Fig. 3. RT-PCR and western blot analysis of the prt^gs mutation. (A) Amplification of the 1747 bp prt cDNA fragment with prt 1F and prt 4R (Fig. 4) primers. rp49 primers were used as a template amount control in the same reaction. (Lane a) molecular weight marker (1 kb ladder, Fermentas), (lane b) wild type female, (lane c) prt^gs female, (lane d) wild-type embryo, (lane e) prt^gs maternal null embryo originated from a cross of a prt^gs homozygous female and a wild-type male, (lane f) maternal and zygotic prt null embryos, the progeny of prt^gs homozygous females and males. (B) Demonstration of an aberrant splice variant in the prt^gs mutant females by RT-PCR. Prt 1F and white 3R primers were used in the RT-PCR reaction. (Lane a) molecular weight marker. An 882 bp long RT-PCR product from the aberrant splice species was amplified in the mutant prt^gs (lane c) but not in the wild type (lane b). (C) Western blot of wild-type (lane a) and prt^gs/prt^gs (lane b) ovary extracts. The blot was probed with an antibody against Drosophila PRT. A doublet of 53-55 kDa is recognised in ovary extracts from wild-type flies, but not from homozygotes for prt^gs/prt^gs. (D) PRT-GFP fusion protein distribution in different stages (st) of developing eggs in wild type.