SgRNA and use thereof in repairing abnormal splicing of introns

Abstract

The invention discloses a sgRNA and use thereof in repairing abnormal splicing of introns. The sgRNA is used for repairing abnormal splicing of introns caused by a HBB (a beta-globin gene) IVS2-654 C>T mutation and comprises a sgRNA-U and a sgRNA-D. A targeting site of the sgRNA-U is located in a range from an upstream 21 bp of the IVS2-654 C>T site to a starting site of a second intron of the HBBgene where the IVS2-654 C>T site is located. A targeting site of the sgRNA-D is located in a range from a downstream 71bp of the IVS2-654 C>T site to a termination site of the second intron of the HBB gene where the IVS2-654 C>T site is located. An existing gene editing technology can be used for repairing a blood transfusion-dependent beta-thalassemia IVS2-654 C>T, has high editing efficiency, and can efficiently modify a persistent and balanced hematopoietic system for autologous hematopoietic stem cells.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to a sgRNA and its application in repairing abnormal splicing of introns. Background technique [0002] β-thalassemia is a common genetic disease caused by abnormal hemoglobin in adults due to β-globin gene defects. There are about 30 million carriers of the "thalassemia" gene in my country, involving nearly 30 million families and 100 million people. Among them, severe and about 300,000 patients with intermediate "thalassemia". Among them, the IVS2-654 C>T genotype is a relatively common type of "thalassemia" in my country. The pathogenesis is that the 654th base in the second intron of the HBB gene has a C>T mutation, resulting in abnormalities. splice site, resulting in an extra 73nt exon in β-globin mRNA and premature termination of translation. [0003] At present, patients with intermediate and severe thalassemia need long-term blood transfusion and iron remo...

AI Technical Summary

Problems solved by technology

Among them, the use of lentiviral vectors for gene therapy has shown great potential, but the semi-random vector integration method has cancer risk

At the same time, the expression elements in the lentivirus will be gradually silenced during the long-term homing and self-renewal process of hematopoietic stem cells, which will reduce the curative effect, that is, it may not be possible to achieve the purpose of lifelong cure

In addition, the high-concentration and high-quality lentivirus required in clinical practice has extremely high requirements on equipment and technology, so it is difficult to reduce the cost

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