• Title/Summary/Keyword: vector-mediated virus

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Foamy Virus Integrase in Development of Viral Vector for Gene Therapy

  • Kim, Jinsun;Lee, Ga-Eun;Shin, Cha-Gyun
    • Journal of Microbiology and Biotechnology
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    • v.30 no.9
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    • pp.1273-1281
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    • 2020
  • Due to the broad host suitability of viral vectors and their high gene delivery capacity, many researchers are focusing on viral vector-mediated gene therapy. Among the retroviruses, foamy viruses have been considered potential gene therapy vectors because of their non-pathogenicity. To date, the prototype foamy virus is the only retrovirus that has a high-resolution structure of intasomes, nucleoprotein complexes formed by integrase, and viral DNA. The integration of viral DNA into the host chromosome is an essential step for viral vector development. This process is mediated by virally encoded integrase, which catalyzes unique chemical reactions. Additionally, recent studies on foamy virus integrase elucidated the catalytic functions of its three distinct domains and their effect on viral pathogenicity. This review focuses on recent advancements in biochemical, structural, and functional studies of foamy virus integrase for gene therapy vector research.

Titer Amplification of GALV (Gibbon Ape Leukemia Virus) Pseudotyped Retrovirus Vectors Produced from PG13 Cells (PG13 Cell로부터 생산된 GALV (Gibbon Ape Leukemia Virus)-pseudotyped Retrovirus Vector의 증폭)

  • 김태완;박윤엽;권모선;염행철;김경화;박영식;박세필
    • Korean Journal of Animal Reproduction
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    • v.21 no.4
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    • pp.397-403
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    • 1997
  • For the ultimate goal of efficient retrovirus vector-mediated transgenic animal production, we tried to increase virus titer by employing three methods: boosting virus production by treating virus-producing cells with sodium butyrate, concentration of virus stock by either filtration or ultracentrifugation. Compared to the control, applications of sodium butyrate (5 mM) treatment and filtration resulted in only 3 and 3. 6 folds of titer increases on bovine EBTr target cells, respectively. However, concentration of virus-containing medium by ultracentrifugation showed 12.5 folds of titer increase compared to the control (10${\times}$10$^5$ LacZ$^+$ TU Im), indicating the best method which can enhance retrovirus vector-mediated transgenic animal production.

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PVY Resistant Transgenic Potato Plants (cv Claustar) Expressing the Viral Coat Protein

  • Gargouri-Bouzid Radhia;Jaoua Leila;Mansour Riadh Ben;Hathat Yemna;Ayadi Malika;Ellouz Radhouane
    • Journal of Plant Biotechnology
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    • v.7 no.3
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    • pp.143-148
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    • 2005
  • The coat protein mediated resistance to potato virus Y is assessed here in transgenic potato plants (Solanum tuberosum L., cv Claustar). Therefore, the corresponding cDNA from tunisian isolate of the virus was cloned into Agrobacterium tumefaciens binary vector. The transgenic lines were subsequently analysed for the presence and expression of the transgene. The CP cDNA copy number was determined for kanamycin resistant plants. Three selected transgenic lines and their S1 progeny resulting from tuber germination showed a high protection level against the virus. These data appear to support the hypothesis that the virus resistance is mediated by the translated viral coat protein expressed in transgenic potato lines.

Gene Transfer into Chicken Embryos using Defective Retroviral Vectors Packaged with Vesicular Stomatitis Virus G Glycoprotein Envelopes (Vesicular Stomatitis Virus G Glycoprotein Envelope으로 포장된 Defective Retroviral Vector를 이용한 닭의 배로의 유전자 전이)

  • 권모선;임은정;허영태;이훈택;이영만;김태완
    • Korean Journal of Animal Reproduction
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    • v.25 no.2
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    • pp.171-180
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    • 2001
  • Compared to other gene transfer system, the advantages of retrovirus-mediated gene transfer are technical ease, efficient expression and genetic stability. Despite the high potency of the retrovirus vector system in gene transfer, one of the drawbacks is a difficulty in concentration of virus stock. To overcome this problem, we tested a new retrovirus vector system producing the progeny retrovirus particles encapsidated with VSV-G (vesicular stomatitis virus G glycoprotein). The infectivity of this virus was not sacrificed by ultracentrifugal concentration and the host cell range extended from all mammalian to fish embryos. Virus titer after 1,000 x concentration was more than 10$^{8}$ CFU/ $m\ell$ on most of the target cell lines. We applied this pantropic viruses in transgenic chicken production by injecting the concentrated (100$\times$) stock into subgerminal cavity of stage X chicken embryos. The survival rate of chicken embryos after injection was about 20% and gene integration rate in surviving embryos was scored almost 100%. Analyses of RT-PCR and fluorescence microscopy, however, showed no evidence of the transgene expression.

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Construction of a Transposon-mediated Baculovirus Vector Hanpvid and a New Cell Line for Expressing Barnase

  • Qin, Qin;Liu, Ying-Le;Zhu, Ying;Li, Shun-Yi;Qi, Yi-Peng
    • BMB Reports
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    • v.38 no.1
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    • pp.41-48
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    • 2005
  • In this study we developed the transposon-mediated shuttle vector 'Hanpvid', which composed of HaNPV (Heliothis armigera nuclear polyhedrosis virus) genomic DNA and a transposon cassette from Bacmid of Bac-to-Bac system. Hanpvid replicates in E. coli in the same way as Bacmid and retains infective function in cotton bollworm cells (Hz-AM1). Using Hanpvid we constructed a recombinant virus, which could infect Hz-AM1 cells and generate recombinant HaNPV (rHa-Bar) containing the barnase gene, a ribonuclease gene from Bacillus amyloliquefaciens. Since the expression vector carrying barnase gene cannot replicate in the absence of barstar, a specific inhibitor of barnase, we constructed a new cotton bollworm cell line (AM1-NB) using the marker rescue method. In AM1-NB barstar was integrated into the cellular chromosome to sustain the replication of rHa-Bar. To screen out recombinant HaNPV for potential use as biopesticide, Hz-AM1 and AM1-NB cell lines were infected with rHa-Bar, respectively. The results obtained indicate that Viral progenies in AM1-NB were 23 and 160 times greater than those in Hz-AM1 48 h and 72 h after infection, respectively. With additional insertion of the polyhedron gene from AcNPV (Autographa californica nuclear polyhedrosis virus) into the Hanpvid genome, rHa-Bar regained the polyhedron phenotype and its pest-killing rate greatly improved. Toxic analysis showed that the lethal dosages ($LD_{50}$) and the lethal time(s) ($LT_{50}$) of rHa-Bar were reduced by 20% and 30%, respectively, compared to wt-HaNPV in the third instar larvae of cotton bollworm. This study shows that in AM1-NB barnase can be effectively produced and used as pest-killing agent for the biological control of cotton pests.

New trends of vaccine development: Recombinant vaccinia viruses (expression vectors) as vaccines (Vaccine개발(開發)의 새로운 동향(動向) : Vaccinia virus를 발견(發見) vector로 이용하는 재조합(再組合) 생(生)vaccine의 작성(作成))

  • Kim, Uh-ho
    • Korean Journal of Veterinary Research
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    • v.29 no.3
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    • pp.407-416
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    • 1989
  • The prospect of live vaccines consisting of genetically modified vaccinia virus expressing foreign genes is exciting, but important issues concerning safety and efficacy need to resolved. Vaccinia virus (VV) is an efficient expression vector with broad host range infectivity and large DNA capacity. This vector has been particularly useful for identifying target antigens for humoral and cell-mediated immunity. The WHO smallpox eradication program, involving the extensive use of VV vaccines, resulted in the late 1970s in the elimination of one of the world's most feared diseases. This achievement is a triumph for preventive medicine and for international collaboration in public health. In 1980, WHO recommended that the routine use of smallpox vaccine should be stopped. Against this background, the prospect of li ve vaccines consisting of genetically modified VV expressing foreign antigens arising from the work of Moss, and Paoletti and their colleagues in 1982 has been greeted with enthusiasm. These investigators have shown that genes coding for immunogenic proteins can be inserted into VV DNA without impairing the ability of the virus to grow in cell culture. Moreover experimental animals infected with VV recombinants containing genes coding for a variety of immunizing proteins have been shown to be protected against challenge infection with the corresponding infectious agent. In this communication, I describe current progress in the construction of a novel plasmid vector that facilitate the insertion and expression of foreign genes in VV as well as the selection of recombinants.

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Replication and encapsidation of recombinant Turnip yellow mosaic virus RNA

  • Shin, Hyun-Il;Kim, In-Cheol;Cho, Tae-Ju
    • BMB Reports
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    • v.41 no.10
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    • pp.739-744
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    • 2008
  • Turnip yellow mosaic virus (TYMV) is a positive strand RNA virus that infects mainly Cruciferae plants. In this study, the TYMV genome was modified by inserting an extra subgenomic RNA promoter and a multiple cloning site. This modified TYMV was introduced into Nicotiana benthamiana using a Agrobacterium-mediated T-DNA transfer system (agroinfiltration). When a gene encoding $\beta$-glucuronidase or green fluorescent protein was expressed using this modified TYMV as a vector, replication of the recombinant viruses, especially the virus containing $\beta$-glucuronidase gene, was severely inhibited. The suppression of replication was reduced by co-expression of viral silencing suppressor genes, such as tombusviral p19, closteroviral p21 or potyviral HC-Pro. As expected, two subgenomic RNAs were produced from the recombinant TYMV, where the larger one contained the foreign gene. An RNase protection assay revealed that the recombinant subgenomic RNA was encapsidated as efficiently as the genuine subgenomic RNA.

An epidemiological survey on seroprevalence of vector-mediated virus infection in cattle bred in a Japanese remote island, Okinawa (일본 오끼나와섬에서 사육된 소의 벡터 매개성 바이러스감염에 대한 혈청학적 역학조사)

  • Sakai, Takeo;Hamakawa, Masaaki;Abe, Sakae;Fujita, Keiichi;Ito, Fumio H.;Lee, Won-chang;Lee, Joong-bok
    • Korean Journal of Veterinary Research
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    • v.38 no.1
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    • pp.167-172
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    • 1998
  • 한국 및 일본을 포함한 동아시아 지역에 벡터 매개성 바이러스에 의한 소 질병으로 인하여 경제적 손실이 몇년을 주기로 극심하게 나타나고 있다. 그러나 본 질병의 정확한 역학에 대하여는 아직까지 확실한 연구가 되어 있지 않은 상태이다. 본 연구에서는 벡터 매개성 질병에 대한 역학을 좀더 명확하게 밝히기 위하여, 일본 열도중 가장 멀리 떨어져 있으며, 아열대 지역에 속하는 오끼나와 섬에서 사육되고 있으면서 백신접종을 받지 않은 소(Japanese black cattle)를 대상으로 하여 1988년도부터 1992년도에 걸쳐 벡터 매개성 바이러스군에 대한 중화항체가를 측정하고 분석하였다. 소의 유행성 바이러스(bovine ephemeral virus)에 대한 항체 양성율은 연도 및 계절별로 크게 변화를 보였고, Ibaraki virus에 대한 항체양성율은 그 어느 다른 계절보다도 5월에 높은 항체가를 보였으며, Akabane virus에 대한 항체 양성율은 이바라키 바이러스에 대한 항체 양성율과 유사한 수준으로 나타났다. 한편 Chuzan virus에 대한 항체양성율은 계절적인 변화가 심하지 않은 것으로 나타났다.

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Transfer and Expression of E. coli LacZ Gene in Boving Embryos by Co-culturing with Retrovirus Vector-Producing Cells (Retrovirus Vector를 생산하는 세포와 공동배양된 소 수정란의 E. coli LacZ 유전자 전이와 발현)

  • 김태완;박세필
    • Korean Journal of Animal Reproduction
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    • v.19 no.2
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    • pp.89-93
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    • 1995
  • In this study was demonstrate that retrovirus-mediated gene transfer is one of the promising alternatives to the conventional pronuclear DNA microinjection approach, especially in transferring the exogenous genes into the boving embryos. By co-culturing of zona of zona-free one-cell stage embryos with the retrovirus-producing cells for 24 hours followed by 6 days of culture in virus-free medium, we could get morulae and blastocysts expressing the E. coli LacZ genes which were transferred by our retrovirus vector. The results obtained in this study are summarized as follows : 1. Addition of 5$\mu\textrm{g}$/ml of polybrene in the embryo and virus-producing cell co-culture medium did not affect development of zona-free one-cell embryo. 2. Compared with the intact embryos removal of zona at one-cell stage before co-culturing with the virus-producing cells for one day caused only slight decrease of embryo develpment. 3. Co-culture of 625 zona-free one-cell stage embryos with the virus-producing cells resulted in 65(10.4%) morulae or blastocysts, and 12.3%(8/65) of the morulae or blastocysts were E. coli LacZ positive.

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Development of Potato Virus Y Resistant Tobacco Plant by Transformation of the Untranslatable Viral Coat Protein Encoding cDNA (감자 바이러스 Y 비전이성 외피단백질 cDNA의 형질전환에 의한 바이러스 저항성 연초품종 개발)

  • 이청호;이영기;강신웅;박성원;김상석;박은경
    • Journal of the Korean Society of Tobacco Science
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    • v.19 no.2
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    • pp.117-123
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    • 1997
  • Viral coat protein (CP) encoding cDNA with artificial start and stop codons was synthesized by reverse-transcriptase polymerase chain reaction (RT-PCR) from the Korean isolate of potato virus Y-vein nectrosis strain (pVY-VN). To make PVY CP cDNA to untranslatable form, three stop codons were inserted near the start codon by "megaprimer-PCR" method. The untranslatable CP cDNA was subcloned to plant expression vector and transferred to N. tabacum cv. NC82 by Agrobacterium-mediated transformation. Highly resistant plants to PVY infection were screened, based on symptom development after mechanical virus inoculation. By genomic PCR and Southern blot analysis, one or more copies of the untranslatable CP gene were found in all transformants. From northern blot analysis, highly resistant transgenic lines had very low level of CP transcript but susceptible lines had high level, suggesting resistance to PVY infection should be related to RNA-mediated mechanism.mechanism.

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