RNA-Seq of human glioblastoma from 64 year old male: Sample Gbm5_rnaseq
RNA from all tumor samples was extracted with Trizol (Invitrogen). rRNA depletion was performed with the Ribo-Zero kit (Epicentre) according to manufacturer instructions then concentrated using either the RNEasy MinElute Kit (Qiagen) or the RNA Clean and Concentrate-5 kit (Zymo). RNA was then fragmented with RNA Fragmentation Reagents (Ambion) for 2 to 4 minutes at 94 degrees and cleaned up with a 1.8x ratio of AmpureXP using a typical SPRI protocol. Fragmented RNA was treated with PNK (NEB) according to manufacturer instructions then cleaned up up using either the RNEasy MinElute Kit (Qiagen) or the RNA Clean and Concentrate-5 kit (Zymo). Libraries were prepared using the NEBNext Small RNA Library Preparation Kit with the following modifications: adapters and RT primer were diluted 1/2 with water, PCR primers were diluted 1/4 with water, and PCR products were purified with 1x AmpureXP. If substantial adapter-dimer product remained in the final library an additional 1x AmpureXP purification was performed.
UTEXAS-phs001389
Gbm5_rnaseq