Generation of Asxl1:nlacZ/nGFP knock-in mice. (A) An nlacZ/nGFP-FRTNeoFRT cassette was introduced 6 bp upstream of the Asxl1 start codon. Rectangular black bars indicate exons. S, StuI. (B) Genotype distribution among live births of Asxl1^+/− intercross. Asxl1^−/− mice are underrepresented. P values are compared with normal Mendelian distribution (25%, 50%, 25%) using a χ² test. (C) Kaplan-Meier curve representing the percent survival of Asxl1^−/− (n = 12), Asxl1^+/− (n = 42), and WT (n = 42) mice vs age in days. (D) The gross appearance of an Asxl1^−/− mouse compared with WT and Asxl1^+/− littermates (4 weeks old). (E) Body weight of WT, Asxl1^+/−, and Asxl1^−/− mice (3-6 weeks old, 9 mice/genotype; ***P < .001). (F) Western blot shows the reduced and deletion of Asxl1 expression in BM cells of representative Asxl1^+/− and Asxl1^−/− mice, respectively, compared with WT. (G) Analyses of Asxl1 mRNA expression levels in BM cells of WT (n = 4), Asxl1^+/− (n = 5), and Asxl1^−/− (n = 5) mice by qPCR with 4 different pairs of primers. The relative Asxl1 mRNA expression was determined by using β-actin as an internal calibrator and reported as relative expression units to the respective Asxl1 expression with each primer pair in WT mice. **P < .01, ***P < .001