• Journal of Sericultural and Entomological Science
• /
• v.39 no.1
• /
• pp.36-43
• /
• 1997

In order to express the FLP recombinase in B. mori cultured cell line, BmN-4, transient expression system using a heat shock protein gene (hsp70) promoter of Dorosophilla melnogaster was constructed. This vector was designated as pHsSV. Activity strength of the hsp70 promoter was compared with that of immediate early gene (IE-1) and polyhedrin gene of BmNPV employing the E. coli $\beta$-galactosidase gene as a reporter gene. The result showed that the pHs $\beta$-gal plasmid vector expressed the $\beta$-galactosidase at 2nd and 3rd day after the transfer of plasmid DNA into BmN-4 cells, which was similar to that of pIE1 $\beta$-gal vector, but different from that of a recombinant virus, vBm $\beta$-gal. For the construction of FLP recombinase transient expression vector, the FLP recombinase gene was cloned by polymerase chain reaction technique. To express the FLP recombinase, this gene was inserted into pHsSV plasmid vector, under the control of the hsp70 promotor, and tranfected in BmN-4 cells. The expressed FLP recombinase was estimated at 44kDa on a 12.5% SDS-PAGE.

• Korean Journal of Microbiology
• /
• v.26 no.4
• /
• pp.318-323
• /
• 1988

MAK18 gene of Saccharomyces cerevisiae, needed for M1 replication, was mapped within 2cM of PET3 on chromosome VIII. From 38kb clone pRE66 carrying SPO11 and PET3, we have localized MAK18 gene whose insert is 2.8kb. MAK18 gene is Iocalized on about 9kb distance from PET3 and about 18kb distance from SPO11 on chromosome VIII.

• Environmental Mutagens and Carcinogens
• /
• v.19 no.1
• /
• pp.28-33
• /
• 1999

The SNF2 family includes proteins from a variety of species with roles I cellular processes such as transcriptional regulation, recombination and various types of DNA repair. Several proteins with unknown function are also included in this family. Here, we report the cloning and characterization of hrp 2+ gene (helicase related gene from S. pombe) which was isolated by PCR amplication using the conserved domain of SNF2 motifs within the ERCC6 gene which encodes a protein involved in DNA excision repair. The hrp2+ gene was isolated by screening with yeast S. pombe genomic library. The isolated cloned contained 6.5 kb insert DNA. Southern blot analysis confirmed that S. pombe chromosome contains the same DNA as hrp2+ gene and this gene exists as a single copy in S. pombe genome. The 4.7 kb transcript of mRNA was identified by Northern blot. To examined the transcriptional regulation of hrp2+ gene, DNA damaging agents were treated. These results indicated that the hrp2+ gene may not be directly involved in DNA replication, but may be involved in damage response pathway.

• Microbiology and Biotechnology Letters
• /
• v.26 no.1
• /
• pp.15-22
• /
• 1998

The dnrF gene, responsible for conversion of aklavinone to $\varepsilon$-rhodomycinone via C-11 hydroxylation, was mapped in the daunorubicin gene cluster of Streptomyces peucetius subsp. caesius ATCC 27952, close to drrAB, one of the anthracycline resistance genes. To characterize the enzymatic properties of the aklavinone 11-hydroxylase, the dnrF gene was overexpressed in Escherchia coli. The pET-22(+) plasmid which has the T7 promoter under the control of lacUV5 gene was used for the overexpression of the dnrF gene, and the recombinant plasmid pET213 that contains the dnrF gene linked to the T7 promoter of pET-22b(+) was introduced into the E. coli BL2l. When the expression of the dnrF gene was induced by IPTG at the final concentration of 1 mM, the induced protein could be detected in SDS-PAGE only in insoluble precipitate. The insoluble protein was electroeluted from the gel and used for the preparation of antiserum in mice. Various culture conditions were tested to maximize the expression of the aklavinone 11-hydroxylase in soluble form. The enzymatic activity was checked by the bioconversion experiment, and the protein was confirmed by the SDS-PAGE and the Western blot analysis. From the analysis of the data, it was concluded that the culture induced with IPTG at the final concentration of 0.02 mM at 37$^{\circ}C$ yielded the best productivity of active form of enzyme.

• Korean Journal of Microbiology
• /
• v.36 no.3
• /
• pp.187-191
• /
• 2000

Sulmonella gyhi is one of important causes of human enteric infections. S @phi I

• Asian Pacific Journal of Cancer Prevention
• /
• v.15 no.3
• /
• pp.1171-1176
• /
• 2014

Objective: The tumor suppressor gene, Ras-association domain family (RASSF)2A, is inactivated by promoter hypermethylation in many cancers. The current study was performed to evaluate the methylation status of RASSF2A in epithelial ovarian cancer (EOC) tissues and plasma, and correlations with gene expression and clinicopathologic characteristics. Method: We detected methylation of the RASSF2A gene in tissues and corresponding plasma samples from 47 EOC patients and 14 patients with benign ovarian tumors and 10 with normal ovarian tissues. The methylation status was determined by methylation-specific PCR while gene expression of mRNA was examined by RT-PCR. The EOC cell line, SKOV3, was treated with 5-aza-2'-deoxycytidine (5-azadC). Results: RASSF2A mRNA expression was significantly low in EOC tissues. The frequency of aberrant methylation of RASSF2A was 51.1% in EOC tissues and 36.2% in corresponding plasma samples, whereas such hypermethylation was not detected in the benign ovarial tumors and normal ovarian samples. The expression of RASSF2A mRNA was significantly down-regulated or lost in the methylated group compared to the unmethylated group (p<0.05). After treatment with 5-aza-dC, RASSF2A mRNA expression was significantly restored in the Skov3 cell line. Conclusion: Epigenetic inactivation of RASSF2A through aberrant promoter methylation may play an important role in the pathogenesis of EOC. Methylation of the RASSF2A gene in plasma may be a valuable molecular marker for the early detection of EOC.

• The Korean Journal of Zoology
• /
• v.36 no.4
• /
• pp.487-495
• /
• 1993

TRH (Thvrotropin-Releasing Hormone) known to regulate the transcription of the TSH (Thyroid-Stimulating Hormones gene in pituitary cells, but little is understood about the mechanism(sl involved. re present study was attempted to elucidate the role of Ca2+ movement through the voltage-gated channels in the regulation of TSH gene transcription. The c-fos is one of immediate early genes and used as model system for the investigation of signaling pathwavs involved in various stimuli. The changes of c-fos mRNA levels were determined after treatment of various agents using Northern and slot hybridization analysis. The c-fos mRNA was rapidly and transiently induced by TRH (about 3-fold) in GH3 cells and this induction was repressed by calcium chelating agent (EGTA), calcium channel blocker (verapamil) anti protein kinase C inhibitor (aminoacridine). The abilities of forskolin (adenvlate cvclase activators, PMA (protein kinase C activator), and A23187 (calcium ionophore) to affect c-ios gene transcription, either alone or in combination with TRH were tested in the same cells. All of them significantly increased the level of c-fos mRUA. However, no additive relationship was observed in all combined treatments except forskolin. These results suggest that TRH action on the c-fos gene activation is mediated by calcium influx as well as through protein kinase C.

• Advances in nano research
• /
• v.4 no.2
• /
• pp.145-156
• /
• 2016

Dendrimers are one of the most appropriate nanocaries for imaging moieties in imaging applications.The purpose of this study was the evalution of cytotoxicity and inducing apoptosis of dendrimers. This study was conducted in order to investigate the metastasis suppression effect of dendrimer in human breast MCF-7 cell line and finding the nanoparticle protein corona in biological enviromental. Dendrimer cytotoxicity effect was assessed by MTT assay. The mRNA experession level of KAI1 as a metastasis suppressor gene, Bax as Pro- apoptotic gene, Bcl-2 as an anti-apoptotic gene and GAPDH as a housekepping gene were determined by real-time PCR assays.concentration-dependent nanoparticle cytotoxicity effect was proofed at range of 1-2 mg/mL in 24 hours, significant upregulation of mRNA expression of Bax, was observed whereas expression of anti-apoptotic Bcl-2 was down-regulated, also expression of metastasis suppressor gene KAI1 was up-regulated. So far a few studies confirmed apoptosis enhancement effect of dendrimers in MCF-7 cell line via bax/bcl-2 pathways. dendrimer nanoparticles was able to act as metastase inhibitor via upregulation of KAI1 gene.

• Asian-Australasian Journal of Animal Sciences
• /
• v.19 no.8
• /
• pp.1100-1105
• /
• 2006

During involution of the mammary gland after the lactation period, the gland undergoes an extensive epithelial cell death. In our previous study, overexpression of an extracellular proteinase inhibitor (Expi) gene accelerated apoptosis of mammary epithelial cells. Here we found that expression of the cathepsin D gene was induced in the Expi-overexpressed apoptotic cells. To understand the role of cathepsin D in apoptosis, we transfected cathepsin D gene into mammary epithelial HC11 cells and established the stable cell lines overexpressing the cathepsin D gene. We found that overexpression of the cathepsin D gene partially induced apoptosis of mammary epithelial cells. Expression patterns of the cathepsin D gene were examined in mouse mammary gland at various reproductive stages. Expression of the cathepsin D gene was increased during involution stages compared to lactation stages, and highest expression levels were shown at involution on day 4. We also examined expression of the cathepsin D gene in various mouse tissues. Mammary gland at involution on day 2 showed highest levels of cathepsin D mRNA of the mouse tissues that we examined. Liver tissues showed high levels of cathepsin D expression. These results demonstrate that cathepsin D may contribute to the apoptotic process of mammary epithelial cells.

• Journal of Aquaculture
• /
• v.13 no.1
• /
• pp.79-85
• /
• 2000

The effects of estrogen and androgens on Vg gene expression were examined in primary hepatocyte culture and livers of the immature male trout. Specific primers of Vg cDNA were designed with already reported Vg gene nucleotide sequences. PCR product was sequenced and verified with Vg cDNA of rainbow trout. Total RNA was extracted from the cultured hepatocytes and livers of steroid-treated rainbow trout and then it was analyzed by reverse transcriptase- polymerase chain reaction (RT-PCR) analysis. The Vg mRNA and Vg protein synthesis were increased in rainbow trout in vivo and in vitro with E$_2$ and methyltestosterone (MT) There were dose and time-related effects of E$_2$ and MT on vitellogesis. Androgens such as progesterone androsterone and testosterone also stimulated Vg mRNA expression in vitro. The results show that androgens as well as E$_2$ can induce expression of Vg mRNA in trout in vivo and in vitro.