• Molecules and Cells
• /
• 제42권1호
• /
• pp.67-78
• /
• 2019

Methylation of HBV cccDNA has been detected in vivo and in vitro; however, the mechanism and its effects on HBV replication remain unclear. HBx derived from a 1.2-mer HBV replicon upregulated protein levels and enzyme activities of DNA methyltransferase 1 (DNMT1), 3a, and 3b, resulting in methylation of the negative regulatory region (NRE) in cccDNA, while none of these effects were observed with an HBx-null mutant. The HBx-positive HBV cccDNA expressed higher levels of HBc and produced about 4-fold higher levels of HBV particles than those from the HBx-null counterpart. For these effects, HBx interrupted the action of NRE binding protein via methylation of the C-1619 within NRE, resulting in activation of the core promoter. Treatment with 5-Aza-2′dC or DNMT1 knock-down drastically impaired the ability of HBx to activate the core promoter and stimulate HBV replication in 1.2-mer HBV replicon and in vitro infection systems, indicating the positive role of HBx-mediated cccDNA methylation in HBV replication.

• 한국미생물·생명공학회지
• /
• 제15권5호
• /
• pp.319-327
• /
• 1987

알코올 생산성이 높은 Zymomonas 균주의 기질 이용성을 넓히기 위한 목적으로 natural replicon을 포함하며 적당한 항생제 저항표지를 갖는 plasmid vector의 제조를 시도하였다. Z. mobilis ATCC10988에서 분리된 몇 개의 plasmid중 3.9kb의 적당한 크기를 갖는 pZM3를 선정하여 수종의 제한효소로 처리하여 절편의 크기에 따라 유전자 지도를 작성하였다. pZM 3의 replicon과 pBR 325의 chloramphenicol 저항유전자를 포함한 재조합 plasmid인 pHZ22를 개발하고 이 plasmid vector가 숙주세포인 Z. mobilis ATCC31821에서 독립적으로 replication됨을 확인하였다. 또 하나의 항생제 저항표지로서 RP4의 tetracycline 저항유전자를 분리하여 pHZ22에 도입함으로써 pHZT224를 제조하였는데 이 plasmid vector도 Zymomonas로 conjugation에 의해 전이되어 안정하게 유지 되었다. 본 연구를 통하여 개발된 plasmid vector는 Z. mobilis와 E. coli에 공히 작용하는 shuttle vector 로서 외부 유전자를 Zymomonas에 도입시킬 수 있는 유용한 유전자 운반체임이 확인되었다.

• Journal of Microbiology and Biotechnology
• /
• 제15권6호
• /
• pp.1229-1239
• /
• 2005

Human immunodeficiency virus-like particles (HIVVLPs) with native conformations similar to that of the wild-type virion could be valid candidates for vaccine development. To this end, we used a Semliki Forest Virus (SFV) expression system to produce HIV- VLPs containing high quantities of native envelope proteins. Here, we described a single SFV replicon containing the HIV gagpol and env genes under the control of separate subgenomic promoters. Mature VLPs incorporating the Gag and Env proteins were detected in the supernatant of replicon-expressing cells by Western blot analysis. The HIV-VLPs showed the expected molecular density (1.14-1.18 g/ml) on a $20-60\%$ sucrose gradient; the particles were 100-120 nm in diameter and Env proteins were observed on their surfaces by immunogold electron microscopy. RT-PCR analysis of VLP-associated RNAs in mature HIV-VLPs revealed two SF V-derived RNA species (full-length and subgenomic). Immunization studies in Balb/c mice showed that these HIV-VLPs were capable of inducing both HIV-specific antibodies and cell-mediated immune responses. Taken together, our results indicate that the SFV replicon system is useful for the production of HIV-VLPs, which may be valuable candidates for an HIV vaccine.

• International Journal of Industrial Entomology and Biomaterials
• /
• 제2권2호
• /
• pp.155-160
• /
• 2001

We have generated a novel baculovirus genome which can be maintained in Escherichia coli that facilitates the rapid and efficient generation of recombinant baculovirus expression vectors. To make $Occ^{+}$ recombinant expression vectors, polyhedrin gene under the control of p10 promoter was inserted to bAcGOZA and this genome was designated bApGOZA. As in bAcGOZA, bApGOZA lacks a portion of the essential ORF1629 gene, but includes a mini-F replicon and selectable kanamycin-resistance marker, This occasion-producing activity of bApGOZA can be used very conveniently for its oral infectivity to insect larvae in mass production of foreign protein and insecticides.

• 식품과학과 산업
• /
• 제50권1호
• /
• pp.67-73
• /
• 2017

Control of food pathogens is critical in food safety field. Norovirus is one of the major causes of gastroenteritis and food poisoning worldwide, however, currently, there is not a vaccine or a specific drug available for its treatment. There are several methods to inactivate norovirus during food processing by chemical and physical treatments, however, the use of natural substance has been suggested as an optional strategy due to their safety and consumer preference. In this study supported by Ministry of Food and Drug Safety in Korea, we identified novel plant-derived substances with significant anti-norovirus activities. The aim of this project was to determine the antiviral activity of a wide range of natural substances, including plant-derived extracts and essential oils, using a norovirus surrogate system, human norovirus replicon-bearing cells, and mouse in vivo experiments. During the activity screening test, we identified novel anti-norovirus substances or oils using plaque assay with MNV-1. Six selected substances were formulated into an optimum mixture and used as an ingredient for salad sauce of which anti-novovirus activity was confirmed(pending for patent and paper submission). The potential application of selected natural substances as a metal surface sanitizer was also tested. Interestingly, the mixture of selected natural compounds showed a significant inhibitory effect against norovirus. These results suggest that these substances may be used as food ingredient with anti-norovirus antivity or components for surface sanitizers to prevent norovirus contamination.

• 한국환경농학회지
• /
• 제1권1호
• /
• pp.48-52
• /
• 1982

수중미생물(水中微生物)의 계수방법(計數方法)은 크게 둘로 나누인다. 첫째 방법(方法)은 replicon개염(槪念)에 기초(基礎)를 둔 것으로 살아있는균(菌)과 죽은 균(菌)을 구별할 수 있으나 사용(使用)되는 배지(培地)가 생리적(生理的)으로 다른 다양(多樣)한 세균(細菌)들로 구성된 수중세균(水中細菌)의 생육(生育)에 적합하지 않아서 전(全) 세균수(細菌數)를 계수(計數)할 수 없다. 둘째 방법(方法)은 직접 검경하여 계수(計數)하는 방법(方法)으로 살아있는 균(菌)과 죽은균(菌), 세균(細菌)과 particle의 구별(區別)이 곤란하다. 그러나 최근 세균염색수(細菌染色術)의 발달(發達)로 수중세균(水中細菌)을 육안(肉眼)으로 쉽게 구별(區別)하여 계수(計數)할 수 있는 방법(方法)이 가능(可能)하게 되었다. 이 방법(方法)은 형광현징경(螢光顯徵鏡)을 사용(使用)하여 acridine orange로 염색(染色)된 수중(水中)의 전세균수(全細菌數)를 측정(測定)하는 방법(方法)이다. 따라서 본(本) 연구(硏究)는 상수도원(上水道源)의 세균오염도(細菌汚染度)를 측정(測定)하는데 새로운 방법(方法)으로서 epifluorcscence microscopy를 제시(提示)하는데 있으며 이의 사용성가능여부(使用性可能與否)를 진단(診斷)하기 위해 chlorine과 chloramine을 시료수(試料水)에 처리(處理)하여 경시적으로 세균수(細菌數)를 조사(調査)하여 평판법(平板法)과 비교(比較)하였다. 시료수(試料水)(sample water)의 전세균수(全細菌數)는 형광검경법(螢光檢鏡法)에 의(依)해 정확(正確)히 측정(測定)되었으며 분리능력(分離能力)(resolution)에 있어서 탁월(卓越)하였고 또한 경제적(經濟的)이고도 간편하였다. 그리고 소독살균제(消毒殺菌劑)로는 chloramine이 감소(監素)보다 수중세균(水中細菌)에 미치는 영향(影響)은 훨씬 컸다.