GSM8890021: FoxP mutant, rep3; Drosophila melanogaster; RNA-Seq
Twelve brains of freshly eclosed males per sample were dissected in PBS at ZT1-2, transferred to RNAIater solution (Sigma), and immediately frozen in liquid nitrogen. Total RNA was extracted using the Arcturus PicoPure RNA Purification kit (Thermo Fisher Scientific). TruSeq RNA Sample Preparation Kit (Illumina) was used to prepare adapter ligated PCR fragments for sequencing. mRNA was purified using polyA selection and was reverse transcribed into a single-stranded cDNA using random hexamer priming. Second strand synthesis was performed to create double-stranded cDNA. The cDNA was then summited to DNA end repair with adenylation followed by ligation of Illumina sequencing adaptors and size selection for 300 bp. Fragments were amplified linearly with 14 PCR cycles and the sample quality of the fragments was validated using the 2100 Agilent Bioanalyzer.