Cloning and transient over-expression of the NRF2 gene in mesenchymal stem cells using adenoviral expression system based on the Gateway technology

Mohammad Mohammadzadeh; Mehryar Habibi Roudkenar

Journal of Kermanshah University of Medical Sciences

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Cloning and transient over-expression of the NRF2 gene in mesenchymal stem cells using adenoviral expression system based on the Gateway technology

Author(s):

Mohammad Mohammadzadeh¹,

Mehryar Habibi Roudkenar^2,*

1Dept. of Basic Sciences, School of Allied Medical Sciences, Gonabad University of Medical Sciences, Gonabad, Iran

2Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran

Journal of Kermanshah University of Medical Sciences:Vol. 17, issue 5; e77056

Published online:Aug 29, 2013

Article type:Research Article

Received:Nov 19, 2012

Accepted:Apr 30, 2013

How to Cite:Mohammad MohammadzadehMehryar Habibi RoudkenarCloning and transient over-expression of the NRF2 gene in mesenchymal stem cells using adenoviral expression system based on the Gateway technology.J Kermanshah Univ Med Sci.17(5):e77056.

Abstract

Background: Increased mesenchymal stem cells (MSCs) survival after transplantation can improve their therapeutic potential. Therefore, manipulation of the MSCs with cytoprotective genes such as NF-E2 Related Factor-2 (NRF2) is important to improve cell therapy efficiency. The aim of this study was cloning and transient over-expression of the NRF2 gene in MSCs by adenoviral expression system.

Methods: NRF2 was isolated from MSCs and cloned into pENTR/TOPO/D vector by TOPO cloning reaction. NRF2 was translocated from pENTR/TOPO/D vector to adenoviral vector using the Gateway technology. The recombinant adenovirus vector was transformed into 293A cells to package recombinant adenovirus particles. Recombinant adenoviruses harboring NRF2 infected MSCs.

Results: NRF2 was successfully isolated, cloned and its accuracy was confirmed by Deoxyribonucleic acid (DNA) sequencing. NRF2 over-expression was evaluated. This over-expression by adenoviral expression system was transient.

Conclusion: NRF2 over-expression in MSCs with adenoviral expression system could be a valuable device because of its transient expression. Stable over-expression of genes in MSCs could not be permitted.

Keywords

Mesenchymal Stem Cell

NRF2

genetic engineering

genetic vectors and cloning vector

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