• Title/Summary/Keyword: infectious clones

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Construction of Infectious cDNA Clone of a Chrysanthemum stunt viroid Korean Isolate

  • Yoon, Ju-Yeon;Cho, In-Sook;Choi, Gug-Seoun;Choi, Seung-Kook
    • The Plant Pathology Journal
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    • v.30 no.1
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    • pp.68-74
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    • 2014
  • Chrysanthemum stunt viroid (CSVd), a noncoding infectious RNA molecule, causes seriously economic losses of chrysanthemum for 3 or 4 years after its first infection. Monomeric cDNA clones of CSVd isolate SK1 (CSVd-SK1) were constructed in the plasmids pGEM-T easy vector and pUC19 vector. Linear positive-sense transcripts synthesized in vitro from the full-length monomeric cDNA clones of CSVd-SK1 could infect systemically tomato seedlings and chrysanthemum plants, suggesting that the linear CSVd RNA transcribed from the cDNA clones could be replicated as efficiently as circular CSVd in host species. However, direct inoculation of plasmid cDNA clones containing full-length monomeric cDNA of CSVd-SK1 failed to infect tomato and chrysanthemum and linear negative-sense transcripts from the plasmid DNAs were not infectious in the two plant species. The cDNA sequences of progeny viroid in systemically infected tomato and chrysanthemum showed a few substitutions at a specific nucleotide position, but there were no deletions and insertions in the sequences of the CSVd progeny from tomato and chrysanthemum plants.

Molecular and Biological Characterization of an Isolate of Cucumber mosaic virus from Glycine soja by Generating its Infectious Full-genome cDNA Clones

  • Phan, Mi Sa Vo;Seo, Jang-Kyun;Choi, Hong-Soo;Lee, Su-Heon;Kim, Kook-Hyung
    • The Plant Pathology Journal
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    • v.30 no.2
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    • pp.159-167
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    • 2014
  • Molecular and biological characteristics of an isolate of Cucumber mosaic virus (CMV) from Glycine soja (wild soybean), named as CMV-209, was examined in this study. Comparison of nucleotide sequences and phylogenetic analyses of CMV-209 with the other CMV strains revealed that CMV-209 belonged to CMV subgroup I. However, CMV-209 showed some genetic distance from the CMV strains assigned to subgroup IA or subgroup IB. Infectious full-genome cDNA clones of CMV-209 were generated under the control of the Cauliflower mosaic virus 35S promoter. Infectivity of the CMV-209 clones was evaluated in Nicotiana benthamiana and various legume species. Our assays revealed that CMV-209 could systemically infect Glycine soja (wild soybean) and Pisum sativum (pea) as well as N. benthamiana, but not the other legume species.

Complete Genome Sequencing and Infectious cDNA Clone Construction of Soybean Mosaic Virus Isolated from Shanxi

  • Wang, Defu;Cui, Liyan;Zhang, Li;Ma, Zhennan;Niu, Yanbing
    • The Plant Pathology Journal
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    • v.37 no.2
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    • pp.162-172
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    • 2021
  • Soybean mosaic virus (SMV) is the predominant viral pathogen that affects the yield and quality of soybean. The natural host range for SMV is very narrow, and generally limited to Leguminosae. However, we found that SMV can naturally infect Pinellia ternata and Atractylodes macrocephala. In order to clarify the molecular mechanisms underlying the cross-family infection of SMV, we used double-stranded RNA extraction, rapid amplification of cDNA ends polymerase chain reaction and Gibson assembly techniques to carry out SMV full-length genome amplification from susceptible soybeans and constructed an infectious cDNA clone for SMV. The genome of the SMV Shanxi isolate (SMV-SX) consists of 9,587 nt and encodes a polyprotein consisting of 3,067 aa. SMV-SX and SMV-XFQ008 had the highest nucleotide and amino acid sequence identities of 97.03% and 98.50%, respectively. A phylogenetic tree indicated that SMV-SX and SMV-XFQ018 were clustered together, sharing the closest relationship. We then constructed a pSMV-SX infectious cDNA clone by Gibson assembly technology and used this clone to inoculate soybean and Ailanthus altissima; the symptoms of these hosts were similar to those caused by the virus isolated from natural infected plant tissue. This method of construction not only makes up for the time-consuming and laborious defect of traditional methods used to construct infectious cDNA clones, but also avoids the toxicity of the Potyvirus special sequence to Escherichia coli, thus providing a useful cloning strategy for the construction of infectious cDNA clones for other viruses and laying down a foundation for the further investigation of SMV cross-family infection mechanisms.

A new serotype confirmed by partial physical mapping of cDNA clones from the infectious pancreatic necrosis virus (IPNV) isolated in Korea (한국에서 분리된 전염성 췌장괴저 바이러스의 새로운 혈청형에 대한 유전자 분석)

  • 이정진;박정우;정가진
    • Korean Journal of Microbiology
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    • v.27 no.3
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    • pp.231-236
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    • 1989
  • The larger segment of double stranded RNA genome from a new serotype of Infectious Pancreatic Necrosis Virus (IPNV), DRT, has been partially cloned at Sma I site in pUC19 and compared with the restriction maps of VR-299 and Sp. Restrction sites found in DRT was distinct and hence a new serotype. The cDNA clones of DRT were about 800, 850, and 1, 400 bp long each and do not share any common restiction site. It is not clear yet if there exist any overlapping sequences among them. This partial cloning, however, was sufficient for the comparison of restriction maps with the other serotypes.

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Pathogenicity of infectious in vitro transcripts and comparison of RNA3 of Alfalfa mosaic virus Korean isolates

  • J.H. Ha;Park, J.K.;K.H. Ryu
    • Proceedings of the Korean Society of Plant Pathology Conference
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    • 2003.10a
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    • pp.146.2-147
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    • 2003
  • Two Korean isolates of Alfalfa mosaic virus (AHV-AZ, AMV-KR) were isolated from azuki bean and potato plants, respectively, and their pathologies were confirmed on some susceptible host plants including pepper, tobacco and red bean plants. Full length cDNAs to RNA1, RNA2 and RNA3 of the two Korean strains were amplified using the long-template reverse transcription (RT)-polymerase chain reaction (PCR) method. RT-PCR products covering entire regions for the three AMV genome RNAs were cloned. RNA transcripts were synthesized in vitro from each clones using T7 RNA polymerase and infectivity test was peformed in 9 reassortment sets of transcripts. All the combinations of reassorted transcripts were found to be infectious when inoculated onto Nicotiana benthamiana plants, and were not distinguishable to those of wild types. The full-length cDNA clones that were confirmed infectious were sequenced their nucleotide sequences. We will discuss sequence analysis of the two Korean isolates of AMV genomic RNA3 and compare reported foreign isolates of AMV.

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Clinical Manifestation and Treatment of Methicillin-resistant Staphylococcus aureus Infections in Children (소아 메티실린내성 황색포도알균 감염증의 임상양상과 치료)

  • Choi, Eun Hwa
    • Pediatric Infection and Vaccine
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    • v.16 no.1
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    • pp.1-5
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    • 2009
  • Methicillin-resistant Staphylococcus aureus (MRSA), a leading cause of nosocomial infections, has been increasingly recognized in communities of the United States. This article will review the clinical spectrum and treatment of MRSA infections in children in the context of recent epidemiological changes of MRSA infections. In general, community-associated (CA) MRSA most frequently causes skin and soft tissue infections and has an increased association with invasive infections, particularly pneumonia and musculoskeletal infections. Hospital-associated (HA) MRSA strains tend to be associated with bloodstream infections, pneumonia, and surgical site infections. Different from the United States, CA-MRSA infections are not common in Korea (only 5.9%); however, there are some CA-MRSA clones that are different from HA-MRSA clones in Korea and from CA-MRSA clones in other countries. The treatment of MRSA infections should be guided by antimicrobial susceptibility testing, the site of infection, and the infection severity. Vancomycin is the treatment of choice for invasive MRSA infections. Other agents such as trimethoprim-sulfamethoxazole, clindamycin, linezolid, quinupristin-dalfopristin, and daptomycin have been used for some conditions.

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A Rapid and Efficient Method for Construction of an Infectious Clone of Tomato yellow leaf curl virus

  • Bang, Bongjun;Lee, Jongyun;Kim, Sunyoung;Park, Jungwook;Nguyen, Thao Thi;Seo, Young-Su
    • The Plant Pathology Journal
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    • v.30 no.3
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    • pp.310-315
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    • 2014
  • Tomato yellow leaf curl virus (TYLCV), a member of the genus Begomovirus, is responsible for one of the most devastating viral diseases in tomato-growing countries and is becoming a serious problem in many subtropical and tropical countries. The climate in Korea is getting warmer and developing subtropical features in response to global warming. These changes are being accompanied by TYLCV, which is now becoming a large problem in the Korean tomato industry. The most effective way to reduce damage caused by TYLCV is to breed resistant varieties of tomatoes. To accomplish this, it is necessary to establish a simple inoculation technique for the efficient evaluation of resistance to TYLCV. Here, we present the rolling circle amplification (RCA) method, which employs a bacteriophage using phi-29 DNA polymerase for construction of infectious TYLCV clones. The RCA method is simple, does not require sequence information for cloning, and is less expensive and time consuming than conventional PCR based-methods. Furthermore, RCA-based construction of an infectious clone can be very useful to other emerging and unknown geminiviruses in Korea.