Cultivation and expansion of erythroblasts from peripheral blood mononuclear cells to assess the efficiency of nucleoporation with plasmid containing the GFP gene

• Grant number: 12/06804-1
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: July 01, 2012
• End date: December 31, 2012
• Field of knowledge: Biological Sciences - Genetics - Human and Medical Genetics
• Principal Investigator: Lygia da Veiga Pereira
• Grantee: Lucas Assis Pereira
• Host Institution: Instituto de Biociências (IB). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

Recently, researchers using murine and human fibroblasts developed a technique for reprogramming somatic cells into induced pluripotent stem cells (iPSC), where four factors were used for cell reprogramming. Since then, several studies are reproducing this technique effectively in different cell types. Reprogramming can be done in several ways, using micro RNAs (miRNA), episomal vectors (which may easily be lost after successive mitosis), the introduction of a small circular DNA (mini-circle) with transcription factors in the cell, cell culture in culture medium with a given set of specific proteins, among others. However, the fibroblasts obtained from patients for generating iPSC use a skin biopsy. This is an invasive method and slows cell culture growth compared to the obtaining of blood cells. Due to this fact, many studies have focused on reprogramming blood cells that are easily obtained and have faster growth than fibroblasts. Thus, blood cells have become a promising source for cell reprogramming in iPSC for developing models of human disease. In this study, we will perform the cultivation and expansion of erythroblasts from peripheral blood cells (mononuclear - MNC) and nucleoporation using a plasmid containing the GFP gene and subsequently analyze the efficiency of this technique.(AU)