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. 2018 May 15;209(3):773–787. doi: 10.1534/genetics.118.301004

Figure 1.

Figure 1

A kdm5140 null allele demonstrates that KDM5 is essential in Drosophila. (A) Lethality of kdm5K06801, kdm510424, and kdm5140 homozygous mutant animals generated from a cross between five female and five male heterozygous parents balanced using CyO-GFP. The column labeled total flies indicates the number of progeny (adult) flies scored from at least three independent crosses. Expected number of progeny is based on Mendelian frequencies and taking into account the lethality of CyO homozygotes, i.e., 33% of total adult flies. * P < 0.01 (chi-squared test). (B) Position of the NP4707, 10424, and K06801 P-element insertions and molecular mapping of the kdm5140 deletion. Ab indicates the region used to generate the rabbit polyclonal anti-KDM5 antibody (Secombe et al. 2007). (C) Real-time PCR using primers to the 5′ end of the gene using RNA from whole third-instar larvae. Animals homozygous for kdm5K06801 or kdm510424 show low levels of transcript while kdm5140 shows none. kdm5 mRNA normalized to wild type (w1118), using rp49. **** P < 0.0001. (D) Real-time PCR using primers to the 3′ end of the gene using RNA from whole third-instar larvae. kdm5140 has wild-type levels of the 3′ end of the transcript. **** P < 0.0001. (E) Western blot from wild-type (w1118) and kdm5140 homozygous mutant wing imaginal discs showing KDM5 and α-tubulin. kdm5140 animals have no detectable full-length or truncated KDM5 proteins. *ns indicates nonspecific band. (F) Schematic of strain genotype for rescue of kdm5140 with a genomic rescue transgene. Flies are homozygous for the kdm5140 mutation on the second chromosome and homozygous for an 11 kb genomic rescue transgene on the third chromosome. (G) Western blot showing KDM5 protein levels from third-instar larval wing imaginal discs from wild-type (w1118) and kdm5140 homozygotes that also have two copies of the kdm5:HA genomic rescue transgene. Anti-KDM5 (top), anti-HA (middle), and anti-histone H3 loading control (bottom). (H) kdm5140 lethality is rescued by a transgene encoding the kdm5 locus. These data were generated by crossing female and male flies heterozygous for kdm5140 and homozygous the wild-type genomic rescue transgene (intercross of kdm5140/CyO-GFP ; kdm5:HA/kdm5:HA males and females).