Open Access
Research (Published online: 05-10-2020)
6. Cloning and expression of Toxoplasma gondii GRA-4 recombinant protein as a toxoplasmosis diagnostic kit candidate
Muhammad Hanafiah, Teuku Zahrial Helmi, Amalia Sutriana, Dwi Priyowidodo and Fihiruddin Fihiruddin
Veterinary World, 13(10): 2085-2091

Muhammad Hanafiah: Parasitology Laboratory, Faculty of Veterinary Medicine, Universitas Syiah Kuala, Banda Aceh, Indonesia.
Teuku Zahrial Helmi: Laboratory of Biochemistry, Faculty of Veterinary Medicine, Universitas Syiah Kuala, Banda Aceh, Indonesia.
Amalia Sutriana: Pharmacology Laboratory, Faculty of Veterinary Medicine, Universitas Syiah Kuala Banda Aceh, Indonesia.
Dwi Priyowidodo: Department of Parasitology, Faculty of Veterinary Medicine, Gadjah Mada University, Yogyakarta, Indonesia.
Fihiruddin Fihiruddin: Department of Medical Laboratory Technology, Politeknik Kemenkes Mataram, Sandubaya Mataram Nusa Tenggara Barat Indonesia.

doi: www.doi.org/10.14202/vetworld.2020.2085-2091

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Article history: Received: 31-03-2020, Accepted: 24-08-2020, Published online: 05-10-2020

Corresponding author: Muhammad Hanafiah

E-mail: hanafi_2015@unsyiah.ac.id

Citation: Hanafiah M, Helmi TZ, Sutriana A, Priyowidodo D, Fihiruddin F (2020) Cloning and expression of Toxoplasma gondii GRA-4 recombinant protein as a toxoplasmosis diagnostic kit candidate, Veterinary World, 13(10): 2085-2091.
Abstract

Aim: The objective of this study was to produce recombinant protein GRA-4 (rGRA-4) of a local Toxoplasma gondii isolate as a candidate for a toxoplasmosis diagnosis kit in Escherichia coli BL21 (DE3) competent cells using pET SUMO plasmid.

Materials and Methods: Samples used were stock T. gondii tachyzoites DNA from the Parasitology Laboratory, Faculty of Veterinary Medicine, Gadjah Mada University, Yogyakarta. Amplified GRA-4 polymerase chain reaction product of T. gondii tachyzoite DNA was cloned in the pET-SUMO TAR cloning vector. The GRA-4 gene from T. gondii local isolate was sequenced, followed by plasmid transformation, recombinant plasmid DNA isolation, and recombinant protein expression in DE3 competent cells.

Results: The amplification product of GRA-4 T. gondii gene was 1036 bp, with 48 kDa molecular weight after expression in DE3 competent cells. An alignment of the amino acid sequence of GRA-4 from the local isolate which was cloned with GRA-4 was obtained from NCBI database and showed 99.61% homology to the predicted GRA-4 from the T. gondii Izatnagar isolate. Amino acid sequence of the predicted GRA-4 protein from local isolate was different at positions 19 and 304.

Conclusion: This research cloned rGRA-4 in pET SUMO plasmid.

Keywords: cloning, expression, GRA-4, pET-SUMO, plasmid, recombinant.