In vitro expression of the recombinant fusion protein of Newcastle disease virus from local Indonesian isolates by using a cell-free protein expression system

https://doi.org/10.22146/ijbiotech.54703

Aris Haryanto(1), Hevi Wihadmadyatami(2), Nastiti Wijayanti(3*)

(1) Department of Biochemistry and Molecular Biology, Faculty of Veterinary Medicine, Universitas Gadjah Mada, Jl. Fauna No. 2, Karangmalang, Yogyakarta 55281
(2) Department of Anatomy, Faculty of Veterinary Medicine, Universitas Gadjah Mada, Jl. Fauna No. 2, Karangmalang, Yogyakarta 55281
(3) Laboratory of Animal Physiology, Faculty of Biology, Universitas Gadjah Mada, Jl. Teknika Selatan, Sekip Utara, Bulaksumur, Yogyakarta 55281
(*) Corresponding Author

Abstract


The aim of this work was the in vitro expression of the recombinant fusion (F) protein of Newcastle disease virus (NDV).  The pBT7-N-His-Fusion-NDV expression plasmid which carries the recombinant F protein encoding gene from local Indonesian isolates, was prepared and transformed into E. coli BL21 (DE3). To detect bacterial colonies carrying the recombinant plasmid, a restriction endonuclease analysis was performed using the EcoRI restriction endonuclease. These results showed that the pBT-N-His-Fusion-NDV plasmid was successfully isolated with a size of 4.601 bp, and three recombinant plasmids carrying the gene coding for the recombinant F protein of NDV were obtained. Selected recombinant plasmids were then in vitro by using a cell-free protein expression system followed by visualization of the recombinant F protein on a 12% SDS-PAGE gel both by Coomassie Brilliant Blue staining and Western blotting. Recombinant F protein was successfully in vitro expressed by using a cell-free protein expression system as indicated by a specific single protein band with a molecular mass of 25.6 kDa.


Keywords


cell-free protein expression; in vitro expression; Newcastle diseases virus (NDV); recombinant F protein

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DOI: https://doi.org/10.22146/ijbiotech.54703

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