Fig. 2.

Mice doubly deficient for miRNA374 and miRNA421 showed no particular phenotype. a Map of the WT, Ftx-deficient (Ftx KO) allele, and miRNA374/421-doubly deficient (miRNA374/421-DKO) allele. The expression of both lncRNA Ftx and two microRNAs—miRNA374 and miRNA421—were lost in the Ftx-targeted allele. b Scheme for generating the miRNA374/421-DKO allele. The neomycin-resistance gene (Neo) was used as a positive selection marker. Yellow and red triangles indicate FRT and loxP sites, respectively. c Homologous recombination was confirmed by PCR. The PCR primers (001, 024, 041 and 042) used for genotyping are depicted in b. R, recombinant allele. d Homologous recombination was confirmed by southern blotting. The probe position is denoted as a thick black rectangle in b. Genomic DNA was digested with SacI (left) and ScaI (right). e Cre-loxP-mediated recombination was confirmed by PCR. The PCR primers (061 and 057) used for genotyping are depicted in b. R, recombinant allele. Marker lane: size marker DNA, ΦX174/Hae III digest. f The expression levels of miRNA374 and miRNA421 were quantified by RT–qPCR in adult mouse kidneys (n = 3) of WT females (+/+), heterozygous females (+/–), homozygous females (–/–), WT males (+/Y) and hemizygous males (–/Y). Error bars, s.d. g The frequency of gross eye abnormalities in Ftx-deficient and miRNA374/421-DKO mice at weaning. The abnormalities were only observed in Ftx-deficient (–/+, +/– and –/–) mice but not in miRNA374/421-DKO (–/+, +/– and –/–) mice (χ²-tests; *P < .05; **P < .01)