GSM6965119: tg_D; Mus musculus; RNA-Seq
Tissue collection: Two days after the last injection of 12A12mAb, animals from three experi-mental groups (wild-type, naive Tg-AD, Tg-AD+mAb) were sacrificed by cervical dis-location, perfused transcardially with ice-cold phosphate buffered saline (PBS), brains and eyes were collected, hippocampi and retinas were dissected, immediately frozen on dry-ice and, then, stored at -80°C until use.   Crude synaptosomal preparations were obtained from mice of three experimental groups (wild-type, naive Tg-AD, Tg-AD+mAb). High-quality RNA was extracted from hippocampi and retinas of wild-type, naive Tg-AD, Tg-AD+mAb mice (n≥4 biological replicates per experimental group) using Trizol reagent (Sigma-Aldrich, United States) according to the manufacturer's protocol. The quantity and quality of the extracted RNA were checked by using the Agilent 4200 Tape Station using the Agilent High Sensitivity RNA Screen Tape (Agilent Technologies, Santa Clara, CA, USA). Libraries were prepared using Ion AmpliSeq Transcriptome Mouse Gene Expression Panel, Chef-Ready Kit on Ion Chef (Thermo Fisher Scientific, Waltham, MA, USA) ac-cording to the manufacturer's instructions. Briefly, 10 ng of total RNA was reverse transcribed using SuperScript VILO cDNA Synthesis Kit (Thermo Fisher Scientific, Waltham, MA, USA) before library preparation on the Ion Chef™ instrument (Thermo Fisher Scientific, Waltham, MA, USA). The resulting cDNA was amplified to prepare barcoded libraries using the Ion Code PCR Plate and the Ion AmpliSeq Transcriptome Mouse Gene Expression Core Panel and Chef-Ready Kit according to the manufacturer's instructions. Barcoded libraries were quantified using the Ion Library TaqManTM Quantitation Kit (Thermo Fisher Scientific, Waltham, MA, USA) and were combined to a final concentration of 100 pM and used to prepare template-positive Ion Sphere Particles to load on Ion 540 Chips using the Ion 540 Kit-Chef. Sequencing was performed on an Ion S5 sequencer with Torrent Suite Software v6 (Thermo Fisher Scientific, Waltham, MA, USA).