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Fig. 7. The pds5^e6 mutant produces a transcript lacking exons 1 through 5. (A) Northern blot of wild-type (Oregon R) total RNA from different developmental stages: EE, early embryo (0 to 30 minutes post egg-laying); L1, first instar larvae; L2, second instar larvae; LL3, late third instar larvae; P1, 0- to 1-day-old pupae. Each lane contained 5 µg of total RNA and the blot was probed sequentially for pds5 (4.6 kb) and rp49 transcripts. The bar graph shows phosphorimager quantification of the northern, with pds5 transcript levels normalized to rp49 levels, and the amount of pds5 transcript present in second instar larvae was set to 1 unit. (B) Northern blot of total RNA from the indicated genotypes: WT, y w; P/P, homozygotes for the viable insertion (P{EPgy2}CG17509^EY06473) used to generate the pds5 mutations; e3/+, pds5^e3/+; e6/+, pds5^e6/+; e3/e3, pds5^e3 homozygotes; e6/e6, pds5^e6 homozygotes. Each lane contained 5 µg of total RNA and the blot was probed for pds5 exon 9. The bar graph to the right shows phosphorimager quantification with pds5 transcript levels normalized to rp49 levels and the amount of 4.6 kb transcript present in wild type (WT) set to 1 unit. Black bars indicate the levels of the 4.6 kb transcript, and gray bars indicate the level of 3.65 kb transcript in heterozygous and homozygous pds5^e6 mutants. (C) Diagram of the pds5 gene and the pds5^e6 mutant 3.65 kb transcript determined by 5' RACE. The 3.65 kb transcript begins 67 nucleotides upstream of the pds5 start site predicted by EST sequences. The 3.65 kb transcript includes the first few nucleotides of exon 1 and 28 nucleotides of P element sequence fused to sequence near the start of pds5 exon 6. The 5' RACE sequence is shown in Fig. S1 in the supplementary material.

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