Fig. 4. Characterization of various mid alleles and the Mid antibody. (A) Mid protein in wild-type Drosophila and in three different mid alleles. The double-headed arrow indicates the region of Mid used to raise the antibody. (B) Western analysis of the Mid protein in the wild type and in various mid alleles and a mid, H15 deficiency. The large arrow indicates the Mid-specific band in the wild type; the small arrow indicates the truncated Mid protein in los¹. (C) There is no maternal deposition of mid transcript to embryos. RT-PCR for mid transcript from unfertilized eggs (Unf) and embryos at 0-4, 4-8, 8-12 and 12-24 hours of development. rp49 was used as control. (D) There is no maternal deposition of Mid protein to embryos. Western blotting of embryos at 0-4, 4-6, 6-8 and 8-10 hours of development. The Mid band (arrow) is separated from a non-specific band in this blot. (E) Mid antibody is specific to Mid protein. Embryos are stained with Mid and Eve antibodies. Anterior is up; vertical lines mark the midline. In the wild type, Mid is expressed in a large number of neurons. The neuron that appears to change into an extra RP2 is indicated by the arrow. In mid¹ embryos, no detectable Mid was present. This was also the case in mid and H15 deficiency (mid^df) embryos.