动物造模,miR-223全基因敲除小鼠 z-bio 2021-09-23 548

　　Goal: Do you want to use dual sgRNA to construct miR-223 knockout mice?

　　Method: Design a dual sgRNA of miR-223 gene, and inject in vitro transcribed sgRNA and Cas9 mRNA into fertilized C57BL/6 mouse eggs? The genomic DNA was amplified and sequenced by PCR to identify the genotype, and the mouse liver was collected at the same time, and the total RNA was extracted after crushing. The expression of miR-223 in the liver was analyzed by real-time PCR.

　　Result: Designed miR-223 gene double sgRNA in vitro transcription purification, microinjection of mouse fertilized eggs to obtain miR-223 gene mutant mice? The sequencing results showed that the mutant mice had three genotypes. The 6 bp deletion mutation does not affect the miR-223 sequence; the other two are 162 bp and 168 bp deletion mutations, which completely delete the miR-223 precursor and mature region sequence? Compared with wild type, these two mouse liver tissues are almost impossible. Is the expression of miR-223 detected?

　　Conclusion: Do you want to design dual sgRNA and apply CRISPR/Cas9 technology to successfully construct miR-223 knockout mice?