title: GSM9282554: Mm_Wnt1-Cre-Nedd4fl-fl-Embryo_Mutant_rep2; Mus musculus; RNA-Seq
library_construction_protocol: RNA was prepared from tissue by lysing in 4M guanidine isothiocyanate solution (Invitrogen) and precipitating with sodium acetate and ethanol. After 75% ethanol wash, RNA pellet was resuspended in 12ml H2O, with 1ml reserved for Bioanalyser quality control of RNA, and 9.5ml used for subsequent cDNA synthesis. Isolated RNA samples were submitted to the ACRF Genomics Facility for library preparation and sequencing. Extracted RNA was quantified using the Bioanalyser Pico chip and 9.5ul (containing 5-17ng of RNA) was input into SMART-Seq v4 Ultra Low Input RNA Kit for Sequencing (Takara Bio) for generation and amplification of double-stranded full-length cDNA (using 9 cycles of LD-PCR). Prior to library preparation, cDNA was sheared to 200-500 bp fragments using the Covaris system. NGS libraries were prepared using the ThruPLEX DNA-seq Kit (Takara Bio) together with the DNA Unique Dual Index Kit (Takara Bio), using 11 cycles of PCR for library amplification.