Selected article for: "blunt end ligation and end ligation"
Author: Wang, Li; Xia, Tian; Guo, Tiantian; Ru, Yi; Jiang, Yanping; Cui, Wen; Zhou, Han; Qiao, Xinyuan; Tang, Lijie; Xu, Yigang; Li, Yijing
Title: Recombinant Lactobacillus casei Expressing Capsid Protein VP60 can Serve as Vaccine Against Rabbit Hemorrhagic Disease Virus in Rabbits
  • Document date: 2019_11_2
    • ID: 1apf9w7j_10
    Snippet: A schematic diagram of the recombinant DNA plasmid is shown in Figure 1 . In brief, recombinant plasmid pMD18-T simple-eGFP was cleaved with SacI and KpnI restriction endonuclease and inserted into the corresponding sites of a pMD18-T simple-VP60 expression vector digested using SacI and KpnI. This generated recombinant plasmid pMD18-T simple-eGFP-VP60. The eGFP-VP60 fragment, cleaved with SacI and ApaI, was inserted into a constitutive expressio.....
    Document: A schematic diagram of the recombinant DNA plasmid is shown in Figure 1 . In brief, recombinant plasmid pMD18-T simple-eGFP was cleaved with SacI and KpnI restriction endonuclease and inserted into the corresponding sites of a pMD18-T simple-VP60 expression vector digested using SacI and KpnI. This generated recombinant plasmid pMD18-T simple-eGFP-VP60. The eGFP-VP60 fragment, cleaved with SacI and ApaI, was inserted into a constitutive expression vector pPG-T7g10-PPT, resulting in the recombinant plasmid pPG-T7g10-eGFP-VP60 (Cm + ). Afterwards, the Cm resistance gene in pPG-T7g10-eGFP-VP60 (Cm + ) was disrupted using MunI and NcoI. Finally, the recombinant plasmid pPG-T7g10-eGFP-VP60 was obtained via blunt-end ligation and transformed into competent LC393 cells by electroporation to generate recombinant L. casei pPG-eGFP-VP60/LC393. ATGGAGGGCAAAACCCGCACAGCGC-3′; 5′-TCAGACATAAGAAAAGCCATTGGCT-3′. pPG-T7g10-PPT, constructed in our previous study [33] , was used as a constitutive expression plasmid in this study; it encoded resistance to chloramphenicol (Cm) and contained an HCE promoter, T7g10 enhancer, PgsA anchor, and the rrnBT1T2 terminator. Plasmids pMD18-T simple-eGFP and pMD18-T simple-VP60 were constructed in our laboratory previously.

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