Design and construction of a recombinant gene construct expressing the cell protective gene

Genetic manipulation is an effective strategy to protect cells against environmental damages and enhance their capabilities for therapeutic usage. In order to avoid unwanted side effects, such as cancers, the expression of genes should be temporary increased. The aim of this study was to clone and temporary increased expression of a cell protective gene, Metallothionein 1 (MT1) in mesenchymal stem cells (MSCs) using plasmid expression system with the pcDNA 3.1 vector. Human bone marrow mesenchymal stem cells were isolated and validated by flow cytometry method. Complete cDNA sequence of MT1 gene was isolated and cloned in a plasmid vector named pcDNA 3.1. Recombinant gene construct was generated and transferred to the human MSCs. MT1 expression was monitored by RT-PCR and western blotting. Recombinant human MT1 gene was successfully cloned in pcDNA vector and the correct format gene in the vector was confirmed by DNA sequencing. By RT-PCR and western blot methods, increased MT1 expression in MSCs was approved. The results showed that the expression of MT1 in the MSCs was transient. Genetic manipulation of MSCs by MT1using pcDNA 3.1 plasmid vector may be one of the protective strategies for transplanted cells from apoptosis and promote a new approach to provide an effective cell therapy after transplantation.