• 환경생물
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• 제23권4호
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• pp.379-382
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• 2005

The new cationic meso-substituted N-quarternized 4-pyridylporphyrins and their metal derivatives were synthesized as novel chemotherapeutics. The level of DNA damage induced by porphyrins TOBut4PyP, TOBut4PyP, TOEt4PyPMn and TOBut4PyPMn and its dependence on the chemical structure of compounds were analyzed by the Comet-assay. On the base of data obtained, the investigated porphyrins may be arranged by their genotoxic activity in the following order: TOEt4PyP>TOEt4PyPMn>TOBut4PyP>TOBut4PyPMn. Thus, i) the genotoxicity of the Mn-derivatives of TOEt4PyP and TOBut4PyP is higher than the original porphyrins and ii) the genotoxicity of TOEt4PyP and TOEt4PyPMn is increased after substitution of a butyl radical for ethyl one. The applied Comet-assay permits to reveal the dependence of DNA damage induction on the chemical structure of porphyrins.

• Journal of Plant Biotechnology
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• 제42권3호
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• pp.154-160
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• 2015

The plant breeding technology was developed with genetic engineering. Many researchers and breeders have turned from traditional breeding to molecular breeding. Genetically modified organisms (GMO) were developed via molecular breeding technology. Currently, molecular breeding technologies facilitate efficient plant breeding without introducing foreign genes, in virtue by of gene editing technology. Gene targeting (GT) via homologous recombination (HR) is one of the best gene editing methods available to modify specific DNA sequences in genomes. GT utilizes DNA repair pathways. Thus, DNA repair systems are controlled to enhance HR processing. Engineered sequence specific endonucleases were applied to improve GT efficiency. Engineered sequence specific endonucleases like the zinc finger nuclease (ZFN), TAL effector nuclease (TALEN), and CRISPR-Cas9 create DNA double-strand breaks (DSB) that can stimulate HR at a target site. RecQl4, Exo1 and Rad51 are effectors that enhance DSB repair via the HR pathway. This review focuses on recent developments in engineered sequence specific endonucleases and ways to improve the efficiency of GT via HR effectors in plants.

• 한국동물학회지
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• 제26권1호
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• pp.19-28
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• 1983

본 연구는 알킬화제이며 항암항생제인 Mitomycin C(MMC)와 Bleomycin(BLM)이 DNA 복제 및 회복에 미치는 영향을 규명하고, 아울러 MMC에 의한 "유발상해회복"이 포유동물세포에 유발되는지 밝히고자 수행하였다. 이를 위해 자기방사법에 의한 비주기 DNA 합성율과 알카리 유출법에 의한 DNA단사절단율을 측정하였다. CHO세포에 MMC를 제 1차 $(MMC_1)$ 처리한 후 5시간 뒤에 제 2차 $(MMC_2)$로 처리하여 얻은 결과는 다음과 같았다. 1. BLM은 비주기 DNA합성을 매우 적게 유발시켰으며, BLM $5\\mug/ml$의 농도에서 부터 비주기 DNA합성율은 증가를 보이지 않았다. BLM은 처리후 1.5시간까지 DNA합성억제를 보였으며, 1.5시간후 DNA합성율이 증가되었으나, 대조군의 60% 수준까지 회복되었을 뿐이다. 2. MMC에 의해 유발된 비주기 DNA합성은 농도에 따라 비례하였으며, 배양후 시간의 간격에 따라 비례하여 감소하였다. 제 1,2차 MMC를 처리한 세포의 비주기 DNA합성은 제 1차만 처리한 세포의 비주기 DNA합성보다 많았다. 3. 알카리 유출법 결과는 MMC에 의하여 유발된 DNA 단백질 연결로 인한 DNA 단사절단율이 농도에 비례함을 나타내었다. DNA 단백질 연결로 인한 DNA 단사절단율은 배양후의 시간에 비례해서 감소되었다. 이러한 결과는 MMC와 BLM 모두가 DNA 상해제임과 MMC에 의한 DNA 상해 부위의 복제율이 어떠한 유발기작에 의하여 촉진됨을 시사한다.촉진됨을 시사한다.

• 한국식품영양과학회지
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• 제12권4호
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• pp.305-309
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• 1983

수종(數種)의 핵산구성성분(核酸構成成分), 아미노산(酸), 요소유도체(尿素誘導體)와 같은 생리활성물질(生理活性物質) 및 함질소화합물(含窒素化合物)의 DNA에 대한 작용(作用)과 그 작용(作用)에 무기(無機)ion의 영향을 검토(檢討)하였다. PTU, Cys-SH는 금속(金屬) ion없이도 DNA 절단능력(切斷能力)을 가지고 있었고, Tyr, Phe, Trp는 $CuSO_4$존재(存在) 하(下)에서 약한 DNA damage 유기작용(誘起作用)이 있었으며 5mM 이상의 Cys-SH와 $500{\mu}M$ $CuSO_4$의 혼합액(混合液)은 강(强)한 DNA 절단작용(切斷作用)을 나타냈다. $Sn^{2+}$를 제외한 $500{\mu}M$의 무기염(無機鹽)은 DNA에 대한 절단능력이 없었으나, $Cu^{2+}$, $Ni^{2+}$, $Mn^{2+}$, $Zn^{2+}$, $Sn^{2+}$ 등은 수종(數種)의 아미노산(酸)의 DNA breaking action에 영향을 미치고 있음을 확인(確認)되었다.

• BMB Reports
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• 제37권4호
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• pp.445-453
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• 2004

We identified apoptosis as being a significant mechanism of toxicity following the exposure of HeLa cell cultures to abrin holotoxin, which is in addition to its inhibition of protein biosynthesis by N-glycosidase activity. The treatment of HeLa cell cultures with abrin resulted in apoptotic cell death, as characterized by morphological and biochemical changes, i.e., cell shrinkage, internucleosomal DNA fragmentation, the occurrence of hypodiploid DNA, chromatin condensation, nuclear breakdown, DNA single strand breaks by TUNEL assay, and phosphatidylserine (PS) externalization. This apoptotic cell death was accompanied by caspase-9 and caspase-3 activation, as indicated by the cleavage of caspase substrates, which was preceded by mitochondrial cytochrome c release. The broad-spectrum caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (zVAD-fmk), prevented abrin-triggered caspase activation and partially abolished apoptotic cell death, but did not affect mitochondrial cytochrome c release. These results suggest that the release of mitochondrial cytochrome c, and the sequential caspase-9 and caspase-3 activations are important events in the signal transduction pathway of abrin-induced apoptotic cell death in the HeLa cell line.

• BMB Reports
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• 제56권2호
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• pp.102-107
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• 2023

Genome editing using CRISPR-associated technology is widely used to modify the genomes rapidly and efficiently on specific DNA double-strand breaks (DSBs) induced by Cas9 endonuclease. However, despite swift advance in Cas9 engineering, structural basis of Cas9-recognition and cleavage complex remains unclear. Proper assembly of this complex correlates to effective Cas9 activity, leading to high efficacy of genome editing events. Here, we develop a CRISPR/Cas9-RAD51 plasmid constitutively expressing RAD51, which can bind to single-stranded DNA for DSB repair. We show that the efficiency of CRISPR-mediated genome editing can be significantly improved by expressing RAD51, responsible for DSB repair via homologous recombination (HR), in both gene knock-out and knock-in processes. In cells with CRISPR/Cas9-RAD51 plasmid, expression of the target genes (cohesin SMC3 and GAPDH) was reduced by more than 1.9-fold compared to the CRISPR/Cas9 plasmid for knock-out of genes. Furthermore, CRISPR/Cas9-RAD51 enhanced the knock-in efficiency of DsRed donor DNA. Thus, the CRISPR/Cas9-RAD51 system is useful for applications requiring precise and efficient genome edits not accessible to HR-deficient cell genome editing and for developing CRISPR/Cas9-mediated knockout technology.

• Journal of Preventive Medicine and Public Health
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• 제37권4호
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• pp.373-380
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• 2004

Objectives : There has been gradually increasing concern about the adverse health effects of electromagnetic radiation originating from cell phones which are widely used in modern life. Cell phone radiation may affect human health by increasing free radicals of human blood cells. This study has been designed to identify DNA damage of blood cells by electromagnetic radiation caused by cell phone use. Methods : This study investigated the health effect of acute exposure to commercially available cell phones on certain parameters such as an indicator of DNA damage for 14 healthy adult volunteers. Each volunteer during the experiment talked over the cell phone with the keypad facing the right side of the face for 4 hours. The single cell gel electrophoresis assay (Comet assay), which is very sensitive in detecting the presence of DNA strand-breaks and alkali-labile damage in individual cells, was used to assess peripheral blood cells (T-cells, B-cells, granulocytes) from volunteers before and after exposure to cell phone radiation. The parameters of Comet assay measured were Olive Tail Moment and Tail DNA %. Results : The Olive Tail Moment of B-cells and granulocytes and Tail DNA % of B-cells and granulocytes were increased by a statistically significant extent after 4-hour use of a cell phone compared with controls. Conclusion : It is concluded that cell phone radiation caused the DNA damage during the 4 hours of experimental condition. Nonetheless, this study suggested that cell phone use may increase DNA damage by electromagnetic radiation and other contributing factors.

• 환경생물
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• 제19권1호
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• pp.19-24
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• 2001

병해충을 막기 위해 농업용 살충제가 광범위하게 사용되고 있다. 농약사용에 따른 생물학적 위해가 우려되며 농약이 또 다른 환경유해요인과 인체에 상승적으로 작용할 경우 농업재해로 이어질 가능성이 있다. 다양한 인자에 의한 DNA손상을 감지하는데 유용한 단세포 겔 전기영동법을 이용하여 살충제와 방사선에 의한 사람 림프구 DNA손상을 평가하였다. 각기 다른 농도로 살충제를 10분간 전처리한 림프구와 정상 림프구에 0-2.0 Gy의 방사선으로 조사한 다음 DNA 손상도를 평가하였다. DNA가닥 절단에 대한 표식인 tail moment의 증가는 감마선에 대해서 뚜렷한 선량-반응 관계를 나타내었다. 단세포 겔 전기영동법을 통한 평가결과 권장 사용농도 이상의 살충제는 림프구에 대한 유전독성을 나타내었을 뿐 아니라 방사선과 함께 상승작용을 일으켜 림프구 DNA손상을 더욱 증가시키는 것이 확인되었다. 이와 같은 결과는 환경재해의 방재 및 예방 조치에 필요한 생물정보를 사전에 제공하는 의미를 가진다.

• Molecular & Cellular Toxicology
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• 제1권2호
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• pp.111-115
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• 2005

Ultraviolet (UV) radiation to mammalian skin is known to alter cellular function via generation of Reactive Oxygen Species (ROS), DNA damage and DNA lesions, such as pyrimidine dimmers and photoproducts, which could lead to DNA mutation if they are not repaired. In this study, we have investigated the reduction of DNA damage and of apoptosis with a particular attention to genetic effect of paeoniflorin in Normal Human Epidermal Keratinocytes (NHEK). After UVB irradiation from $10\;to\;500mJ/cm^{2}$ to NHEK, Mean Tail Moments (MTM) were increased with UVB dose increase. The greatest amount of strand breaks was induced at $500mJ/cm^{2}$ of UVB. Even at the lowest dose of UVB ($10mJ/cm^{2}$), change in MTM was detected (P<0.0001). Pretreated cell with 0.1% paeoniflorin maximally reduced the level of DNA damage to about 21.3%, compared to untreated cell. In the lower concentrations less than 0.01% of paeoniflorin, MTM had a small increase but paeoniflorin still had reductive effects of DNA damage. We measured the apoptosis suppression of paeoniflorin with annexin V flous staining kit. As we observed under the fluorescence microscopy to detect apoptosis in the irradiated cell, the fluorescence intensity was clearly increased in the untreated cell, but decreased in treated cells with paeoniflorin. These results suggest that paeoniflorin reduces the alteration of cell membranes and prevents DNA damage. Therefore, the use of paeoniflorin as a free radical scavenger to reduce the harmful effects of UV lights such as chronic skin damage, wrinkling and skin cancer can be useful to prevent the formation of photooxidants that result in radical damage.

• Molecules and Cells
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• 제26권1호
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• pp.5-11
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• 2008

Stalled DNA replication forks activate specific DNA repair mechanism called post-replication repair (PRR) pathways that simply bypass DNA damage. The bypassing of DNA damage by PRR prevents prolonged stalling of DNA replication that could result in double strand breaks (DSBs). Proliferating cell nuclear antigen (PCNA) functions to initiate and choose different bypassing pathways of PRR. In yeast, DNA replication forks stalled by DNA damage induces monoubiquitination of PCNA at K164, which is catalyzed by Rad6/Rad18 complex. PCNA monoubiquitination triggers the replacement of replicative polymerase with special translesion synthesis (TLS) polymerases that are able to replicate past DNA lesions. The PCNA interaction motif and/or the ubiquitin binding motif in most TLS polymerases seem to be important for the regulation of TLS. The TLS pathway is usually error-prone because TLS polymerases have low fidelity and no proofreading activity. PCNA can also be further polyubiquitinated by Ubc13/ Mms2/Rad5 complex, which adds an ubiquitin chain onto monoubiquitinated K164 of PCNA. PCNA polyubiquitination directs a different PRR pathway known as error-free damage avoidance, which uses the newly synthesized sister chromatid as a template to bypass DNA damage presumably through template switching mechanism. Mammalian homologues of all of the yeast PRR proteins have been identified, thus PRR is well conserved throughout evolution. Mutations of some PRR genes are associated with a higher risk for cancers in mice and human patients, strongly supporting the importance of PRR as a tumor suppressor pathway.