Open AccessExpression of Recombinant Fusion Protein of NDV from Escherichia coli BL-21 C-1b Clone using AccurapidTM Cell-Free Protein Expression System
Author(s) -
K N Andrian,
Aris Tri Haryanto
Publication year - 2021
Publication title -
the epic series in biological sciences
Language(s) - English
Resource type - Conference proceedings
ISSN - 2398-5291
DOI - 10.29007/9mr9
Subject(s) - recombinant dna , newcastle disease , microbiology and biotechnology , biology , fusion protein , virology , virus , restriction enzyme , expression vector , plasmid , gene , biochemistry
Newcastle Disease (ND) is an infectious disease that infected poultry caused by Newcastle Disease Virus (NDV). NDV is a virus from family Paramyxoviridae which enveloped, unsegmented single strand (ss) RNA negative sense. Fusion (F) protein is an NDV protein that has an important role in the infection process of NDV to the host cell. F protein of NDV can induce the immune response of the host so that it has the potential to become an NDV vaccine. Recombinant plasmid pBT7-N-His-F has been designed by inserting the F protein-encoding gene of NDV. The inserted gene is an F protein of NDV from a local isolate from Galur, Kulon Progo, Yogyakarta. This study aimed to investigate the in vitro expression of the recombinant F of NDV from recombinant plasmid pBT7-N-His-F NDV from E. coli BL-21 clone C-1b. Samples of recombinant plasmid pBT7-N-His-F NDV showed a DNA fragment in size of 4.642 bp. The DNA fragment was then digested by the EcoRI enzyme to separate the pBT7-N-His vector with the F protein-encoding gene as a DNA insert. Restriction digest generated two DNA fragments in the size of 4.001 bp and 642 bp. The in vitro expression of recombinant F protein was conducted using the AccuRapidTM Cell-free Expression system. The visualization of the expressed recombinant F protein using the SDS-PAGE method showed a recombinant protein fragment in size of 25.6 kDa. The Western blotting on the polyvinyl difluoride (PVDF) membrane showed a recombinant protein fragment in size of 25.6 kDa. The recombinant F protein of NDV was successfully in vitro expressed from clone C-1b of recombinant plasmid pBT7-N-His-FNDV by AccuRapidTM Cell-free protein expression system.