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:: Scientific Journal of Kurdistan University of Medical Sciences- NO 4, 2024 -Articles In press ::
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Expression of the Gene Encoding the Brucella abortus BAB1-0278 Antigen in Probiotic Lactococcus lactis Bacteria
Donya Kazemi1 , Abbas Doosti 2, Mostafa Shakhsi niaee
1- Islamic Azad University, Shahrekord Branch,
2- Islamic Azad University, Shahrekord Branch, , abbasdoosti@yahoo.com
Abstract:   (158 Views)
Background and Aim: Brucella abortus BAB1-0278 antigen is one of the leading causes of brucellosis. This antigen encodes essential cell cycle regulator GcrA, which plays a key role in transcription of target genes across the cell cycle. Therefore, BAB1-0278 has potential to be used in therapeutic strategies against brucellosis. The objective of this study is the engineered Lactococcus lactis expressing recombinant Brucella abortus BAB1-0278 protein.
Materials and Methods: The gene construct containing a peptide signal sequence in the nisin-based expression vector (pNZ8148-Usp45-BAB1-0278) was designed and synthesized by GENEray. The recombinant plasmid was transformed in Escherichia coli (E. coli) bacteria. Recombinant plasmid was extracted from transformed bacteria. Lactococcus lactis was transformed by electroporation both with the recombinant plasmid containing the target gene pNZ8148-Usp45-BAB1-0278 and with the plasmid without the target gene. Confirmation of  BAB1-0278 expression was performed by reverse polymerase chain reaction (RT-PCR) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) techniques.
Results: Brucella abortus BAB1-0278 was expressed in Lactococcus lactis bacteria and confirmed by RT-PCR. SDS-PAGE analysis of proteins isolated from Lactococcus lactis transformed with recombinant plasmid compared to untransformed Lactococcus lactis showed a protein with a molecular weight of 13 kDa.
Conclusion: PCR, restriction enzyme digestion and sequencing results by GENEray indicated that the BAB1-0278 correctly cloned in pNZ8148-Usp45-BAB1-0278. BAB1-0278 protein expression was analyzed by SDS-PAGE. Transformed Lactococcus lactis could be an important step towards development of oral recombinant vaccine against Brucella abortus.
 
Keywords: Brucella abortus, BAB1-0278, Lactococcus lactis, Vaccine, Electroporation
     
Type of Study: Original Research | Subject: Molecular Medicine and Genetics
Received: 2023/01/5 | Accepted: 2024/02/4
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مجله علمی دانشگاه علوم پزشکی کردستان Scientific Journal of Kurdistan University of Medical Sciences
مجله علمی دانشگاه علوم پزشکی کردستان Scientific Journal of Kurdistan University of Medical Sciences
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