ABSTRACT
OBJECTIVE: To identify MARCO gene knockout mouse model by genotyping,sequencing and Western blotting.METHODS: A total of 16 base-knockout MARCO~(-/-)C57 BL/6 mice( 8 female and 8 male) were obtained by clustered regularly interspaced short palindromic repeats( CRISPR)/CRISPR associated protein 9( Cas9) technique with MARCO~(+/+)C57 BL/6 mice(8 female),and their offspring MARCO~(+/-)mice were obtained. Then MARCO~(+/-)mice were inter-crossed to get a sufficient number of MARCO~(-/-)homozygous mice. The genotypes of mice were identified by gene sequencing and the relative expression of MARCO protein was detected by Western blotting. RESULTS: After one year of breeding,a total of 5 generations were bred. There were 5 types of MARCO~(-/-)genotypes(-11,-25,-36,-46,-61 bp) stably inherited; MARCO~(-/-)∶ MARCO~(+/-)∶ MARCO~(+/+)= 1 ∶ 2 ∶ 1,which was consistent with Mendelian's law of heredity. Using MARCO(-11 bp) as an example,42 MARCO~(-/-)mice,92 MARCO~(+/-)mice and 48 MARCO~(+/+)mice were obtained from the 5 th generation( F5 generation); and there was no significant difference in body mass of the above 3 genotypes of F5 generation mice at the 4 th,the 6 th and the 8 th weeks after birth( P > 0. 05). The relative expression of MARCO protein in MARCO~(-/-)(-11 bp) and MARCO~(-/-)(-46 bp) mice was significantly down-regulated,compared with that of MARCO~(+/+),MARCO~(-/-)(-36 bp) and MARCO~(-/-)(-61 bp) mice(P < 0. 05). MARCO~(-/-)(-11 bp)and MARCO~(-/-)(-46 bp) mice were chosen as the MARCO gene knockout mice. CONCLUSION: MARCO gene knockout mice were successfully identified,which laid a foundation for further study on the role and regulatory mechanism of MARCO gene in silicotic fibrosis in mice.