Construction method of point mutation glanzmann's thrombasthenia (GT) mouse model

Abstract

The invention provides a construction method of a point mutation glanzmann's thrombasthenia (GT) mouse model. The construction method comprises the following steps of: (1), performing carrier design and construction, and in-vitro transcription: (1-1), designing sequences of gRNA and Donor Oligo according to gene information and experimental requirements; (1-2), synthesizing the Donor Oligo, and constructing a gRNA carrier; and (1-3), performing in-vitro transcription on the gRNA vector and a Cas9 vector; (2), performing microinjection and identification of F0-generation mice; (3), breeding andidentifying F1-generation mice: respectively breeding the sexually mature positive F0 mice in the step (2) with wild mice for one generation, and finally, providing at least three F1-generation pointmutation GT mouse models verified by PCR (Polymerase Chain Reaction) and sequencing; and (4), freezing and preserving breeds. A CRISPR/Cas9 technology is used for accurately editing and constructingthe point mutation GT mouse model for the first time; the mouse gene is accurately edited through the model; compared with a GT mouse model constructed through antibody injection induction or a gene knockout technology in the past, the model is more accurate; and a good model is provided for deeply researching the pathogenesis of the GT and exploring a new treatment method.

Description

technical field [0001] The invention belongs to the field of medical technology, and in particular relates to a method for constructing a mouse model of point mutation thrombocytopenia. Background technique [0002] Glanzmann's thrombosthenia (GT) is a monogenic genetic disease. The ITGA2B or ITGB3 gene defect of chromosome 17 is the cause of the disease. When ITGA2B or ITGB3 is mutated, the platelet membrane surface glycoprotein αIIbβ3 will be abnormal in quality or quantity, which will lead to abnormal platelet aggregation function and cause bleeding. At present, the GT mouse model constructed by antibody injection induction or gene knockout technology cannot precisely edit the mouse gene. Contents of the invention [0003] The technical problem to be solved by the present invention is to provide a method for constructing a mouse model of point mutation thrombocytopenia, which can precisely edit the gene of the mouse, and provides a good small tool for in-depth researc...

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Problems solved by technology

At present, the GT mouse model constructed by antibody injection indu

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