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- Title
Expression of Avian Reovirus (ARV) σA Protein in HEK293T Cells.
- Authors
Ren Hongyu; Xie Zhixun; Xie Liji; Wang Sheng; Huang Jiaoling; Fan Qing; Luo Sisi; Zhang Yanfang; Zeng Tingting; Zhang Mingxiu; Xie Zhiqin; Deng Xianwen
- Abstract
`[Objective] The paper was to construct eukaryotic expression vector of Avian reovirus (ARV)σA gene and express σA protein accurately in HKK293T cells. [Methodl The specific primers of ARV σA gene were designed according to the gene sequence of ARV S2 gene in GenBank (accession number KF741763.1). With pMD18-T-σA recombinant vector as the template, the specific sequence of aA gene was amplified by PCR and cloned into pMD18-T vector to construct recombinant plasmid. The cloning vector pMD18-T-σA and eukaryotic expression plasmid pKKla-HA were double digested by restriction enzymes Kpn I and iNol I. The purified <tA gene was connected with pEF1α-HA to construct eukaryotic expression plasmid pEF1α-HA-aA. After colony 1'CH, double enzyme digestion and sequencing, the recombinant plasmid pEFlα-HA-σA was tansfected into HEK293T colls. The proteins were collected at 24 h after tansfeetion and verified by Western-blol. [Result] The ARV σA gene was successfully cloned in the test. The eukaryotic expression plasmid pEFlα-HA-aA was constructed, which could be expressed in I1EK293T cells. [Conclusion] The protein could be accurately expressed in HEK293T cells.
- Subjects
DNA primers; PROTEINS; GENETIC vectors; CELLS
- Publication
Animal Husbandry & Feed Science, 2019, Vol 11, Issue 5/6, p145
- ISSN
1943-9911
- Publication type
Article
- DOI
10.19578/j.cnki.ahfs.2019.05-06.005