Thalassaemia gene therapy

β-thalassemia is one of the most common genetic diseases in the world, caused by mutations in the human hemoglobin beta(HBB) gene. HBB −28 (A>G) mutations is one of the five most common mutations in China patients with β-thalassemia. However, there are few studies on this mutated of thalassemia, due to the rarity sample and ethics. Genome editing technologies for disease modeling and developing cellular therapies has been extensively documented. Therefore, firstly we combined CRISPR/Cas9 and asymmetric single strand oligo DNA to mutant HBB gene in K562 cell lines and correct the HBB mutation with high efficiency and seamless. Then in order to study how this mutation affected the hemoglobin expression, we use RPMI 1640 medium without glutamine supplemented with 10 % (v/v) FBS and 1 mM sodium butyrate to erythroid differentiation. Finally we see the impact of the mutation at the transcriptome level by RNA-Seq which is the first time to study the HBB(28A>G) differences between the isotype cell lines. The 28(A>G) affects hemoglobin expression, opens the NOTCH pathway, and affects cell morphology and size and so on. Collectively, Our study provides an effective approach to mutation and correct HBB mutations without leaving any genetic footprint in the cells, and we also have the mutation cells can provide the diease model to study the disease without ethics. At the transcriptome level ,we show the difference between the mutation and the wild type which Provide data support for subsequent studies.