• Asian-Australasian Journal of Animal Sciences
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• v.7 no.3
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• pp.427-434
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• 1994

The primordial germ cells (PGCs) were transfected in vitro and expressed the exogenous RSVLTR/${\beta}G2$ plasmid, suggesting thaI PGC is a possible vector for direct gene transfer into the germ line. Transfection efficiency of cell suspensions containing PGCs was 1.5% by liposome mediated DNA transfection. By microinjection of the transfected PGCs into the host germinal crescent, PGCs migrated via blood vessel to the future gonad and these transfected PGCs resulted in the RSVLTR/${\beta}G2$ expression in the gonad. The results from the seeding of PGCs on the chorioallantoic membrane were insufficient to test the hypothesis that PGCs can penetrate or invade the chorioallantoic membrane for transport via the circulatory system.

• Journal of Microbiology and Biotechnology
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• v.23 no.1
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• pp.131-135
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• 2013

A DOTAP analog labeled by NBD on the head group (DTNBD) was designed and synthesized to label DOTAP liposome. The structure was confirmed by $^1H$ NMR and FAB-MS, and the fluorescence of the newly synthesized DT-NBD was observed by fluorescent microscopy. The transfection efficiency of DOTAP liposome containing DT-NBD was comparable to commonly used NBD PE in COS7 and MCF7 cells. Furthermore, the level of cellular uptake and fluorescent intensity of fluorescent liposome containing DT-NBD was higher than NBD PE. Therefore, the novel NBD-labeled DOTAP analog seems to be effectively used for investigation of the cellular interaction and transfection mechanism of DOTAP liposome.

• KSBB Journal
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• v.16 no.6
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• pp.579-591
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• 2001

The cell growth inhibitor effect of cervical cancer cells was investigated by liposome mediated transfection (pRcCMVp53/lipofectin) and by transfection using adenovirus (AdCMVp57). The papilloma virus cancer cell lines we used in this study were HPV16 positive, having inhibiter gene, wild p53 gene, CaSki, SiHa, HPV18 positive HeLa, HeLaS3 and HPV negative C33A, HT3. LacZ gene of E.coli was used as the marker gene for the transfection efficiency. The effect on the inhibition of tumor cell growth was measured by cell count and cell viability though ELISA analysis and MTT assay. The inhibition of tumor cell growth was confirmed by measuring each assay for six days, comparing with the normal control cell growth. The cell growth of cervical cancer calls by transfection was significantly reduced and showed tittle differences among the cell lines. To eliminate the potential problem of Ad(adenovirus) contamination during rAAV production, rAAV can be produced by a triple transfection of vector plasmic, packaging plasmid, and adenovirus helper plasmid. To examine the helper functions of Ad plasmids on the production of rAAV vector, we carried out cotransfection of three plasmids, AAV vector, packaging construct, and Ad helper plasmids. The optimized transfection condition for calcium phosphate method is 25ug of total DNA per 10-cm-diameter plate of 293 cell. We found that rAAV yields peaked at 48hr after Ad infection. The titer of rAAV was measured by the dot blot analysis to measure the number of particles/ml based on the quantification of viral DNA. Recent1y, Kombucha(fungi) was identified as a very potent antileukefic agent. In the present study, effect of natural toxin(plankton) and Kombucha is PSP(GTXI-3, neoSTX), on various MTT assay cervical cancer cell line. Toxin(GTX 1-3, neoSTX) also inhibited the proliferation in primary cervical cancer calls in a dose-dependent toxin concentration. These results showed that toxin was very potent in inhibiting the proliferation of cervical cancer calls in vitro. Toxins and Kombuoha exhibited a dose dependent inhibition of cellular proliferation in cancer cell line.

• Reproductive and Developmental Biology
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• v.33 no.2
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• pp.107-111
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• 2009

The transfection efficiency of a transgene into pig and goat fetal fibroblast cells (PFF and GFF, respectively) was tested using co-transfection of a human tissue-type plasminogen activator (tPA) transgene and neomycin-resistant ($Neo^r$) gene, followed by G418 selection. To initially test G418 resistance, GFF and PFF were incubated in culture medium containing different concentration of G418 for 2 weeks, and cell survival was monitored over time. Based on the obtained results, the concentrations chosen for G418 selection were 800 ug/ml and 200 ug/ml for GFF and PFF, respectively. For co-transfection experiments, the pBC1/tPA and $Neo^r$ vectors were co-transfected into GFF and PFF ($1{\times}10^6$ cells in each case) using the FuGENE6 transfection reagent, and resistant colonies were obtained following 14 days of G418 selection. We obtained 96 and 93 drug-resistant colonies of GFF and PFF, respectively, only 54 and 39 of which, respectively, continued proliferating after drug selection. PCR-based screening revealed that 23 out of 54 analyzed GFF colonies and 5 out of 39 analyzed PFF colonies contained insertion of the tPA gene. Thus, the experimentally determined transfection efficiencies for tPA gene co-transfection with the $Neo^r$ gene were 42.6% for GFF and 12.8% for PFF. These findings suggest that co-transfection of a transgene with the $Neo^r$ gene can aid in the successful integration of the transgene into fetal fibroblast cells.

• Journal of Microbiology and Biotechnology
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• v.10 no.1
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• pp.91-94
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• 2000

The suspension cells of Taxus cuspidata were transformed with Agrobacterium tumefaciens harboring binary vector pCAMBIE1302 encoding mgfp. Transient transfection efficiency was compared by using the fluoremetric measurement. The transient transfection efficiency was improved by transformation with DMSO and/or sonication treatment. Optimum conditions for DMSO and sonication treatment were 3% and 30sec, respectively. selection and maintenance of transformed cells were continued for 3 months. An insertion of the mgfp gene in transformed cells was detected by PCR and an expression of GFP confirmed by the western blot analysis.

• Proceedings of the Polymer Society of Korea Conference
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• 2006.10a
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• pp.178-178
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• 2006

Chitosan has been investigated as a non-viral vector because it has several advantages such as biocompatibility, biodegradability and low toxicity with high cationic potential. However, low specificity and low transfection efficiency of chitosan as a DNA carrier need to be overcome for clinical trials. In this study, chemical modification for enhancement of cell specificity and transfection efficiency was investigated. Also, the chitosan derivative formulations in vivo were included.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.9
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• pp.4435-4439
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• 2012

Purpose: This study aimed to explore the role of the Twist gene in the epithelial-mesenchymal transition of ovarian cancer. Methods: An RNA interference plasmid expressing a small interfering RNA (siRNA)-targeting Twist (Twist siRNA vector) was designed, constructed, and transfected into the human ovarian cancer cell line A2780. Transfection efficiency was assessed under a fluorescence microscope. Changes in the expression of Twist mRNA in A2780 after transfection with the pGenesil Twist shRNA plasmid were analyzed through RT-PCR. MTT assays and adhesion experiments were applied to determine changes in proliferation and adhesion ability of A2870 after transfection with the Twist shRNA plasmid. Changes in the expression of the E-cadherin and N-cadherin proteins in A2780 after transfection with the Twist shRNA plasmid were analyzed using Western blotting. Result: The restructuring plasmid pGenesil-Twist shRNA was constructed successfully. After 48 h of culture, 80% of the cells expressed high-intensity GFP fluorescence and stability. The expression of Twist decreased significantly after the transfection of the Twist shRNA plasmid (P<0.05). Proliferation of the transfected Twist shRNA cells showed no difference with that of the A2780-nontransfection or A2780-si-control groups (P>0.05) but the adhesion ability of A2780 decreased dramatically (P<0.05). Expression of the E-cadherin protein increased, whereas that of the N-cadherin protein decreased compared with that in the A2780-nontransfection or A2780-si-control groups (P<0.05). Conclusion: Twist is essential for epithelial-mesenchymal transition, invasion, and metastasis of ovarian cancer.

• Journal of the Korean Chemical Society
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• v.43 no.2
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• pp.193-198
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• 1999

The transfection efficiency of plasmid DNA was inspected using multi-cationic polymer, 5, 10, 25 and 50KD polyethylenimine (PEI). The optimal neutralization ratio of PEI/DNA complexes by agarose assay was 1.5-2.0 (nmol/nmol) without much difference in molecular weight of PEI.In vitro transfection assay, most of PEI-mediated plasmid delivery was better compared to the naked DNA. Especially, 25KD PEI at optimal condition gave higher transfection rather than the standard assay of DEAE-dextran or Lipofectin. To enhance the cell targeting delivery, the liposome formulations were introduced using phospholipids. As a result, PC/PE liposomes increased 2-2.5 times of the transfection efficiency of PEI single or PC/PE single delivery, but not the case of 25KD PEI. Moreover, the DOTAP/PE-introduced PEI delivery reduced the transfection of DOTAP/PE single delivery. All these results proved that the PEI can be used not only good transfectants and but also good DNA condensing agents in neutral/anionic liposome for cell targeting delivery.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.17 no.11
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• pp.640-647
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• 2016

By minimizing fluorescence interference phenomena, aequorin-based luminescence technology can provide a relatively sensitive detection platform with integration of $G{\alpha}16$ protein in order to track internal calcium mobilization by G protein-coupled receptors (GPCR). In this type of cell-based functional assay format, it is essential to optimize the transfection process of a receptor and $G{\alpha}16$ protein. For this study, corticotropin releasing factor receptor subtype 2(CRF2) was set as a model system to generate three stable cells with CRF2 and $G{\alpha}16$ in addition to transiently transfected cells under three different conditions. Agonist (sauvagine) and antagonist (K41498) responses in those cells were analyzed to develop the optimum transfection process. As a result, the effective signal ratio in the dose response experiments of sauvagine and K41498 were at least 10-fold higher (z'=0.77) in CRF2-$G{\alpha}16$ stable cells. For the transient transfection cells, stable expression of $G{\alpha}16$ prior to the CRF2 represented a two-fold higher signal (z'=0.84) than the other cases of transient transfection. In conclusion, for the utilization of transient transfection processes to develop a cell-based GPCR functional assay system, it is suggested to introduce various target receptors after stable expression of $G{\alpha}16$ protein.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.12
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• pp.4915-4918
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• 2014

Background and Aims: Advances in the treatment of cervical cancer over the last decade have predominantly involved the development of genes directed at molecular targets. Gene therapy is recognized to be a novel method for the treatment of cervical cancer. Genes can be administered into target cells via nanocarriers. This study aimed to develop systemically administrable nano-vectors. Floate (Fa) containing gene loaded nanoparticles (NPs) could target HeLa human cervical cancer cells through combination with receptors on the cells to increase the nuclear uptake of genetic materials. Methods: Fa was linked onto Poly (ethylene glycol)-b-poly (D, L-lactide) (PEG-PLA) to form Fa-PEG-PLA, and the resulting material was used to load plasmids of enhanced green fluorescence protein (pEGFP) to obtain gene loaded nanoparticles (Fa-NPs/DNA). Physical-chemical characteristics, in vitro release and cytotoxicity of Fa-NPs/DNA were evaluated. The in vitro transfection efficiency of Fa-NPs/DNA was evaluated in HeLa cells and human umbilical vein endothelial cells (HUVEC). PEG-PLA without Fa was used to load pEGFP from NPs/DNA as a control. Results: Fa-NPs/DNA has a particle size of 183 nm and a gene loading quantity of 92%. After 72h of transfection, Fa-NPs/DNA displayed over 20% higher transfection efficiency than NPs/DNA and 40% higher than naked DNA in HeLa cells. However, in HUVECs, no significant difference appeared between Fa-NPs/DNA and NPs/DNA. Conclusions: Fa-PEG-PLA NPs could function as excellent materials for gene loading. This nano-approach could be used as tumor cell targeted medicine for the treatment of cervical cancer.