Generation of miR-132/212 knockout mice.

<p>miR-132/212 knockout mice were generated by insertion of LoxP sites in the 1^st intron and exon2 of the small non coding RNA gene that contains miR-132 and miR-212 (A). Targeted C57BL/6 ES cells were generated as described in the Methods section, and identified by Southern analysis of Kpn I digests using a probe 3′ to the sequence used in the targeting vector (B). Correctly targeted ES cells were used to generate a conditional miR-132/212 allele in mice using standard techniques. Mice were crossed to Flp transgenic mice to excise Neomycin resistance cassette and then Cre expressing mice to delete miR-132 and miR-212. Routine genotyping was carried out using 3 primers (p1, p2 and p3) which resulted in bands of 373 bp for the wild-type allele, 420 for the floxed allele and 550 for the deleted allele (C). Litter sizes (D) and pre weaning mortality (E) for matings of male and female knockout (n = 50 for litter size, n = 39 for pre weaning mortality), floxed (n = 20 or 17) or heterozygous (n = 53 or 41) mice are shown. Litters where pups were used for neuronal cultures were excluded from the pre weaning mortality values. Error bars represent standard deviation.