparative and bioinformatic analyses revealed that GbMECPS showed extensive homology with MECPSs from other species and contained conserved residues owned by the MECPS protein family. Phylogenetic analysis indicated that GbMECPS was more ancient than other plant MECPSs. Tissue expression pattern analysis indicated that GbMECPS expressed the highest in roots, followed by in leaves, and the lowest in seeds. The color complementation assay indicated that GbMECPS could accelerate the accumulation of $\beta$-carotene. The cloning, characterization and functional analysis of GbMECPS will be helpful to understand more about the role of MECPS involved in the ginkgolides biosynthesis at the molecular level.

• Korean Journal of Ecology and Environment
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• v.53 no.3
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• pp.286-294
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• 2020

The Chironomus(Diptera: Chironomidae) is a freshwater benthic invertebrate that is an important indicator organism used for environmental pollution and water quality monitoring. In this study, we performed morphological classification and genetic species identification using the cytochrome c oxidase subunit I (COI) gene on mitochondrial DNA for an accurate species classification of Chironomus larvae found in tap water purification plants in Incheon, Korea. Twenty larvae in six water purification plants consist of four species, including twelve Chironomus kiiensis, six Chironomus flaviplumus, one Chironomus dorsalis, and one Polypedilum yongsanensis (not included Genus Chironomus). Morphological characteristics of each larvae were identified based on the head capsule, the mentum, the mandible, the antenna, and the claw. Based on the COI sequences of 21 individuals of 17 Chironomus species registered in NCBI Genbank, phylogenetic analysis indicated that the 20 individuals investigated in this study consist of the same clade with corresponding species of the high homology (99~100%) including C. kiiensis, C. flaviplumus, C. dorsalis, and P. yongsanensis. These results will be used as main classification indicator for monitoring freshwater ecosystems by providing integrated morphological and genetic information for the species identification of Korean Chironomus.

• Korean Journal of Environmental Biology
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• v.21 no.2
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• pp.194-202
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• 2003

To isolate alga-lytic bacteria, a number of samples were collected from Lake of Sukchon and Pal'tang reservoir where cyanobacteria blooming occurred. HY0210-AK1, which exhibited high alga-lytic activity, was isolated using Anabaena cylindrica lawn. The morphological and biochemical characteristics of the isolate HY0210-AK1 were very similar to that of the genus Rhizobium. Taxonomic identification including 16S rDNA base sequencing and phylogenetic analysis indicated that the isolate Hy0210-AK1 had a 99.1% homology in its 16S rDNA babe sequence with Sphingobium herbicidovorans. A. cylindrica NIES-19 was susceptible to the alga-lytic bacterial attack. The growth-inhibiting offset of the bacterium was not different on A. cylindrica NIES-19 when Sphingobium herbicidovorans HY0210-AK1 was in the lag, exponential, and stationary growth phase, although the alga-Iytic effect of S. herbici-dovorans HY0210-AK1 that in stationary growth phase was somewhat pronounced at the first time of inoculation. When S. herbicidovorans HY0210-AK1 was inoculated was inoculated with $1\times 10^{8}$ CFU $ml^{-1}$ together with A cylindrica NIES-19, the bacterium proliferated and caused algal lysis. A. cylindrica NIES-19 died when S. herbicidovorans HY0210 AKl was added to the algal culture but not when duly the filtrates from the bacterial culture was added. This suggests that extracellular substances are not responsible for inhibition of A. cylindrica NIES-19 and that algal Iysis largely attributed to direct interaction between S. herbicidovorans HY0210-AK1 and A. cylindrica NIES-19. The alga-lytic bacterium HY0210-AK1 caused cell lysis and death of three strain of Micro-cystis aeruginosa, but revealed no alga-Iytic effects on the Stephanodiscus hantzschii.