• Title/Summary/Keyword: complementation

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Vibrio vulnificus Metalloprotease VvpE has no Direct Effect on Iron-uptake from Human Hemoglobin

  • Sun, Hui-Yu;Han, Song-Iy;Choi, Mi-Hwa;Kim, Seong-Jung;Kim, Choon-Mee;Shin, Sung-Heui
    • Journal of Microbiology
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    • v.44 no.5
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    • pp.537-547
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    • 2006
  • This study was designed to determine whether or not Vibrio vulnificus metalloprotease VvpE can promote iron uptake via the proteolytic cleavage of human hemoglobin. We found that V. vulnificus utilized hemoglobin as an iron source more efficiently via the vulnibactin-mediated iron-uptake system than via the HupA-mediated iron-uptake system and, of the proteases produced by V. vulnificus, VvpE was found to be the only protease capable of destroying hemoglobin. However, VvpE expression, on both the transcriptional and protein levels, was suppressed in iron-limited media. However, vvpE transcription, but not extracellular VvpE production, was reactivated by the addition of hemoglobin or inorganic iron into iron-limited media. Moreover, vvpE transcription began only in the late growth phase when V. vulnificus had already consumed most of the iron for growth. In addition, neither vvpE mutation nor in trans vvpE complementation affected the ability of V. vulnificus to acquire iron or to grow in iron-limited media or in cirrhotic ascites containing hemoglobin. Hemoglobin added into iron-limited media was not destroyed, but gradually formed an insoluble aggregate during culture; this aggregation of hemoglobin occurred regardless of vvpE mutation or complementation. These results indicate that VvpE is not required for efficient iron uptake from hemoglobin. On the contrary, hemoglobin or iron is required for efficient vvpE transcription. In addition, a discrepancy exists between vvpE transcription and extracellular VvpE production in iron-limited media containing inorganic iron or hemoglobin, which suggests that additional unknown posttranscriptional events may be involved in the extracellular production of VvpE.

Molecular Cloning and Characterization of Outer Envelope Membrane Protein from Salicornia herbacea (퉁퉁마디로부터 색소체 외막 단백질 유전자의 분리 및 발현분석)

  • Ermawati Netty;Cha, Joon-Yung;Liang, Yingshi;Jung, Min-Hee;Shin, Dongjin;Lee, Byung-Hyun;Lee, Kon-Ho;Son, Daeyoung
    • Journal of Plant Biotechnology
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    • v.31 no.4
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    • pp.273-278
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    • 2004
  • Complementary DNA encoding chloroplast outer envelope membrane protein (OEP) from the halophyte Salicornia herbacea has been cloned and sequenced. The full length cDNA is 596 bp and encodes a polypeptide of 91 amino acid residues with a molecular mass of 8.9 kDa. The expression level of ShOEP increased by salt, drought and ABA treatments. ShOEP expression was largely induced in roots and shoots by high salts. The biological function of ShOEP was examined by yeast complementation. ShOEP can suppress Na$^{+}$ sensitivity of yeast mutant (cnb$\Delta$) in the presence of salt. These results suggest that ShOEP is a salt inducible gene and may have functions in the regulation of plant salt stress.ant salt stress.

Cloning of Acetyl CoA Carboxylase (fabE) in Escherichia coli (대장균의 acetyl CoA carboxylase유전자의 클로닝)

  • Park, Wan;Song, Bang-Ho;Hong, Soon-Duk
    • Microbiology and Biotechnology Letters
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    • v.14 no.2
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    • pp.181-186
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    • 1986
  • A defective lambda transducing phase carrying acetyl CoA carboxylase gene (fabE) from Escherichia coli chromosome (72 min on the current linkage map) has been isolated. A restriction map of the chromosomal region from defective transducing phage was established by digestion with combination of the restriction enzymes. No cleavage site for the enzyme EcoRI was found in this region. Restriction fragments were cloned from defective transducing phage into high copy number plasmid vector pACYC184 to generate hybrid plasmids which were capable of complementation of fabE temperature sensitive mutation. We show here that the fabE gene is located on a 3.4 megadalton Bam HI-SalI fragment with a HindIII site, which lies within the 7.4 megadalton BglIIfragment, by complementation analysis.

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Understanding the Soteriology of the Film : Focusing on the 'The Human Complementation Project' (의 구원관 연구: '인류보완계획'을 중심으로)

  • Lee, Kil-Yong
    • Cartoon and Animation Studies
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    • s.19
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    • pp.1-16
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    • 2010
  • The purpose of this Paper is a studying the soteriology of the film 'The End of Evangelion'(1997). In many religions the human being is recognized as negative and defect existence, on this count human being needs the salvation. Therefor the word salvation implies the meaning of liberation or overcoming. On this point Christianity, Buddhism, and Islam are of one accord. The christianity sees the original sin as the human limits, and the buddhism points out the ignorance(無明, ajnana) of human being. The soteriology of the film 'The End of Evangelion' is similar to the above-mentioned religions. For that the terminology of 'the Human Complementation Project' is used in film. In many religions the salvation is fulfilled in this world, in other words 'now' and 'here' are emphasized fot the accomplishment of the salvation. In this point the film 'the End of Evangelion' stands on the same road with the other religions.

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Meta-analysis of Excision Repair Cross-complementation Group 1 (ERCC1) Association with Response to Platinum-based Chemotherapy in Ovarian Cancer

  • Li, Feng-Ying;Ren, Xiao-Bin;Xie, Xin-You;Zhang, Jun
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.12
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    • pp.7203-7206
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    • 2013
  • Recent studies suggested that the ovarian cancers with negative excision repair cross-complementation group 1 enzyme (ERCC1) expression have a better response to platinum-based chemotherapy than those with positive ERCC1 expression. The objective of this study was to evaluate whether ERCC1 expression is associated with response to platinum-based chemotherapy in ovarian cancers. MEDLINE, PubMed, Web of Science and CNKI databases were used for searching studies relating to ERCC1 protein expression and response to platinum-based chemotherapy in ovarian cancers. Statistical analysis was based on the method for a fixed effects meta-analysis. Pooled odds ratios (ORs) with 95% confidence intervals for ERCC1 protein expression and response to platinum-based chemotherapy were generated. Publication bias was investigated with Begg's test. Five studies involving 306 patients with ovarian cancer were included. Compared to patients with positive ERCC1 expression, those with negative ERCC1 expression had a better response to platinum-based chemotherapy. The pooled OR was 5.264 (95% CI: 2.928-9.464, P < 0.001) and publication bias was not found (P = 0.904). The result was similar in both in Asians and Caucasians (P < 0.001 and P = 0.028, respectively). ERCC1 protein expression status is significantly associated with response to platinum-based chemotherapy in ovarian cancers.

Elucidation of Function and Isolation of Trans-acting Factors Regulating the Basal Level Expression of Eukaryotic Genes (진핵세포 유전자의 기초대사 발현을 조절하는 trans 작용인자의 기능해석과 새로운 인자의 분리)

  • 황용일
    • Microbiology and Biotechnology Letters
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    • v.19 no.1
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    • pp.37-44
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    • 1991
  • - I aimed to isolate trans-acting factors involved in the basal expression level of eukaryotic genes. One of the yeast histidine biosynthetic gene, HIS5 was taken as a model for this study. HIS5 gene has a substantial basal level in amino acid rich medium and is derepressed if starved for any single amino acid. The derepression is mediated by cis-acting DNA sequences 5'-TGACTC-3' found in 5' non-transcribed region of the gene and trans-acting factors including GCN4 as positive factor and its negative factor GCDI 7, and GCNZ as a negative factor of GCD17. I first investigated the role of these trans-acting factors in HIS5 basal expression level by using HIS5-pH05 fusion in which expression of pH05 gene encoding inorganic phosphate-repressible acid phosphatase (APase) is regulated by HIS5 promoter. Strain with gcn2 or gcn4 mutation showed 3 to 4 fold lower APase activity than wild type. The level of APase activity was similar in gcn2 and gcn4 mutants. Trans-acting factors involved in basal level were identified by isolating 14 mutants showing increased expression of HISSPH05 fusion from gcn4 background. All the mutants carry a single nuclear recessive mutation and fall into four complementation groups, designated as bell (basal expression level), be12, be23 and be14.

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Antibiotic Biosynthesis in bldA-like Mutant of Strptomyces coelicolor (Streptomyces coelicolor blAA-like Mutant에서의 항생물질 생합성)

  • Park, Unn-Mee
    • Korean Journal of Microbiology
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    • v.32 no.1
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    • pp.70-77
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    • 1994
  • The author isolated 7 mutant candidates which mapped around cysA (which was 10 o'clock). They were divided into two groups. One of them was located counterclockwise to cysA, and the other was clockwise to cysA. Since bldA was mapped counterclockwise to cysA, the candidate which mapped counterclockwise to cysA was transduced with phage containing wild type bldA gene clone. The candidates might be the alleles of bldA, because they were complemented by bldA clone. However some of such mutants sporulated very well and developed as much pigment as wild type on rich media plate. Their phenotype was not like bld mutant at all on such conditions. There were real antibiotics gene expressions, since transcriptional reporter gene xylE had shown high activities. Majority of the bldA like mutants showed act gene expressions when they were transformed with high copy number plasmid containing actII-ORF4.

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Generation of Embryonic Stem Cell-derived Transgenic Mice by Using Tetraploid Complementation

  • Park, S.M.;Song, S.J.;Uhm, S.J.;Cho, S.G.;Park, S.P.;Lim, J.H.;Lee, H.T.
    • Asian-Australasian Journal of Animal Sciences
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    • v.17 no.12
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    • pp.1641-1646
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    • 2004
  • The objective of this study was to generate transgenic mice expressing human resistin gene by using the tetraploidembryonic stem (ES) cell complementation method. Human resistin gene was amplified from human fetal liver cDNA library by PCR, cloned into $pCR^{(R)}$ 2.1 $TOPO^{(R)}$ vector and constructed in pCMV-Tag4C vector. Mammalian expression plasmid containing human resistin was transfected into D3-GL ES cells by Lipofectamine 2,000, and then after 10-12 days of transfection, the human resistin-expressing cells were selected with G418. In order to produce tetraploid embryos, blastomeres of diploid embryos at the two-cell stage were fused with two times of electric pulse using 60 V 30 $\mu$sec (fusion rate: 2,114/2,256, 93.5%) and cultured up to the blastocyst stage (development rate: 1,862/2,114, 94.6%). The selected 15-20 ES cells were injected into tetraploid blastocysts, and then transferred into the uteri of E 2.5 d pseudopregnant recipient mice. To investigate the gestation progress, two E 19.5 mused fetuses were recovered by Cesarean section of which one fetus was confirmed to contain human resistin gene by genomic DNA-PCR. Therefore, our findings demonstrate that tetraploid-ES mouse technology can be considered as a useful tool to produce transgenic mice for the rapid analysis of gene function in vivo.

Predictive Value of Excision Repair Cross-complementing Rodent Repair Deficiency Complementation Group 1 and Ovarian Cancer Risk

  • He, Shan-Yang;Xu, Lin;Niu, Gang;Ke, Pei-Qi;Feng, Miao-Miao;Shen, Hong-Wei
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.5
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    • pp.1799-1802
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    • 2012
  • Objective: We aimed to analyze the association between excision repair cross-complementing rodent repair deficiency complementation group 1 (XRCC1) and ovarian cancer risk. Methods: We performed a hospital-based case-control study with 155 cases and 313 controls in China. All Chinese cases with newly diagnosed primary ovarian cancer between May 2005 to May 2010 in our hospital were invited to participate within 2 months of diagnosis. Controls were randomly selected from people who requested general health examinations in the same hospital during the same period. SNPs in EXCC1, ERCC1 C8092A and ERCC1 T19007C, were analyzed by PCR-RFLP method. Results: We observed a non-significantly increased risk of ovarian cancer among individuals with ERCC1 8092TT compared with those with the 8092CC genotype (adjusted OR=1.55, 95% CI%=0.74-2.97). Moreover, 19007TT genotype carriers also showed a non-significant increased risk of ovarian cancer over those with the 19007CC genotype (adjusted OR=1.78, 95% CI%=0.91-3.64). Conclusion: Our firstly investigation of links between polymorphisms in the ERCC1 gene and the risk of ovarian cancer in Chinese population demonstrated no significant association. Further large sample studies in Chinese populations are needed.

Molecular Cloning of the Arginine Biosynthetic Genes from Corynebacterium glutamicum

  • Chun, Jae-Shick;Jung, Sam-Il;Ko, Soon-Young;Park, Mee-Young;Kim, Soo-Young;Lee, Heung-Shick;Cheon, Choong-Ill;Min, Kyung-Hee;Lee, Myeong-Sok
    • Journal of Microbiology
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    • v.34 no.4
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    • pp.355-362
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    • 1996
  • Complementation cloning of the argC, E, B, D, F, and G genes in Corynebacterium glutamicum was done by transforming the genomic DNA library into the corresponding arginine auxotrophs fo Escherichia coli. Recombinant plasmids containing 6.7 kb and 4.8kb fragments complementing the E. coli argB mutant were also able to complement the E. coli argC, E, A, D, and F mutants, indicating the clustered organization of the arginine biosynthetic genes within the cloned DNA fragments. The insert DNA fragments in the recombinant plasmids, named pRB1 AND pRB2, were physically mapped with several restriction enzymes. By further subcloning the entire DNA fragment containing the functions and by complementation analysis, we located the arg genes in the order of ACEBDF on the restriction map. We also determined the DNA nucleotide sequence of the fragment and report here the sequence of the argB gene. When compared to that with the mutant strain, higher enzyme activity of N-acetylglutamate kinase was detected in the extract of the mutant carrying the plasmid containing the putative argB gene, indicating that the plasmid contains a functional argB gene. Deduced amino acid sequence of the argB gene shows 45%, 38%, and 25% identity to that from Bacillus strearothermophilus, Bacillus substilus, and E. coli respectively. Our long term goal is genetically engineering C. glutamicum which produces more arginine than a wild type strain does.

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