Gene Editing by CRISPR-Cas9 System

Background and Aim: CRISPR-Cas9 is an acquired immune system inprokaryotes that help them for resistance against foreign genetic elementsuch as phages. Today researchers are using CRISPR-Cas9 for in vivoand in vitro gene editing. Duchenne muscular dystrophy (DMD) is agenetic disorder characterized by progressive muscle degeneration andweakness. It is one of nine types of muscular dystrophy. DMD is causedby an absence of dystrophin. The aim of our study was in vitro dmd geneeditingMethods: HEK-293 cell was cultured in DMED high glucose supplementedwith 10% FBS (Fetal bovine serum), 500 μL Pen/Strep. gRNAs weredesigned to target Exons 48 to 53. This gRNAs were inserted in a vectorthat carrying Cas9 and GFP. After cloning, this vector transected to thecells by lipofectamine 2000. According to the gRNAs that we designed,we expected a500 bp band after deletion in dmd gene.Results: Cells that were transfected with the vector were isolated. AfterDNA extraction and PCR, we demonstrated 500 bp band in edited cells.To confirmation of edition, Sanger sequencing was done.Conclusion: In conclusion, CRISPR-Cas9 can be used as a powerful toolfor gene editing and helpful for a genetic disorder and cancer treatment.

کلیدواژه ها:

Gene editing ، dmd ، CRISPR/Cas9