• BMB Reports
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• v.38 no.3
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• pp.354-359
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• 2005

Open reading frame 29 (ha29) is a gene specific for Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus (HearSNPV). Sequence analyses showed that the transcription factor Tfb2 motif, bromodomain and Half-A-TPR (HAT) repeat were present at aa 66-82, 4-76, 55-90 of the Ha29 protein respectively. The product of Ha29 was detected in HearSNPV-infected HzAM1 cells at 3 h post-infection. Western blot analysis using a polyclonal antibody produced by immunizing a rabbit with purified GST-Ha29 fusion protein indicates that Ha29 is an early gene. The size of Ha29 product in infected HzAM1 cells was about 25 kDa, which was larger than the presumed size of 20.4 kDa. Tunicamycin treatment of HearSNPV-infected HzAM1 cells suggested that the Ha29 protein is N-glycosylated. Fluorescent confocal laser scanning microscope examination, and Western blot analysis of purified budded virus (BVs), occlusion-derived virus (ODVs), cell nuclear and cytoplasmic fraction, showed that the Ha29 protein was localized in the nucleus. Our results suggested that ha29 of HearSNPV encodes a non-structurally functional protein that may be associated with virus gene transcription in Helicoverpa hosts.

• The Journal of Natural Sciences
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• v.10 no.1
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• pp.33-37
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• 1998

Cytokinin is one of major growth regulators in plants. In this study, the gene isopentenyl transferase (jpt) which encodes a key enzyme involved in the biosynthesis of the growth regulator cytokinin isolated from Agrobacterium tumefaciens was introduced ito tobacco plant via Agrobacterium-mediated transformation. The jpt gene was modulated using the proteinase inhibitor II (PI-IIK) promotor. In general, this promoterlipt gene fusion resulted in overproduction of cytokinins throughout the transgenic plants. The overproduction of cytokinin caused dramatic changes in morphology of the plant, including stem thickness and reduced root development. The studies reported in this paper were initiated to examine the consequences of overproduction of cytokinin in plant.

• International Journal of Industrial Entomology and Biomaterials
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• v.15 no.1
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• pp.35-38
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• 2007

In this study, ORF 46 of Bombyx mod nucleopolyhedrovirus(Bm46) fused with EGFP was expressed in Bombyx mod cell lines, larvae and pupae by BmNPV Bacmid system. Bm46 and EGFP were cloned into donor plasmid pFastBacHTb, which was transformed to competent DH10B cells containing helper and BmNPV bacmid by site-specific transposition. Recombinant bacmid was used to transfected BmN-4 cells to produce the recombinant baculovirus vBm-Bm46-EGFP. Recombination virus was injected into silkworm larvae and pupae. The expression of the fusion protein was monitored by examining green fluorescence using a fluorescent microscope. Intense fluorescence in cells and silkworm was observed at 4 days post-infection, indicating the Bm46-EGFP fusion gene was expressed successfully.

• Journal of Industrial Technology
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• v.29 no.B
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• pp.115-119
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• 2009

Green fluorescent protein (GFPuv) was displayed on the surface of ethanol-producing bacteria Zymomonas mobilis using N-terminal domain of ice nucleation protein (INP) as an anchoring motif. To evaluate the ice nucleation protein as plausible anchor motif in Z. mobilis, GFPuv gene was subcloned into Zymomonas expression vector yielding pBBR1MCS-3/pPDC/INPN/GFPuv plasmid., INP-GFPuv fusion protein was expressed in Z. mobilis and its fluorescence was verified by confocal microscopy. The successful display of GFPuv on Zymomonas mobilis suggest that INP anchor motif could be used for future fusion partner in Z. mobilis strain improvement.

• Journal of Microbiology
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• v.44 no.1
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• pp.35-41
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• 2006

The unique gene encoding thioredoxin reductase (TrxR) was previously cloned and characterized from the fission yeast Schizosaccharomyces pombe, and its expression was induced by oxidative stress. To elucidate tbe regulatory mechanism of the S. pombe TrxR gene, three fusion plasmids were generated using polymerase chain reaction: pYUTR20, pYUTR30, and pYUTR40. Plasmid pYUTR20 has an upstream region of 891 base pairs, pYUTR30 has 499 in this region, and pYUTR40 has an 186 bp upstream region. Negatively acting sequence is located between $-1,526\;\~\;-891bp$ upstream of the gene. The upstream sequence, responsible for the induction of TrxR by menadione (MD), is situated on the $-499\;\~\;-186bp$ region, which is also required for TrxR induction by mercuric chloride. The same region also appeared to be required for Pap1-mediated transcriptional regulation of the TrxR gene, which contains the two plausible Papl binding sites, TTACGAAT and TTACGCGA. Consistently, basal and inducible expression of the TrxR gene was markedly lower in the Pap1-negative TP108-3C cells than in wild-type yeast cells. In summary, up-regulation of the S. pombe TrxR gene is mediated by Pap1 via the transcriptional motif(s) located on the $-499\;\~\;-186bp$ region.

• Journal of Microbiology
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• v.42 no.3
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• pp.233-238
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• 2004

Transcriptional regulation of the Schizosaccharomyces pombe y-glutamylcysteine synthetase (GCS) gene was examined using the two GCS-lacZ fusion plasmids pUGCS101 and pUGCS102, which harbor 607 bp and 447 bp upstream regions, respectively. The negatively-acting sequence was located in the -607 - -447 bp upstream region of the GCS gene. The upstream sequence responsible for induction by menadione(MD) and L-buthionine-(S, R)-sulfoximine (BSO) resides in the -607 - -447 bp region, whereas the sequence which codes for nitric oxide induction is located within the -447 bp region, measured from the translational initiation point. Carbon source-dependent regulation of the GCS gene appeared to be dependent on the nucleotide sequence within -447 bp region. The transcription factor Papl is involved in the induction of the GCS gene by MD and BSO, but not by nitric oxide. Induction of the GCS gene occurring due to low glucose concentration does not depend on the presence of Pap1. These data imply that induction by MD and BSO may be mediated by the Pap1 binding site, probably located in the -607 - -447 region, and also that the nitric oxide-mediated regulation of the S. pombe GCS gene may share a similar mechanism with its carbon-dependent induction.

• Journal of Life Science
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• v.14 no.4
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• pp.657-663
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• 2004

Tobacco transformants harboring cinnamic acid 4-hydroxylase gene (C4H) fused with susquiterpene cyclase promoter was developed in order to regulate biosynthesis of phenolic compounds by the expression of the introduced gene. Twenty transformants for each specific promoter were used to analyze the incorporation of the chimeric genes by PCR and Southern blot analysis. PCR products of NPTII(neomycin phosphotransferase) gene (553bp) were detected in the transgenic tobacco plants. The incorporation of the chimeric gene was confirmed in the Southern blot analysis. C4H activity in the transgenic plants was elevated by UV-irradiation and its level was higher compared to that of control plants.

• Korean Journal of Microbiology
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• v.40 no.2
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• pp.87-93
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• 2004

Dynamic modification of cytoplasmic and nuclear proteins by O-linked N-acetylglucosamine (O-GlcNAc) on Ser and Thr residues is ubiquitous in higher eukaryotes. And this modification may serve as a signaling mod-ification analogous to protein phosphorylation. Addition and cleavage of O-GlcNAc are catalyzed by O-linked GlcNAc transferase (OGT) and O-linked N-acety1glucosaminidase (O-GlcNAcase), respectively. Two types of human O-GlcNAcase gene were cloned and expressed as three fusion proteins in Escherichia coli. O-GlcNA-case activity showed in the order of thioredoxin fusion> $6{\times}His$ tag> GST fusion. O-GlcNAcase had enzy-matic activity against only ${\rho}$NP-GlcNAc of seven tested substrate analogs. Blast search revealed that O-GlcNAcase has two conserved domains, amino terminal hyaluronidase-like domain and carboxy terminal N-acetyltransferase domain. Extensive deletion studies were done to define catalytically important domains. The deletions of hyaluronidase-like domain and N-acetyltransferase domain abolished enzyme activity. But, N-ter-minal 55 amino acid deletion and C-terminal truncation showed lower activity. Based on deletion analysis, we suggest that hyaluronidase-like domain is essential for enzyme activity and carboxy terminal N-acetyltrans-ferase domain may be modulatory function.

• Journal of Life Science
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• v.29 no.3
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• pp.376-381
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• 2019

To construct useful yeast strain for bioethanol production, we improved yeast harboring various phenotypes by using yeast protoplast fusion method. In this study, S. cerevisiae BYK-F11 strain which have ethanol tolerance, thermotolerance and ${\beta}-glucanase$ activity and P. $stipitis{\Delta}ura$ strain which has xylose metabolism pathway were fused by genome shuffling. P. $stipitis{\Delta}ura$ strain was constructed for protoplast fusion by URA3 gene disruption, resulting in uracil auxotroph. By protoplast fusion, several fused cells were selected and BYKPS-F8 strain (fused cell) showing both karyotypes from two parent strains (S. cerevisiae BYK-F11 and P. $stipitis{\Delta}ura$ strain) among 22 fused cells was finally selected. Sequentially, various phenotypes such as ${\beta}-glucanase$ activity, xylose utility, ethanol tolerance, thermotolerance and ethanol productivity were analyzed. The BYKPS-F8 strain obtained ${\beta}-glucanase$ activity from BYK-F11 strain and 1.2 fold increased xylose utility from P. $stipitis{\Delta}ura$ strain. Also, the BYKPS-F8 strain showed thermotolerance at $40^{\circ}C$ and increased ethanol tolerance in medium containing 8% ethanol. In this fused cell, 7.5 g/l ethanol from 20 g/l xylose was produced and the multiple phenotypes were stably remained during long term cultivation (260 hr). It was proved that novel biological system (yeast strains) is easily and efficiently bred by protoplast fusion among yeasts having different genus.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.7
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• pp.3349-3355
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• 2012

Background and objectives: Chromosomal abnormalities play an important role in genesis of acute lymphoblastic leukemia (ALL) and have prognostic implications. Five major risk stratifying fusion genes in ALL are BCR-ABL, MLL-AF4, ETV6-RUNX11, E2A-PBX1 and SIL-TAL1. This work aimed to detect common chromosomal translocations and associated fusion oncogenes in adult ALL patients and study their relationship with clinical features and treatment outcome. Methods: We studied fusion oncogenes in 104 adult ALL patients using RT-PCR and interphase-FISH at diagnosis and their association with clinical characteristics and treatment outcome. Results: Five most common fusion genes i.e. BCR-ABL (t 9; 22), TCF3-PBX1 (t 1; 19), ETV6-RUNX1 (t 12; 21), MLL-AF4 (t 4; 11) and SIL-TAL1 (Del 1p32) were found in 82/104 (79%) patients. TCF3-PBX1 fusion gene was associated with lymphadenopathy, SIL-TAL1 positive patients had frequent organomegaly and usually presented with a platelets count of less than $50{\times}10^9/l$. Survival of patients with fusion gene ETV6-RUNX1 was better when compared to patients harboring other genes. MLL-AF4 and BCR-ABL positivity characterized a subset of adult ALL patients with aggressive clinical behaviour and a poor outcome. Conclusions: This is the first study from Pakistan which investigated the frequency of5 fusion oncogenes in adult ALL patients, and their association with clinical features, treatment response and outcome. Frequencies of some of the oncogenes were different from those reported elsewhere and they appear to be associated with distinct clinical characteristics and treatment outcome. This information will help in the prognostic stratification and risk adapted management of adult ALL patients.