Human osteosarcoma U-2OS tool cell strain capable of stably expressing Cas9 protein as well as preparation method and application thereof

Abstract

The invention discloses a human osteosarcoma U-2OS tool cell strain capable of stably expressing Cas9 protein as well as a preparation method and an application thereof. The preparation method comprises the following steps: introducing a Cas9 protein expression gene and an antibiotic selection marker gene through a CRISPR/Cas9 system to construct the human osteosarcoma cell line U-2OS tool cell strain capable of stably expressing the Cas9 protein; the cell strain can stably express Cas9 protein, related gene editing work can be carried out as long as single sgRNA is transfected, only one smallsgRNA fragment needs to be transfected when specific gene editing is carried out, and the gene editing efficiency can be greatly improved. Osteosarcoma is the most common primary malignant osteosarcoma, U-2OS is a common osteosarcoma cell model, and the genetically engineered cell strain provided by the invention has important significance in researching pathogenesis and regulation and control networks of osteosarcoma and preventing and treating osteosarcoma as soon as possible.

Description

technical field [0001] The invention belongs to the field of genetic engineering and genetic modification, and in particular relates to a human osteosarcoma U-2OS tool cell strain capable of stably expressing Cas9 protein and a preparation method and application thereof. Background technique [0002] Osteosarcoma is the most common primary malignant bone tumor in the skeletal system originating from the mesenchymal tissue. The incidence rate is relatively high in adolescents and the elderly over 60 years old, and can reach 35% of primary bone tumors. Moreover, osteosarcoma has the characteristics of high malignancy, early metastasis and poor prognosis. Currently, the treatment for osteosarcoma mainly consists of surgery and chemotherapy. With the development of neoadjuvant chemotherapy, the overall survival rate of patients with osteosarcoma has now increased from 10% to 70%. However, about 30% of patients still experience recurrence and metastasis, leading to treatment fa...

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Problems solved by technology

[0004] However, since the Cas9 protein-coding gene is relatively large, close to 4000bp, the transfection efficiency is often low due to the large plasmid during gene editing, which seriously affects the efficiency of gene editing, making the construction of gene editing cell lines / gene editing models The success rate of animals is greatly reduced

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