كلون سازي cDNA گيرنده فعال كننده پراكسي زومي نوع γ1 موشي در وكتور بياني pEGFP-C1

Cloning of mouse PPARyl cDNA in pEGFP-CI expression vector

Peroxisome proliferated activated receptor y type 1 contains various functions In the cells including regulation of cell growth and development, regulation of immune responses and energy homeostasis and regulation of stem cell differentiations. The aim of this study was to clone PPARyl cDNA in a mammalian expression vector in a chimeric cDNA type, encompassing PPARyl cDNA with EGFP eDNA and in later study, transfering into the stem cells to see the role of PPARyl in the process of stem cell differentiations. At the first step, total RNA was extracted from fat tissue of an adult mouse. Using specific primers, PPARy]· cDNA was amplified to produce the entire length of ORF. RT-PCR products containing PPARyl cDNA were treated by enzymatic digestion and inserted into the pEGFP-CI downstream the EGFP cDNA and were used for transformation into bacterial competent cells. The positive colonies which showed inserted PPARy1 cDNA were selected for plasmid preparations and additional analysis was performed to ensure that PPARy1 cDNA was inserted properly. The results from enzymatic digestion and sequencing, confirmed, as expected, that PPARyl cDNA was amplified and cloned correctly. This cDNA gene encompasses 1428 bp.