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RISS 인기검색어
- 검색키워드
- 저자 : Davoud Koolivand (검색결과 4 건)
- 무료
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Koolivand, Davoud,Bashir, Nemat Sokhandan,Behjatnia, Seyed Aliakbar,Joozani, Raziallah Jafari The Korean Society of Plant Pathology 2016 Plant Pathology Journal Vol.32 No.5
The genomic region of Grapevine fanleaf virus (GFLV) encoding the movement protein (MP) was cloned into pET21a and transformed into Escherichia coli strain BL21 (DE3) to express the protein. Induction was made with a wide range of isopropyl-${\beta}$-D-thiogalactopyranoside (IPTG) concentrations (1, 1.5, and 2 mM) each for duration of 4, 6, or 16 h. However, the highest expression level was achieved with 1 mM IPTG for 4 h. Identity of the expressed protein was confirmed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) followed by Western blotting. The expressed 41 kDa protein was purified under denaturing condition by affinity chromatography, reconfirmed by Western blotting and plate-trapped antigen enzyme-linked immunosorbent assay (PTA-ELISA) before being used as a recombinant antigen to raise polyclonal antibodies in rabbits. Purified anti-GFLV MP immunoglobulines (IgGs) and conjugated IgGs detected the expressed MP and GFLV virions in infected grapevines when used in PTA-ELISA, double antibody sandwich-ELISA, and Western blotting. This is the first report on the production of anti-GFLV MP polyclonal antibodies and application for the virus detection.
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Davoud Koolivand,Nemat Sokhandan Bashir,Seyed Aliakbar Behjatnia,Raziallah Jafari Joozani 한국식물병리학회 2016 Plant Pathology Journal Vol.32 No.5
The genomic region of Grapevine fanleaf virus (GFLV)encoding the movement protein (MP) was clonedinto pET21a and transformed into Escherichia colistrain BL21 (DE3) to express the protein. Inductionwas made with a wide range of isopropyl-β-D-thiogalactopyranoside(IPTG) concentrations (1, 1.5, and2 mM) each for duration of 4, 6, or 16 h. However,the highest expression level was achieved with 1 mMIPTG for 4 h. Identity of the expressed protein wasconfirmed by sodium dodecyl sulphate polyacrylamidegel electrophoresis (SDS-PAGE) followed by Westernblotting. The expressed 41 kDa protein was purifiedunder denaturing condition by affinity chromatography,reconfirmed by Western blotting and platetrappedantigen enzyme-linked immunosorbent assay(PTA-ELISA) before being used as a recombinant antigento raise polyclonal antibodies in rabbits. Purifiedanti-GFLV MP immunoglobulines (IgGs) and conjugatedIgGs detected the expressed MP and GFLV virionsin infected grapevines when used in PTA-ELISA,double antibody sandwich-ELISA, and Western blotting. This is the first report on the production of anti-GFLV MP polyclonalantibodies and application forthe virus detection.
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Abadkhah, Mahsa,Koolivand, Davoud,Eini, Omid The Korean Society of Plant Pathology 2018 Plant Pathology Journal Vol.34 No.6
Tomato spotted wilt virus (TSWV; Genus Orthotospovirus: Family Tospoviridae) is one of the most destructive viruses affecting a wide range of horticultural crops on a worldwide basis. In 2015 and 2016, 171 leaf and fruit samples from tomato (Solanum lycopersicum) plants with viral symptoms were collected from the fields in various regions of Iran. ELISA test revealed that the samples were infected by TSWV. The results of RT-PCR showed that the expected DNA fragments of about 819 bp in length were amplified using a pair of universal primer corresponding to the RNA polymerase gene and DNA fragments of ca 777 bp and 724 bp in length were amplified using specific primers that have been designed based on the nucleocapsid (N) and non-structural (NSs) genes, respectively. The amplified fragments were cloned into pTG19-T and sequenced. Sequence comparisons with those available in the GenBank showed that the sequences belong to TSWV. The high nucleotide identity and similarities of new sequences based on the L, N, and NSs genes showed that minor evolutionary differences exist amongst the isolates. The phylogenetic tree grouped all isolates six clades based on N and NSs genes. Phylogenetic analysis showed that the Iranian isolates were composed a new distinct clade based on a part of polymerase, N and NSs genes. To our knowledge, this is the first detailed study on molecular characterization and genetic diversity of TSWV isolates from tomato in Iran that could be known as new clade of TSWV isolates.
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Mahsa Abadkhah,Davoud Koolivand,Omid Eini 한국식물병리학회 2018 Plant Pathology Journal Vol.34 No.6
Tomato spotted wilt virus (TSWV; Genus Orthotospovirus: Family Tospoviridae) is one of the most destructive viruses affecting a wide range of horticultural crops on a worldwide basis. In 2015 and 2016, 171 leaf and fruit samples from tomato (Solanum lycopersicum) plants with viral symptoms were collected from the fields in various regions of Iran. ELISA test revealed that the samples were infected by TSWV. The results of RTPCR showed that the expected DNA fragments of about 819 bp in length were amplified using a pair of universal primer corresponding to the RNA polymerase gene and DNA fragments of ca 777 bp and 724 bp in length were amplified using specific primers that have been designed based on the nucleocapsid (N) and non-structural (NSs) genes, respectively. The amplified fragments were cloned into pTG19-T and sequenced. Sequence comparisons with those available in the GenBank showed that the sequences belong to TSWV. The high nucleotide identity and similarities of new sequences based on the L, N, and NSs genes showed that minor evolutionary differences exist amongst the isolates. The phylogenetic tree grouped all isolates six clades based on N and NSs genes. Phylogenetic analysis showed that the Iranian isolates were composed a new distinct clade based on a part of polymerase, N and NSs genes. To our knowledge, this is the first detailed study on molecular characterization and genetic diversity of TSWV isolates from tomato in Iran that could be known as new clade of TSWV isolates.
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