title: GSM5914891: D_1; Homo sapiens; RNA-Seq
library_construction_protocol: A JoVE visual experiment video and protocol are available by the lead author that describes in detail how the purkinje cells were removed from post-mortem human brain. PMID: 30735198 Total RNA was extracted from approximately ~2000 Purkinje cells per patient using a Qiagen Rneasy Micro kit (Cat# 74004) with repeated loading of the spin column in step 3. Samples were eluted in 14uL of water with RNase inhibitors. RNA quality was checked using an Agilent bioanalyzer pico chip. Library was constructed following manufacturers protocols of the SMARTer Stranded RNA-seq kit v2 Pico input (Cat# 634413), including indexing primers for 96 samples. 5ng inputs of total RNA were fragmented for 4 mins. Next, first strand synthesis and addition of illumina adapters, followed by AMpure bead cleanup. Samples underwent rRNA depletion with ZapR and R-probes. Library was amplified with 13 cycles and cleaned a second time with AMpure beads. Library cDNA was eluted in 15uL of Tris buffer.