Prokaryotic Expression and Purification of Tobacco Mosaic Virus Specific <i>P</i>54<italic/> Protein

LI Cong; TIAN Peijie; ZHANG Yu; ZHANG Deyong; LIU Yong; ZHANG Songbai

doi:10.19303/j.issn.1008-0384.2021.02.011

Volume 36 Issue 2

Feb. 2021

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LI C, TIAN P J, ZHANG Y, et al. Prokaryotic Expression and Purification of Tobacco Mosaic Virus Specific P54 Protein [J]. Fujian Journal of Agricultural Sciences，2021，36（2）：209−214 doi: 10.19303/j.issn.1008-0384.2021.02.011

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LI C, TIAN P J, ZHANG Y, et al. Prokaryotic Expression and Purification of Tobacco Mosaic Virus Specific P 54 Protein [J]. Fujian Journal of Agricultural Sciences，2021，36（2）：209−214 doi: 10.19303/j.issn.1008-0384.2021.02.011

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Prokaryotic Expression and Purification of Tobacco Mosaic Virus Specific P54 Protein

doi: 10.19303/j.issn.1008-0384.2021.02.011

LI Cong^1
,,
TIAN Peijie²,
ZHANG Yu³,
ZHANG Deyong^{1, 3},
LIU Yong^{1, 3},
ZHANG Songbai^{1, 3
,
,}

1.
Longping Branch, Graduate College, Hunan University, Changsha, Hunan　410125, China
2.
Plant Protection College, Hunan Agricultural University, Changsha, Hunan　410120, China
3.
Plant Protection Institute, Hunan Academy of Agricultural Science, Changsha, Hunan　410125, China

Received Date: 2020-10-27
Rev Recd Date: 2020-11-20

Available Online: 2021-02-08

Publish Date: 2021-02-28

Abstract

Abstract

Objective Pathogenicity and molecular mechanisms of tobacco mosaic virus (TMV), a typical member belonging to Tobamovirus of Vigaviridae that infect more than 400 species in 36 families of plants causing 20%-30% reduction or complete loss on crop yield, were studied. Methods Sequence of the 1 425 bp TMV specific P54 protein was amplified by RT-PCR from the cDNA of Nicotiana tabacum var. Samsun NN infected with TMV and cloned into prokaryotic expressing plasmid, pEASY^®-Blunt E1, followed by expressing in E. coli BL21 (DE3) by IPTG induction. Subsequently, the expression products were retrieved and purified by Ni-NTA chromatography and confirmed by western-blotting identification. Results The genome of TMV was a positive-sense single-stranded RNA of 6 400 bp that encoded one structural protein and two nonstructural proteins. The amplified 1 425 bp P54 was inserted in the virus replication associated P183 gene. The prokaryotic expressed recombinant P54 was insoluble. It was retrieved and purified to show a molecular weight of approximately 60 kDa and verified by means of western blot. Conclusions TMV P54 protein was successfully expressed and purified for further study on the devastating diseases on plants caused by TMV.
- Tobacco mosaic virus,
- P54 gene,
- prokaryotic expression,
- protein purification

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References(18)

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