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Fig. 1. Identification of Hox downstream genes during early embryogenesis. (A) Outline of microarray analysis. Scanning electron micrographs of embryos at late stage 10-early stage 11, late stage 11-early stage 12, and late stage 12-early stage 13. arm::2xEGFP embryos demonstrate ubiquitous transgene expression beginning at stage 10. Time scale shows hours of embryogenesis. (B) Quantitative real-time PCR shows similar levels of overexpression of Dfd, Scr, Antp, Ubx, Abd-A and Abd-B transgenes. Numbers of genes regulated by the different Hox proteins are indicated. (C) Average fluorescent intensity (in arbitrary units) of 20 independent nuclei at different locations in wild-type, arm::Dfd and arm::Abd-B embryos stained either with -Dfd or -Abd-B antibody. Due to variable expression levels of endogenous Hox proteins, fluorescence from nuclei in different expression domains was measured (marked as `endo weak' and `endo strong'). (D) Representative embryo used for measuring fluorescent intensity of nuclei is shown. The upper two rows show wild-type and arm::Dfd embryos stained with -Dfd antibody, the lower two rows wild-type and arm::Abd-B embryos stained with -Abd-B antibody. Red boxes mark the areas used for fluorescence analysis. s, strong endogenous expression domain; w, weak endogenous expression domain. (E) Ubiquitous overexpression of Hox proteins in stage 11 and 12 was confirmed by antibody staining on arm::lacZ and arm::Hox embryos.

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