Deletion of Mettl3 at the Pro-B Stage Marginally Affects B Cell Development and Profibrogenic Activity of B Cells in Liver Fibrosis

Figure 1

Construction and characterization of B cell-specific Mettl3 knockout mice using Cd19-Cre. (a) Strategy of Cd19-Cre-mediated Mettl3 knockout mouse construction. Floxed-Mettl3 allele was generated by flanking exons 2 and 4 with loxP sites. B cell-specific Mettl3 knockout mice (Mettl3 cKO) were obtained by crossing floxed-Mettl3 mice with Cd19-Cre transgenic mice. (b) Schematic diagram of breeding strategies of Mettl3 cKO mice. (c) Genomic PCR for the tail of indicated genotype. The top lane (floxdel) showed the exon 2-4 deleted alleles (amplified using Mettl3-F1 and Mettl3-R2 primer shown in (a)). The middle lane (Cd19-Cre) showed the effective insertion of Cd19 promoter-driven Cre. The bottom lane displayed genotyping of heterozygous (Mettl3^flox/-) or homozygous (Mettl3^flox/flox) flox flanking alleles (amplified by Mettl3-F1 and Mettl3-R1 primer shown in (a)). (d) Genomic PCR analysis for CD19⁺ B cells and CD19^- cells isolated from the spleen of mice with indicated genotype. Intact (upper) and floxdel (lower, with the exon 2-4 deleted alleles) lanes were amplified using Mettl3-F1 and Mettl3-R2 primer shown in (a). (e) qRT-PCR for Mettl3 of CD19⁺ B cells and CD19^- cells isolated from WT and Mettl3 cKO mouse spleen. (f) Western blot for Mettl3 of CD19⁺ B cells and CD19^- cells isolated from WT and Mettl3 cKO mouse spleen. Data in (e) were presented as with the indicated significance (; student’s -test).

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