Effect of human β-Globin second intron on transient gene expression  in mammalian cell lines

Kevin Kumar Vijayakumar; Humera Khathun Abdul Hameed; Shakila Harshavardhan

doi:10.18006/2023.11(4).663.670

Authors

Kevin Kumar Vijayakumar Department of Molecular Microbiology, School of Biotechnology, Madurai Kamaraj University, Palkalai Nagar, Madurai, Tamil Nadu
Humera Khathun Abdul Hameed Department of Molecular Microbiology, School of Biotechnology, Madurai Kamaraj University, Palkalai Nagar, Madurai, Tamil Nadu
Shakila Harshavardhan Department of Molecular Microbiology, School of Biotechnology, Madurai Kamaraj University, Palkalai Nagar, Madurai, Tamil Nadu

DOI:

https://doi.org/10.18006/2023.11(4).663.670

Keywords:

Human β-Globin (hBG) second intron, Gene therapy, Intron-Mediated Enhancement (IME), Nonsense-Mediated Decay (NMD), pVAX

Abstract

Exogenous protein expression in mammalian cells is necessary to produce therapeutic proteins and modern medical applications like developing DNA vaccines and gene therapy. This study examines the human-Globin (hBG) second intron's capacity for intron-mediated enhancement (IME) in various mammalian cell lines. Our study's main aim is to investigate the effect of the incorporation and arrangement of the second intron of the human Beta-globin gene into the pVAX-1 expression cassette on improving the expression of foreign genes. Two plasmids were constructed, one with the hBG second intron positioned upstream and the other downstream in the expression cassette. EGFP expression was evaluated at the mRNA and protein levels after transfection using Lipofectamine 2000 using One-way ANOVA analysis. Results showed that the pVAX-1 harbouring the hBG second intron did not lead to enhanced transient EGFP expression and did not exhibit Intron Mediated Enhancement (IME) in tested mammalian cell lines. Further investigations are necessary to understand factors contributing to the lack of enhancement and explore alternative intron options for optimizing foreign gene expression in cell lines.

Author Biographies

Kevin Kumar Vijayakumar, Department of Molecular Microbiology, School of Biotechnology, Madurai Kamaraj University, Palkalai Nagar, Madurai, Tamil Nadu

Humera Khathun Abdul Hameed, Department of Molecular Microbiology, School of Biotechnology, Madurai Kamaraj University, Palkalai Nagar, Madurai, Tamil Nadu

Shakila Harshavardhan, Department of Molecular Microbiology, School of Biotechnology, Madurai Kamaraj University, Palkalai Nagar, Madurai, Tamil Nadu

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