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Fig. 1. Generation of a dark⁸² null mutation. (A) Schematized view of the genomic structure of the dark locus, relevant alleles and the dark⁸² null mutation. The dark transcript spans 6.6 kb. dark⁸² is a 6324 bp deletion (dashed line) generated by imprecise excision of the indicated P-element in the dark^CD4 strain (Rodriguez et al., 1999). The allele was mapped by sequencing a 1.3 kb genomic PCR fragment (see B) using a primer pair (designated 1 and 2) spanning the junctional interval. In dark⁸², sequences from -1277 bp (upstream of the translation start codon) to 19 bp downstream of the stop codon are absent such that the entire dark ORF and part of the untranslated first exon are missing. Note that 396 bp of sequence from the CD4 transposon remain at this junction. (C) RT-PCR with primer pair 3 and 4, using total RNA from prepupae, confirms complete loss of the dark transcript in the dark⁸² allele. Two different isolates of dark⁸² from the screen were assayed here, 82 (1) and 82 (2). rp49 is a control.

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