SRX3380757 GSM2849528 SRP124720 SRS2676329 GSM2849528 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849528 GSM2849528 GEO Accession GSM2849528 SRX3380758 GSM2849529 SRP124720 SRS2676330 GSM2849529 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849529 GSM2849529 GEO Accession GSM2849529 SRX3380759 GSM2849530 SRP124720 SRS2676332 GSM2849530 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849530 GSM2849530 GEO Accession GSM2849530 SRX3380760 GSM2849531 SRP124720 SRS2676331 GSM2849531 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849531 GSM2849531 GEO Accession GSM2849531 SRX3380761 GSM2849532 SRP124720 SRS2676333 GSM2849532 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849532 GSM2849532 GEO Accession GSM2849532 SRX3380762 GSM2849533 SRP124720 SRS2676334 GSM2849533 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849533 GSM2849533 GEO Accession GSM2849533 SRX3380763 GSM2849534 SRP124720 SRS2676335 GSM2849534 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849534 GSM2849534 GEO Accession GSM2849534 SRX3380764 GSM2849535 SRP124720 SRS2676336 GSM2849535 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849535 GSM2849535 GEO Accession GSM2849535 SRX3380765 GSM2849536 SRP124720 SRS2676337 GSM2849536 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849536 GSM2849536 GEO Accession GSM2849536 SRX3380766 GSM2849537 SRP124720 SRS2676338 GSM2849537 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849537 GSM2849537 GEO Accession GSM2849537 SRX3380767 GSM2849538 SRP124720 SRS2676339 GSM2849538 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849538 GSM2849538 GEO Accession GSM2849538 SRX3380768 GSM2849539 SRP124720 SRS2676340 GSM2849539 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849539 GSM2849539 GEO Accession GSM2849539 SRX3380769 GSM2849540 SRP124720 SRS2676341 GSM2849540 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849540 GSM2849540 GEO Accession GSM2849540 SRX3380770 GSM2849541 SRP124720 SRS2676342 GSM2849541 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849541 GSM2849541 GEO Accession GSM2849541 SRX3380771 GSM2849542 SRP124720 SRS2676343 GSM2849542 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849542 GSM2849542 GEO Accession GSM2849542 SRX3380772 GSM2849543 SRP124720 SRS2676344 GSM2849543 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849543 GSM2849543 GEO Accession GSM2849543 SRX3380773 GSM2849544 SRP124720 SRS2676345 GSM2849544 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849544 GSM2849544 GEO Accession GSM2849544 SRX3380774 GSM2849545 SRP124720 SRS2676346 GSM2849545 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849545 GSM2849545 GEO Accession GSM2849545 SRX3380775 GSM2849546 SRP124720 SRS2676348 GSM2849546 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849546 GSM2849546 GEO Accession GSM2849546 SRX3380776 GSM2849547 SRP124720 SRS2676347 GSM2849547 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849547 GSM2849547 GEO Accession GSM2849547 SRX3380777 GSM2849548 SRP124720 SRS2676350 GSM2849548 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849548 GSM2849548 GEO Accession GSM2849548 SRX3380778 GSM2849549 SRP124720 SRS2676349 GSM2849549 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849549 GSM2849549 GEO Accession GSM2849549 SRX3380779 GSM2849550 SRP124720 SRS2676351 GSM2849550 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849550 GSM2849550 GEO Accession GSM2849550 SRX3380780 GSM2849551 SRP124720 SRS2676352 GSM2849551 RNA-Seq TRANSCRIPTOMIC cDNA Prior to RNA-seq, the AcfC flies were backrossed with the wild-type OrR strain for 8 generations. Embryos were collected 0-45 min AEL and allowed to develop at 25°C until approximately 30 min before the desired stage (around 1 h for Bownes Stage 3 and 4 h for Bownes Stage 8). Without prior dechorionation, embryos were hand-picked and submerged into a drop of Voltalef 10s halocarbon oil (Lehman and Voss Co.) placed on a microscope slide. After about 5 min, the embryonic structures become visible under the stereomicroscope. Embryos were allowed to develop further under the halocarbon oil until the desired stage. Single embryos were picked and crushed with a 26G needle into 200 µl of Lysis Buffer (supplemented with Proteinase K) from the Agencourt RNAdvance Tissue Kit (Beckman Coulter, Cat No A32645). After the addition of 10 µl of 1:100 ERCC Spike-in RNA mix (Ambion, Cat No 4456740), the samples were incubated at 37°C for 20 min, snap-frozen in liquid nitrogen and stored at -80°C. Total RNA was extracted from the single-embryo homogenate using the same Agencourt RNAdvance Tissue Kit, following standard protocol but utilizing half of the volumes recommended. The RNA integrity was checked on a Bioanalyzer 2100 (Agilent). Ribosomal RNA depletion was achieved using rRNA Depletion Kit (Human/Mouse/Rat) (New Englands Biolab, Cat No E6310) and the rRNA-depleted RNA was stored at -80°C until further processing. Non-directional libraries were prepared using NEBnext Ultra RNA Library Prep Kit for Illumina (New Englands Biolab, Cat No E7530S) following standard protocol. 6 replicates per genotype and stage were sequenced on an Illumina HiSeq1500 instrument. Illumina HiSeq 1500 gds 302849551 GSM2849551 GEO Accession GSM2849551